During the revision lecture, the unit coordinator told us not to assume that there wouldn't be a short-answer question on Lecture 1. And then I realised that I hadn't blogged about it. I thought I was done blogging about pharmacology, but apparently not!
Define the term “pharmacology” & the scope of the discipline
Pharmacology is essentially the study of how drugs and other biologically active substances affect the body. We don't worry so much about how drugs and so forth are actually prescribed, that's pharmacy's job.
Show awareness of the importance of plant-based medicines to the historical emergence of pharmacology, identifying 3 drugs that came from botanical sources
Originally we used plants as medicine due to the presence of many bioactive compounds.
One of the most well-known examples of a modern drug that originally came from a plant is aspirin. Aspirin originally came from willow bark, used as far back as ~200AD in Ancient Rome. In 1828 Büchner isolated salicin, the active component of willow bark. In 1897, aspirin, or acetylated salicylic acid, was patented and synthesised by the drug and dye firm Bayer.
A lesser-known example of a drug that came from a plant is atropine. Atropine originally came from deadly nightshade, a.k.a. belladonna. "Belladonna" actually means "beautiful woman," and the name came about because it can cause rosy cheeks and dilated pupils, which people thought was beautiful. (It does eventually cause death though, so there's that.) In 1831 Mein isolated atropine from belladonna, and in 1867 it was found to block cardiac vagal stimulation. (Atropine blocks muscarinic cholinergic receptors, as mentioned in my post about the autonomic nervous system.)
A third example is artemisinin, used to treat malaria. In 340AD, Sweet Annie (Artemisia annua) was used by Ge Hong to alleviate fever. In 1596 another Chinese herbalist, Li Shizhen, reported using it to treat malarial fever. It wasn't until the 1970s that the active ingredient (artemisinin) was isolated, however. One of the scientists working on this discovery, Tu Youyou, won a Nobel Prize for her work.
Appreciate how rising awareness of the limitations of plant-based medicines drove the growth of modern pharmacology
Plants are great, but they aren't perfect. For example, it's very hard to determine how much of an active ingredient you have in a plant. There may also be other compounds in the plant that might interfere with the activity of the active ingredient. Combined, these make it difficult to test drugs and determine a safe dose.
Another limitation of using plants is that you're limited to what you can find in nature. With technology, you can tweak molecules around to make drugs that are safer and/or more effective. So next time someone comes up to you saying "why take nasty synthetic aspirin when you can just chew on some lovely natural willow bark?" you've got a few things that you can yell right back at them.
Understand how 3 different classes of names are commonly assigned to drugs.
Drugs have at least three different names. They have a chemical name, a generic name and one or more brand names.
The chemical name of a drug is simply the name given by IUPAC. These are extremely descriptive, but are often quite a mouthful. For example, if you have depression you might be prescribed some (S)-1-[3-(Dimethylamino)propyl]-1-(4-fluorophenyl)-1,3-dihydroisobenzofuran-5-carbonitrile. Have fun asking the chemist for a refill of your (S)-1-[3-(Dimethylamino)propyl]-1-(4-fluorophenyl)-1,3-dihydroisobenzofuran-5-carbonitrile, though.
The generic name is a much simpler name given to the drug molecule that sounds somewhat sophisticated and classy, but not catchy enough to be a brand name. Normally they have prefixes or suffixes that are related to their function- for example, all of the penicillins end with -cillin. Just in case you were curious, (S)-1-[3-(Dimethylamino)propyl]-1-(4-fluorophenyl)-1,3-dihydroisobenzofuran-5-carbonitrile is also known as escitalopram. I still have no idea if I'm pronouncing it correctly (I've heard three different pronunciations of that name, two from the same person), but at least it's a lot easier to pronounce than the chemical name.
Finally, drugs have a brand name that sounds kinda catchy so that it sticks in people's minds. For example, paracetamol is often sold as Panadol, and escitalopram is often sold as Lexapro. After the drug is off its patent, other companies can sell the same drug under different brand names. For example, Lexapro is also sold under the brand name Lexam (they kept the first four letters the same, how original).
Show a basic appreciation of the importance of “pharmacokinetics” and “pharmacodymanics” to understanding the effects of drugs on the human body.
Pharmacodynamics is what a drug does to the body, and pharmacokinetics is what the body does to the drug. For more information, just read all of my other pharmacology posts :P
Showing posts with label PHAR2210. Show all posts
Showing posts with label PHAR2210. Show all posts
Wednesday, June 1, 2016
Sunday, May 29, 2016
PHAR2210 Drug Encyclopaedia
A little birdie (*cough*ConfessionsatUWA*cough*) told me that apparently last year's exam had a lot of very specific questions relating to drugs mentioned throughout the lectures, even if they had a minor mention on a random slide somewhere. This is my attempt to summarise all of the drugs that were covered in order to make revision a little easier. I will also include certain substances that are naturally produced by the body (hormone etc.), biologically-active substances that aren't used as drugs but were covered in the lectures due to toxicity or whatever, and general classes of drugs.
(Note: The unit coordinator raised concerns that I may have made some of you guys unnecessarily worried about the exam. So I guess you should just use this as a guide and not as something you need to stress out about? I'm actually the biggest stress-head, so maybe I shouldn't be giving advice on how to handle exam stress. Okay, I'm going to shut up now.)
(Note: The unit coordinator raised concerns that I may have made some of you guys unnecessarily worried about the exam. So I guess you should just use this as a guide and not as something you need to stress out about? I'm actually the biggest stress-head, so maybe I shouldn't be giving advice on how to handle exam stress. Okay, I'm going to shut up now.)
| Drug Name | Lecture(s) | Important Information |
| Acetylcholine | 12, 13 |
|
| Acrolein | 7 |
|
| Acyclovir | 19 |
|
| Adrenaline | 10 |
|
| Aminoglycaside (class) | 18 |
|
| Amphetamine | 12 |
|
| Amphotericin | 20 |
|
| Artemisinin | 1 |
|
| Aspirin | 1, 9 |
|
| Atropine | 1 |
|
| Azathioprine | 22-1 |
|
| Azidothymidine (AZT) | 19 |
|
| Azoles (class) | 20 |
|
| Benzene | 6 |
|
| Benzodiazepine (class) | 12 |
|
| Carbamazepine | 3 |
|
| Carbapenem (class) | 18 |
|
| Celecoxib | 9 |
|
| Cephalosporin (class) | 18 |
|
| Chloramphenicol | 18 |
|
| Cimetidine | 4 |
|
| Clavulanic acid | 18 |
|
| Clozapine | 3 |
|
| Cobicistat | 3 |
|
| Codeine | 3, (4) |
|
| Colistin | 17, 18 |
|
| Cortisone | 12 |
|
| Debrisoquine | 3 |
|
| Deutetrabenazine | 3 |
|
| Dexamethasone | 12 |
|
| Didanosine | 19 |
|
| Diethylstilbesterol (DES) | 6 |
|
| Diphenhydramine | 11 |
|
| Echinocandins (class) | 20 |
|
| Enalapril | 2 |
|
| Enflurane | 10 |
|
| Esomeprazole | 10 |
|
| Ethanol | 7, 8 |
|
| Fluorouracil | 12 |
|
| Fluoxetine | 1, 3, 12 |
|
| GABA (γ-aminobutyric acid) | 13 |
|
| Glucocorticoid (family) | 16 |
|
| Glutamate | 13 |
|
| Glycine | 13 |
|
| HIV Protease Inhibitors (class) | 19 |
|
| Ibuprofen | 10 |
|
| Imipramine | 5 |
|
| Insulin | 12, 15, 21 |
|
| Isoniazid | 17 |
|
| Ketamine | 10 |
|
| Lumiracoxib | 7, 9 |
|
| Lenalidomide | 8 |
|
| Macrolide (class) | 18 |
|
| Meperidine (MPPP) | 7 |
|
| Methoxychlor | 10 |
|
| Mianserin | 10 |
|
| NNRTI (class) | 19 |
|
| Nystatin | 20 |
|
| Omeprazole | 10 |
|
| Pancuronium | 12, 13 |
|
| Paracetamol/ acetaminophen | 7, 12 |
|
| Penicillin | 17, 18 |
|
| Phenylephrine | 11 |
|
| Phenytoin | 8 |
|
| Prednisolone | 12 |
|
| Probenecid | 4 |
|
| Prontosil | 17 |
|
| Salbutamol | 11, 14 |
|
| Salvarsan | 17 |
|
| Serotonin (5-hydroxytryptamine) | 13 |
|
| Sparteine | 3 |
|
| Stribild | 3 |
|
| Strychnine | 13 |
|
| Suxamethonium | 13 |
|
| Tetracycline (class) | 17, 18 |
|
| Thalidomide | 8, 10 |
|
| Theophylline | 3 |
|
| Thiopental | 2 |
|
| Tri-o-cresylphosphate (TOCP) | 6 |
|
| Valproate | 9 |
|
| Warfarin | 2, 5 |
|
Human Pharmacology III: Clinical Trials
This is just a quickie (hopefully) on all the steps that drugs have to go through before they get to market. Once this post is over, I'll be done blogging about pharmacology for the semester (and probably for the year given that I don't have any pharmacology units next semester)!
Understand the purposes of clinical drug trialling
Surely this should be pretty obvious, unless you're A-OK with heaps of people getting drugs that haven't been proven to be safe and effective...
Know the phases of clinical drug trials, the intention of each phase and what groups are recruited into them
Know the important characteristics that distinguish each phase, know the difference between observational and controlled trials and know how power and bias are managed in clinical trials.
There are four main phases, imaginatively named I, II, III and IV. Sometimes there is also a "phase 0."
Oh yeah, before I forget, I need to tell you the basic stuff about the differences between observational and controlled trials and whatnot.
Phase IV is an observational trial, where you are basically relying on reports and so forth. This might be biased because not all drug reactions may be reported, or perhaps people might report something as being a drug reaction when it was caused by something else. This is because unfortunately we are all human and prone to fallacies like "post hoc ergo propter hoc" (i.e. thinking that two things are related simply because they happened at the same time), confirmational bias (i.e. "I think vaccines are bad, therefore I'm going to be 100% vigilant for any sniffle or cough or mild change in behaviour that might be a 'vaccine injury'") and so on. However, observational trials like this are probably our best bet at observing drug reactions in an entire population.
Controlled trials, such as those done in phases II and III, attempt to avoid as much bias as possible. The gold standard is a double-blind placebo-controlled study. Double-blind means that neither the patient nor the doctor knows what drug the patient is getting- that's only revealed right at the end for the purposes of data analysis. Placebo-controlled means that the drug is either tested against a sugar pill or against a drug that's already shown to work. It might seem weird testing a drug against another drug, but sometimes that's our most ethical option. It's not nice to leave some people untreated with only sugar pills when there is an effective drug available. Speaking of ethics...
Understand the important ethical considerations surrounding human drug trialling and the function of a Human Research Ethics Committee
Ethics committees are important in making sure that researchers don't do unethical stuff, like take blood samples at kids' birthday parties *cough*Mr Andrew Wakefield*cough*. Here are the main criteria of the Human Research Ethics Committee in Australia. These criteria are probably pretty similar worldwide.
Be aware of the purposes of the Trial Protocol, the Investigator’s Brochure and the Patient Information and Consent documents
This is basically all the stuff you have to submit to get an ethics approval. The Trial Protocol is basically outlining what you're going to do, the Investigator's Brochure includes as much information about the drug as is currently known and the Patient Information and Consent Documents are the handouts that you're going to give to the patients to let them know that participation is voluntary, they can leave at any time, yada yada yada.
Aaaaaaand I'm done! (Except I am going to have a bonus post or two... stay tuned!)
Understand the purposes of clinical drug trialling
Surely this should be pretty obvious, unless you're A-OK with heaps of people getting drugs that haven't been proven to be safe and effective...
Know the phases of clinical drug trials, the intention of each phase and what groups are recruited into them
Know the important characteristics that distinguish each phase, know the difference between observational and controlled trials and know how power and bias are managed in clinical trials.
There are four main phases, imaginatively named I, II, III and IV. Sometimes there is also a "phase 0."
- Initial human pharmacokinetic and pharmacodynamic studies. Very low doses are used.
- Finding the safe dose. Only a few healthy volunteers are tested. They are exposed to gradually increasing doses of the drug while being carefully monitored.
- Finding out if the drug might work. 10s to 100s of volunteers with the condition are tested. They are compared against some kind of control group to see if the drug has any benefit over placebo (or a drug that has been shown to work).
- Finding out of the drug works. Similar to phase II, but now many more participants with the condition are tested. The number tested is statistically determined using power calculations and whatnot. (I don't really understand the statistics, sorry.)
- Continuous safety testing. Once the drug is marketed, reports of adverse effects are collected. These are generally effects that are rare so they might not have shown up in the previous three phases.
Oh yeah, before I forget, I need to tell you the basic stuff about the differences between observational and controlled trials and whatnot.
Phase IV is an observational trial, where you are basically relying on reports and so forth. This might be biased because not all drug reactions may be reported, or perhaps people might report something as being a drug reaction when it was caused by something else. This is because unfortunately we are all human and prone to fallacies like "post hoc ergo propter hoc" (i.e. thinking that two things are related simply because they happened at the same time), confirmational bias (i.e. "I think vaccines are bad, therefore I'm going to be 100% vigilant for any sniffle or cough or mild change in behaviour that might be a 'vaccine injury'") and so on. However, observational trials like this are probably our best bet at observing drug reactions in an entire population.
Controlled trials, such as those done in phases II and III, attempt to avoid as much bias as possible. The gold standard is a double-blind placebo-controlled study. Double-blind means that neither the patient nor the doctor knows what drug the patient is getting- that's only revealed right at the end for the purposes of data analysis. Placebo-controlled means that the drug is either tested against a sugar pill or against a drug that's already shown to work. It might seem weird testing a drug against another drug, but sometimes that's our most ethical option. It's not nice to leave some people untreated with only sugar pills when there is an effective drug available. Speaking of ethics...
Understand the important ethical considerations surrounding human drug trialling and the function of a Human Research Ethics Committee
Ethics committees are important in making sure that researchers don't do unethical stuff, like take blood samples at kids' birthday parties *cough*Mr Andrew Wakefield*cough*. Here are the main criteria of the Human Research Ethics Committee in Australia. These criteria are probably pretty similar worldwide.
- Research merit and integrity: Is the experiment soundly designed? You don't want to put participants through a trial and then find out that your results don't mean anything because you had a crappy experimental design.
- Justice: All patients have equal chance of benefit or harm. So for example participants should be randomly selected for the placebo and experimental groups- you can't just put all your mates into one group and all the people you don't like into the other.
- Beneficence: Participants' well-being has priority. Be nice to your participants!
- Respect: Participation should be voluntary and participants should be free to leave the trial at any time.
Be aware of the purposes of the Trial Protocol, the Investigator’s Brochure and the Patient Information and Consent documents
This is basically all the stuff you have to submit to get an ethics approval. The Trial Protocol is basically outlining what you're going to do, the Investigator's Brochure includes as much information about the drug as is currently known and the Patient Information and Consent Documents are the handouts that you're going to give to the patients to let them know that participation is voluntary, they can leave at any time, yada yada yada.
Aaaaaaand I'm done! (Except I am going to have a bonus post or two... stay tuned!)
Human Pharmacology II: Pharmacogenetics
Spoilers: 5' -> 3'! 5' -> 3'! Translation, transcription! Mutations! Etc etc etc!
Understand the basis of genetic variability generally and understand the concept of polymorphism in coding and regulatory regions of genes
Genetic variability occurs through two main mechanisms: mutations and crossing over (plus all the other stuff that happens during meiosis). For more information about mutations and their effects, see the following posts. (Kinda regretting not blogging about ANHB1101 now because the stuff in that was probably most relevant to this post. Ah well. On the upside, I didn't waste my time blogging about hominids. *shudders*)
Be aware that people can be genetically different in many aspects of drug kinetics (metabolism, especially) and in drug response
When mutations occur, enzymes might fail to function properly (if the mutation is in a gene coding for an enzyme) or someone might lack or have too much of some other important molecule. Enzymes are pretty important in regard to drug metabolism, so someone's levels of a particular enzyme might influence how much drug you need to give them, and so on and so forth.
Be aware that genetic polymorphism is just one part of the overall variability that exists in drug response. Age, diet, smoking, drug interaction and disease are others.
Okay cool, I'm aware now. Moving on...
Understand the example of thiopurine methyl transferase polymorphism
Thiopurine methyl transferase? That's a long name for an enzyme. Good thing it's also known as TPMT.
I'm not going to talk about TPMT straight off the bat though- instead I'm going to tell you a story about another drug called azathioprine.
Azathioprine is a drug used to treat leukaemia and inflammation. The way it does this is by becoming metabolised to form 6-mercaptopurine, which looks a helluva lot like guanine and as such sometimes gets substituted into DNA in place of guanine. This impairs DNA synthesis, which stops leukocytes (white blood cells) from replicating haphazardly. If too much 6-mercaptopurine is produced, however, too many leukocytes might die, leaving the patient open to infection.
Aside from being converted into 6-mercaptopurine, azathioprine can also be converted into two other metabolites. When it is metabolised by xanthine oxidase, an enzyme that everyone has roughly the same level of, it becomes thiouric acid, which is inactive. When azathioprine is metabolised by our old friend TPMT, which is genetically polymorphic (i.e. everyone has differences in TPMT activity), azathioprine becomes methylated and thus inactivated. Both xanthine oxidase and TPMT stop too much azathioprine from being metabolised to form 6-mercaptopurine. Since TPMT levels vary from person to person, it's TPMT that we have to watch out for. Patients with low TPMT activity may find themselves getting sick from too much azathioprine becoming 6-mercaptopurine.
The clinical significance of this is, of course, that screening patients by either measuring blood TPMT or sequencing their genome to see what their TPMT alleles are like (the former being much more practical!) might be important to find a safe and effective starting dose.
Understand the basis of genetic variability generally and understand the concept of polymorphism in coding and regulatory regions of genes
Genetic variability occurs through two main mechanisms: mutations and crossing over (plus all the other stuff that happens during meiosis). For more information about mutations and their effects, see the following posts. (Kinda regretting not blogging about ANHB1101 now because the stuff in that was probably most relevant to this post. Ah well. On the upside, I didn't waste my time blogging about hominids. *shudders*)
- Nucleic Acids- Mutations
- Mutation and Repair of DNA
- Enzymes Involved in DNA Replication and Repair
- Factors Influencing Development
- Molecular Cell Biology and Disease
Be aware that people can be genetically different in many aspects of drug kinetics (metabolism, especially) and in drug response
When mutations occur, enzymes might fail to function properly (if the mutation is in a gene coding for an enzyme) or someone might lack or have too much of some other important molecule. Enzymes are pretty important in regard to drug metabolism, so someone's levels of a particular enzyme might influence how much drug you need to give them, and so on and so forth.
Be aware that genetic polymorphism is just one part of the overall variability that exists in drug response. Age, diet, smoking, drug interaction and disease are others.
Okay cool, I'm aware now. Moving on...
Understand the example of thiopurine methyl transferase polymorphism
Thiopurine methyl transferase? That's a long name for an enzyme. Good thing it's also known as TPMT.
I'm not going to talk about TPMT straight off the bat though- instead I'm going to tell you a story about another drug called azathioprine.
Azathioprine is a drug used to treat leukaemia and inflammation. The way it does this is by becoming metabolised to form 6-mercaptopurine, which looks a helluva lot like guanine and as such sometimes gets substituted into DNA in place of guanine. This impairs DNA synthesis, which stops leukocytes (white blood cells) from replicating haphazardly. If too much 6-mercaptopurine is produced, however, too many leukocytes might die, leaving the patient open to infection.
Aside from being converted into 6-mercaptopurine, azathioprine can also be converted into two other metabolites. When it is metabolised by xanthine oxidase, an enzyme that everyone has roughly the same level of, it becomes thiouric acid, which is inactive. When azathioprine is metabolised by our old friend TPMT, which is genetically polymorphic (i.e. everyone has differences in TPMT activity), azathioprine becomes methylated and thus inactivated. Both xanthine oxidase and TPMT stop too much azathioprine from being metabolised to form 6-mercaptopurine. Since TPMT levels vary from person to person, it's TPMT that we have to watch out for. Patients with low TPMT activity may find themselves getting sick from too much azathioprine becoming 6-mercaptopurine.
The clinical significance of this is, of course, that screening patients by either measuring blood TPMT or sequencing their genome to see what their TPMT alleles are like (the former being much more practical!) might be important to find a safe and effective starting dose.
Human Pharmacology I: Adverse Drug Reactions
And now we're onto our last topic! This is a fairly relatable topic- the possibility of adverse drug interactions. Nasty stuff, but necessary to know.
Be able to provide a working definition of adverse drug reactions (ADRs) and demonstrate basic appreciation of the contribution ADRs make to the burden of human disease
Adverse drug reactions are nasty, unwanted reactions to drugs. In contrast to toxicity responses, which occur when too much of a drug is taken, adverse drug reactions occur when the normal dose of a drug is taken. Of course, this is bad for two reasons: firstly, the patient (who is probably already sick to be taking a drug) is made even sicker by the adverse reaction, and secondly it might make it harder to treat the patient's original condition as it limits the number of drugs that you can use.
Show awareness of the main categories of patients that are at most risk of ADRs, including the elderly, children and pregnant women
As the heading says, the elderly, children and pregnant women are most at risk of adverse drug reactions. Let's have a brief look at why.
The A in "Type A" stands for "augmented." Basically, Type A reactions are simply exaggerated effects of what the drug is normally meant to do. Hence, these are kind of predictable and can usually be worked around by switching to a different drug or different dosing regimen.
An example of a Type A drug reaction is insulin hypoglycaemia. Insulin is a hormone that lowers glucose levels in the blood. As you're likely well aware, patients with diabetes have to take insulin because their bodies either don't produce it or don't respond to it, leaving them at risk for hyperglycaemia. If they take too much insulin though, their glucose levels can drop considerably, leaving them hypoglycaemic, which is also pretty undesirable.
Identify 2 subcategories of immune-mediated Type B ADRs, namely immediate (IgE-mediated) and non-immediate (T-cell mediated) responses
Type B drug reactions are "bizarre" drug reactions. These tend to be unpredictable, though there may be certain genetic sensitivities to drugs (e.g. differences in HLA alleles). Allergic reactions to drugs are also classified as Type B drug reactions. Usually these involve some sort of priming, or prior exposure to the drug.
Type B reactions can also be further classified into "immediate" and "non-immediate" reactions. Let's take a look at them one at a time.
Immediate drug reactions occur within one hour of exposure. These tend to be mediated by specific IgE antibodies. IgE is produced by antigen-specific B-lymphocytes. It can then bind to Fc receptors on mast cells and basophils, which are both involved in mediating immune responses (or at least that's my basic understanding). Initially, this occurs during a period of sensitisation in which no symptoms occur. Later on, when the patient is re-exposed to the drug, IgE cross-links, which stimulates the release of histamine and a bunch of other stuff involved in allergic reactions. (I've never done any immunology, so I can't really go into more detail.)
Non-immediate drug reactions occur over an hour after the time of exposure (though within a few days). These tend to be mediated by T-cells. Firstly, dendritic cells process the drug antigen before internalising it and sending it to lymph nodes. Here, naïve T-cells get excited, and antigen-specific T-cells eventually begin making their way around the body. (Once again, this occurs during a period of sensitisation where no symptoms occur.) Upon re-exposure, those primed T-cells go nuts and the patient gets sick.
The last two paragraphs were really just skimming the surface of what happens. Once again, I've never done immunology, so I can't go into much more detail. Maybe we'll learn about this next semester!
Be able to provide a working definition of adverse drug reactions (ADRs) and demonstrate basic appreciation of the contribution ADRs make to the burden of human disease
Adverse drug reactions are nasty, unwanted reactions to drugs. In contrast to toxicity responses, which occur when too much of a drug is taken, adverse drug reactions occur when the normal dose of a drug is taken. Of course, this is bad for two reasons: firstly, the patient (who is probably already sick to be taking a drug) is made even sicker by the adverse reaction, and secondly it might make it harder to treat the patient's original condition as it limits the number of drugs that you can use.
Show awareness of the main categories of patients that are at most risk of ADRs, including the elderly, children and pregnant women
As the heading says, the elderly, children and pregnant women are most at risk of adverse drug reactions. Let's have a brief look at why.
- Elderly: More likely to receive multiple drugs for various chronic conditions. Also there are other physical changes that might affect pharmacokinetics, including reduced drug metabolism, reduced drug protein binding and reduced renal excretion.
- Children: Different capacities for renal excretion and hepatic metabolism as compared to adults. There is also limited information on the safety of many drugs in children.
- Pregnant women: Altered activity of drug metabolising enzymes. A whole bunch of physiologic changes, such as increased fat and decreased protein binding, which might affect pharmacokinetic parameters such as volume of distribution. There is also a risk of teratogenicity with regards to the woman's baby.
The A in "Type A" stands for "augmented." Basically, Type A reactions are simply exaggerated effects of what the drug is normally meant to do. Hence, these are kind of predictable and can usually be worked around by switching to a different drug or different dosing regimen.
An example of a Type A drug reaction is insulin hypoglycaemia. Insulin is a hormone that lowers glucose levels in the blood. As you're likely well aware, patients with diabetes have to take insulin because their bodies either don't produce it or don't respond to it, leaving them at risk for hyperglycaemia. If they take too much insulin though, their glucose levels can drop considerably, leaving them hypoglycaemic, which is also pretty undesirable.
Identify 2 subcategories of immune-mediated Type B ADRs, namely immediate (IgE-mediated) and non-immediate (T-cell mediated) responses
Type B drug reactions are "bizarre" drug reactions. These tend to be unpredictable, though there may be certain genetic sensitivities to drugs (e.g. differences in HLA alleles). Allergic reactions to drugs are also classified as Type B drug reactions. Usually these involve some sort of priming, or prior exposure to the drug.
Type B reactions can also be further classified into "immediate" and "non-immediate" reactions. Let's take a look at them one at a time.
Immediate drug reactions occur within one hour of exposure. These tend to be mediated by specific IgE antibodies. IgE is produced by antigen-specific B-lymphocytes. It can then bind to Fc receptors on mast cells and basophils, which are both involved in mediating immune responses (or at least that's my basic understanding). Initially, this occurs during a period of sensitisation in which no symptoms occur. Later on, when the patient is re-exposed to the drug, IgE cross-links, which stimulates the release of histamine and a bunch of other stuff involved in allergic reactions. (I've never done any immunology, so I can't really go into more detail.)
Non-immediate drug reactions occur over an hour after the time of exposure (though within a few days). These tend to be mediated by T-cells. Firstly, dendritic cells process the drug antigen before internalising it and sending it to lymph nodes. Here, naïve T-cells get excited, and antigen-specific T-cells eventually begin making their way around the body. (Once again, this occurs during a period of sensitisation where no symptoms occur.) Upon re-exposure, those primed T-cells go nuts and the patient gets sick.
The last two paragraphs were really just skimming the surface of what happens. Once again, I've never done immunology, so I can't go into much more detail. Maybe we'll learn about this next semester!
Chemotherapy IV: Antifungal Drugs
Last post for this topic!
Show an awareness of the 3 broad classes of fungal species
The three broad classes of fungal species are yeasts, moulds and dimorphic fungi.
Yeasts are often called "sprouting fungi." They are the sorts of moulds you find if you leave an orange on the bottom of the Lost Property box for too long (yup, found this when helping to clean out the Lost Property box back in primary school). Their cells are oval-shaped and contain granules, and often vacuoles as well. They grow by forming buds which increase in size before separating out.
Moulds are what you tend to find on stale bread and so forth. They grow as multicellular filaments known as hyphae, which branch to form a dense mat called a mycelium. The three main genera of moulds are pencillum, aspergillus and mucor.
Dimorphic fungi display characteristics of yeasts or moulds, depending on temperature. They tend to be more filamentous and mould-like at lower temperatures, and more yeast-like at higher temperatures. They can grow as single cells or as rudimentary filaments known as pseudohyphae. These pseudohyphae are responsible for the invasive properties of these fungi.
Show appreciation of the four major classes of fungal infections in humans, the factors driving their rising significance, and the challenges accompanying their control with drugs.
The four major classes of fungal infections, or mycoses (singular mycosis), are simply based off where they occur. Superficial mycoses appear on the epidermis of the skin, cutaneous appear a bit deeper, subcutaneous appear in the dermal and underlying layers of the skin and systemic mycoses are very widespread. As a general rule, the deeper the mycosis is, the more severe it is. Normally, fungi do their damage by blocking things (e.g. blood vessels), rather than secreting toxins or anything like that.
Fungi normally aren't that damaging to our bodies. Most fungal infections are opportunistic infections- that is, healthy people can clear them without an issue, but they can be quite nasty towards immunocompromised individuals. This is important to note, because there are several factors that can affect a person's immune system. For example, medications such as cancer chemotherapy and corticosteroids can impair a person's immune system. Heavy antibiotic use can also worsen the impact of a fungal infection as non-pathogenic bacteria, which normally help us eliminate fungi, may be eliminated. Pre-existing conditions such as HIV/AIDS and diabetes may also increase an individual's vulnerability towards fungi.
The tricky thing about treating fungi is that they are also eukaryotic, and therefore have many more similarities to us as compared to bacteria (which are prokaryotic) and viruses (which are essentially just packages of nucleic acids). Hence, there are fewer "safe targets" for antifungal drugs. The main differences that are targeted in drug therapy are distinctive sterols in the cell membrane (we have cholesterol, whereas they have other funky things like ergosterol and lanosterol) and the fungal cell wall which is comprised of molecules called glucans.
Demonstrate awareness of 3 main classes of antifungal agents, including their basic mechanism of action, clinical uses, and most significant side-effects.
Amphotericin
Amphotericin is one of the earlier antifungal drugs. It works by binding to ergosterol, which is a sterol found only in fungal cell walls, as mentioned above. When it binds to ergosterol, it creates pores through which essential ions such as K+ can escape. This leads to death of the fungus.
Azoles
Azoles, so called because they have special rings like imidazole and triazole rings, also interfere with ergosterol. Instead of binding to ergosterol directly, however, they block an enzyme called 14-α-sterol demethylase, which converts lanosterol (another fungus-only sterol) to ergosterol. This causes depletion of ergosterol, which causes membrane dysfunction, impaired replication, and other stuff that is nasty for the fungus and good for us. (Azoles aren't that nasty though- they're fungostatic, not fungocidal, which means that they just slow down the fungus rather than kill it outright.)
Echinocandins
Echinocandins work via a different pathway altogether. They inhibit 1,3-β-D-glucan synthase (what a mouthful), which blocks the synthesis of 1,3-β-D-glucan. This is one of the glucans that is important in the synthesis of the cell wall. Hence, inhibiting this enzyme is bad for the fungus. That poor fungus.
Show an awareness of the 3 broad classes of fungal species
The three broad classes of fungal species are yeasts, moulds and dimorphic fungi.
Yeasts are often called "sprouting fungi." They are the sorts of moulds you find if you leave an orange on the bottom of the Lost Property box for too long (yup, found this when helping to clean out the Lost Property box back in primary school). Their cells are oval-shaped and contain granules, and often vacuoles as well. They grow by forming buds which increase in size before separating out.
Moulds are what you tend to find on stale bread and so forth. They grow as multicellular filaments known as hyphae, which branch to form a dense mat called a mycelium. The three main genera of moulds are pencillum, aspergillus and mucor.
Dimorphic fungi display characteristics of yeasts or moulds, depending on temperature. They tend to be more filamentous and mould-like at lower temperatures, and more yeast-like at higher temperatures. They can grow as single cells or as rudimentary filaments known as pseudohyphae. These pseudohyphae are responsible for the invasive properties of these fungi.
Show appreciation of the four major classes of fungal infections in humans, the factors driving their rising significance, and the challenges accompanying their control with drugs.
The four major classes of fungal infections, or mycoses (singular mycosis), are simply based off where they occur. Superficial mycoses appear on the epidermis of the skin, cutaneous appear a bit deeper, subcutaneous appear in the dermal and underlying layers of the skin and systemic mycoses are very widespread. As a general rule, the deeper the mycosis is, the more severe it is. Normally, fungi do their damage by blocking things (e.g. blood vessels), rather than secreting toxins or anything like that.
Fungi normally aren't that damaging to our bodies. Most fungal infections are opportunistic infections- that is, healthy people can clear them without an issue, but they can be quite nasty towards immunocompromised individuals. This is important to note, because there are several factors that can affect a person's immune system. For example, medications such as cancer chemotherapy and corticosteroids can impair a person's immune system. Heavy antibiotic use can also worsen the impact of a fungal infection as non-pathogenic bacteria, which normally help us eliminate fungi, may be eliminated. Pre-existing conditions such as HIV/AIDS and diabetes may also increase an individual's vulnerability towards fungi.
The tricky thing about treating fungi is that they are also eukaryotic, and therefore have many more similarities to us as compared to bacteria (which are prokaryotic) and viruses (which are essentially just packages of nucleic acids). Hence, there are fewer "safe targets" for antifungal drugs. The main differences that are targeted in drug therapy are distinctive sterols in the cell membrane (we have cholesterol, whereas they have other funky things like ergosterol and lanosterol) and the fungal cell wall which is comprised of molecules called glucans.
Demonstrate awareness of 3 main classes of antifungal agents, including their basic mechanism of action, clinical uses, and most significant side-effects.
Amphotericin
Amphotericin is one of the earlier antifungal drugs. It works by binding to ergosterol, which is a sterol found only in fungal cell walls, as mentioned above. When it binds to ergosterol, it creates pores through which essential ions such as K+ can escape. This leads to death of the fungus.
Azoles
Azoles, so called because they have special rings like imidazole and triazole rings, also interfere with ergosterol. Instead of binding to ergosterol directly, however, they block an enzyme called 14-α-sterol demethylase, which converts lanosterol (another fungus-only sterol) to ergosterol. This causes depletion of ergosterol, which causes membrane dysfunction, impaired replication, and other stuff that is nasty for the fungus and good for us. (Azoles aren't that nasty though- they're fungostatic, not fungocidal, which means that they just slow down the fungus rather than kill it outright.)
Echinocandins
Echinocandins work via a different pathway altogether. They inhibit 1,3-β-D-glucan synthase (what a mouthful), which blocks the synthesis of 1,3-β-D-glucan. This is one of the glucans that is important in the synthesis of the cell wall. Hence, inhibiting this enzyme is bad for the fungus. That poor fungus.
Thursday, May 26, 2016
Chemotherapy III: Antiviral Drugs
Now we're going to move onto another target: viruses!
Understand basic viral structure, biology, life cycle and pathogenicity.
Viruses are essentially just packages of DNA or RNA. They don't have organelles or anything like that to sustain themselves: instead they have to go into a cell and hijack the cell's machinery in order to replicate and thrive. (This is why designing antiviral drugs can be hard: you want to stop the virus from replicating, but you don't want to stop normal DNA replication from occurring either.)
Viruses have a nucleic acid core, surrounded by a coat, or capsid. This capsid is made up of proteins called capsomeres. The nucleic acid and capsid together are collectively known as a nucleocapsid. The capsomeres are of importance as their binding to receptors on the cell allows the virus to be taken up by the cell.
Here's what happens, in a little more detail than in the first paragraph: the capsomere proteins bind to receptors on the cell, the virus gets taken up and sheds its capsid, and viral DNA or RNA is produced. This viral DNA/RNA goes on to code the structural, enzymatic and regulatory proteins that the virus needs. These include capsomeres, allowing viral proteins to be packaged up and exocytosed from the cell. This allows the virus to go on and infect more cells.
Understand the mechanistic basis for the use of acyclovir as the first effective antiviral drug.
Acyclovir, as its name suggests, does not have a ring (a = without, cyclo = ring). Acyclovir is actually similar to deoxyguanosine (one of the bases of DNA), but it is missing that fundamental cyclic ring structure. This also means that it's missing a 3'-OH, so DNA synthesis cannot continue if acyclovir is added instead of deoxyguanosine. In this way, acyclovir acts as a purine analogue inhibitor- it's a purine analogue (i.e. it looks like a purine, in this case guanosine), and by doing so, it inhibits viral DNA replication. This is because it either competitively inhibits DNA polymerase, or as alluded to a couple of sentences ago, it can bind in place of a guanosine and prevent further synthesis from occurring.
Acyclovir can't do this on its own, however. As mentioned in a previous post, nucleosides lacking phosphate groups aren't added onto a chain- instead, nucleoside triphosphates are required. Hence, acyclovir is actually a prodrug that requires some more processing to be able to do its job. Each processing step simply involves addition of a phosphate group. Firstly, this is done by viral thymidine kinase (a kinase produced by the virus, so essentially the virus is being complicit in its own murder!). The second and third phosphorylations are simply carried out by kinases normally present in the cell.
Demonstrate a basic understanding of AIDS.
AIDS stands for Acquired Immune Deficiency Syndrome and is defined by a loss of CD4+ T-cells (important in our immune system). It comes about as a result of immune system damage from the Human Immunodeficiency Virus (HIV). The symptoms of HIV can be quite varied, as the weakened immune system lays down the foundation for opportunistic infections to occur. (Opportunistic infections are those that wouldn't normally hurt a healthy person, but can be quite deadly in people without functional immune systems.)
HIV is an RNA retrovirus, which means that it has RNA as well as an enzyme called reverse transcriptase that can turn that RNA into DNA. It exists in two forms: HIV-1, which is more common, and HIV-2, which is less virulent. HIV can attach to its target cells through the interaction of HIV glycoproteins with surface receptors on a variety of cells, including T-lymphocytes expressing CD4 glycoprotein (i.e. those CD4+ T-cells that I referred to earlier).
The main factors to take into consideration when deciding to implement drug therapy are CD4+ T-lymphocyte numbers as well as HIV RNA copy number. CD4+ T-lymphocyte numbers decrease as the disease progresses. HIV RNA copy number, indicative of viral load, increases as the disease progresses.
Show appreciation for major drug classes used to treat HIV-infected patients.
The key targets for treating HIV include receptor binding (e.g. CCR5 receptor blockers), fusion, reverse transcriptase, integrase (inserts viral DNA into the host DNA) and proteases (which in this case can actually aid viral maturation- more on these later). Here I will be focusing on reverse transcriptase inhibitors and HIV Protease inhibitors.
First off, a quick look at reverse transcriptase inhibitors. These come in two classes: NRTIs (Nucleoside Reverse Transcriptase Inhibitors- they work by mimicking nucleosides, kinda like the purine analogue inhibitors I talked about earlier), and NNRTIs (Non-Nucleoside Reverse Transcriptase Inhibitors- do not look like nucleosides). The NRTI class includes AZT (azidothymidine), which was the first anti-HIV drug.
HIV Protease is kinda unique. Remember how I said that viral DNA/RNA codes for coat proteins that help package up more virus and send it out of the cell? Well, HIV protease, weirdly enough, plays a role in the synthesis of the coat proteins. You see, HIV makes "polyproteins," which are essentially multiple proteins all mashed together. Proteases break them up into individual coat proteins. Drugs that target this pathway include saquinavir and atazanavir.
Usually, HIV patients are put on several drugs to manage their symptoms. HAART (Highly Active Antiretroviral Therapy) is the fancy name given to these cocktails of drugs. HAART usually consists of an NRTI, an NNRTI and a protease inhibitor. So far, this has been our most effective treatment against HIV.
Understand how drug resistance can limit the effectiveness of anti-HIV drugs
Unfortunately, unlike bacteria, HIV is also prone to building up resistance mechanisms. This is partly because HIV's reverse transcriptase has a low fidelity (i.e. very error-prone), so mutations can occur readily. Some of these mutations may confer resistance to a drug. There are now some drugs available against the most common mutant variants, but unfortunately these are still quite expensive.
Understand basic viral structure, biology, life cycle and pathogenicity.
Viruses are essentially just packages of DNA or RNA. They don't have organelles or anything like that to sustain themselves: instead they have to go into a cell and hijack the cell's machinery in order to replicate and thrive. (This is why designing antiviral drugs can be hard: you want to stop the virus from replicating, but you don't want to stop normal DNA replication from occurring either.)
Viruses have a nucleic acid core, surrounded by a coat, or capsid. This capsid is made up of proteins called capsomeres. The nucleic acid and capsid together are collectively known as a nucleocapsid. The capsomeres are of importance as their binding to receptors on the cell allows the virus to be taken up by the cell.
Here's what happens, in a little more detail than in the first paragraph: the capsomere proteins bind to receptors on the cell, the virus gets taken up and sheds its capsid, and viral DNA or RNA is produced. This viral DNA/RNA goes on to code the structural, enzymatic and regulatory proteins that the virus needs. These include capsomeres, allowing viral proteins to be packaged up and exocytosed from the cell. This allows the virus to go on and infect more cells.
Understand the mechanistic basis for the use of acyclovir as the first effective antiviral drug.
Acyclovir, as its name suggests, does not have a ring (a = without, cyclo = ring). Acyclovir is actually similar to deoxyguanosine (one of the bases of DNA), but it is missing that fundamental cyclic ring structure. This also means that it's missing a 3'-OH, so DNA synthesis cannot continue if acyclovir is added instead of deoxyguanosine. In this way, acyclovir acts as a purine analogue inhibitor- it's a purine analogue (i.e. it looks like a purine, in this case guanosine), and by doing so, it inhibits viral DNA replication. This is because it either competitively inhibits DNA polymerase, or as alluded to a couple of sentences ago, it can bind in place of a guanosine and prevent further synthesis from occurring.
Acyclovir can't do this on its own, however. As mentioned in a previous post, nucleosides lacking phosphate groups aren't added onto a chain- instead, nucleoside triphosphates are required. Hence, acyclovir is actually a prodrug that requires some more processing to be able to do its job. Each processing step simply involves addition of a phosphate group. Firstly, this is done by viral thymidine kinase (a kinase produced by the virus, so essentially the virus is being complicit in its own murder!). The second and third phosphorylations are simply carried out by kinases normally present in the cell.
Demonstrate a basic understanding of AIDS.
AIDS stands for Acquired Immune Deficiency Syndrome and is defined by a loss of CD4+ T-cells (important in our immune system). It comes about as a result of immune system damage from the Human Immunodeficiency Virus (HIV). The symptoms of HIV can be quite varied, as the weakened immune system lays down the foundation for opportunistic infections to occur. (Opportunistic infections are those that wouldn't normally hurt a healthy person, but can be quite deadly in people without functional immune systems.)
HIV is an RNA retrovirus, which means that it has RNA as well as an enzyme called reverse transcriptase that can turn that RNA into DNA. It exists in two forms: HIV-1, which is more common, and HIV-2, which is less virulent. HIV can attach to its target cells through the interaction of HIV glycoproteins with surface receptors on a variety of cells, including T-lymphocytes expressing CD4 glycoprotein (i.e. those CD4+ T-cells that I referred to earlier).
The main factors to take into consideration when deciding to implement drug therapy are CD4+ T-lymphocyte numbers as well as HIV RNA copy number. CD4+ T-lymphocyte numbers decrease as the disease progresses. HIV RNA copy number, indicative of viral load, increases as the disease progresses.
Show appreciation for major drug classes used to treat HIV-infected patients.
The key targets for treating HIV include receptor binding (e.g. CCR5 receptor blockers), fusion, reverse transcriptase, integrase (inserts viral DNA into the host DNA) and proteases (which in this case can actually aid viral maturation- more on these later). Here I will be focusing on reverse transcriptase inhibitors and HIV Protease inhibitors.
First off, a quick look at reverse transcriptase inhibitors. These come in two classes: NRTIs (Nucleoside Reverse Transcriptase Inhibitors- they work by mimicking nucleosides, kinda like the purine analogue inhibitors I talked about earlier), and NNRTIs (Non-Nucleoside Reverse Transcriptase Inhibitors- do not look like nucleosides). The NRTI class includes AZT (azidothymidine), which was the first anti-HIV drug.
HIV Protease is kinda unique. Remember how I said that viral DNA/RNA codes for coat proteins that help package up more virus and send it out of the cell? Well, HIV protease, weirdly enough, plays a role in the synthesis of the coat proteins. You see, HIV makes "polyproteins," which are essentially multiple proteins all mashed together. Proteases break them up into individual coat proteins. Drugs that target this pathway include saquinavir and atazanavir.
Usually, HIV patients are put on several drugs to manage their symptoms. HAART (Highly Active Antiretroviral Therapy) is the fancy name given to these cocktails of drugs. HAART usually consists of an NRTI, an NNRTI and a protease inhibitor. So far, this has been our most effective treatment against HIV.
Understand how drug resistance can limit the effectiveness of anti-HIV drugs
Unfortunately, unlike bacteria, HIV is also prone to building up resistance mechanisms. This is partly because HIV's reverse transcriptase has a low fidelity (i.e. very error-prone), so mutations can occur readily. Some of these mutations may confer resistance to a drug. There are now some drugs available against the most common mutant variants, but unfortunately these are still quite expensive.
Chemotherapy II: Antibacterial Drugs
As promised in my last post, this post will discuss antibiotics in more detail!
Explain key concepts relevant to the action of antibacterial drugs, including spectrum of action, mechanism of action, individuation of dosing, drug resistance, etc.
Uhh, I'm fairly sure I've done this already...
Show awareness of major drugs such as the penicillins and related drug classes that interfere with the synthesis of the bacterial cell wall.
Firstly, a few points on what the bacterial cell wall looks like and how it is synthesised. Bacterial cell walls are made up of peptidoglycans, which are essentially lattices of glycan (sugar) chains. These chains are crosslinked by short peptide chains. The peptidoglycan layer varies in thickness depending on the type of bacteria- gram-positive bacteria have thicker peptidoglycans than gram-negative. (Gram-positive/negative simply refers to whether or not they are stained by Gram's stain.)
The actual cell wall production process is kinda complex, but the main gist of it is that the cross-linking peptide chains are synthesised on the inside of the cell and are dragged across the cell membrane by binding to a 55-carbon lipid. While it's being dragged across, some other stuff gets added, including the crosslinking peptide chains. After being towed across the membrane, the crosslinking chains can do their job and crosslink stuff, helped along by enzymes like transpeptidase.
The important part for you to know is that transpeptidase catalyses crosslinkages, and without crosslinkages a stable cell wall does not form. β-lactam drugs, so called because they have a 4-carbon β-lactam ring, form covalent bonds with transpeptidases, irreversibly stopping them from forming crosslinks. β-lactam drugs include the penicillins, the cephalosporins and the carbapenems.
There are several problems emerging with the use of penicillins. Firstly, people are developing allergies to penicillin. This is because the β-lactam ring of penicillin can open up to form penicilloic acid, which can then bind to other stuff, creating new and interesting antigens that some people's immune systems don't seem to like. Penicillin can result in some nasty allergic responses in these people.
Another problem with the use of penicillins (or with antibiotics in general) is the emergence of resistance. Bacteria can build up resistance to penicillin via the formation of β-lactamases (enzymes that break down the all-important β-lactam ring), stopping the drug from reaching the target area of the cell or by creating new proteins that penicillin can't bind to. The first mechanism of resistance that I mentioned, formation of β-lactamases, can be circumvented by the use of clavulanic acid. This is an irreversible inhibitor of β-lactamases. This is why amoxicillin (one of the penicillins) is often co-administered with clavulanic acid.
Show an understanding of the basic features of bacterial protein synthesis, including an appreciation for how specific antibacterial drugs block specific steps.
Protein synthesis inhibitors can inhibit pretty much every stage of the process of protein synthesis. (I have a post here specifically on prokaryotic protein synthesis, though it might be too much detail for the purposes of this post.) The four main types that I am going to talk about are the aminoglycasides, tetracyclines, amphenicols and macrolides.
Aminoglycosides are amino sugars (amino = amino, glyco = sugar). As both amino groups and sugars tend to be quite polar and hydrophilic, they don't cross cell membranes so well and thus they tend to have poor absorption. Their mechanism of action is by binding to the decoding site on the 30S ribosome, leading to misreading of the mRNA template and the insertion of the wrong amino acid. This, in turn, leads to non-functional proteins, making things quite difficult for the bacteria. Aminoglycosides also happen to be bactericidal- they are the only bactericidal protein synthesis inhibitors (other protein synthesis inhibitors are bacteriostatic).
Tetracyclines have four rings (tetra = four, cycl = ring). They prevent tRNAs from binding to the mRNA-ribosome complex, halting protein synthesis.
Chloramphenicol (a major player in the amphenicols) binds to the 50S ribosomal subunit, preventing the formation of peptide bonds between adjacent amino acids.
Macrolides are fairly large molecules that bind to the 50S subunit, preventing the tRNA from moving from the A site to the P site (these sites are mentioned in a previous post). They do this by plugging up the "tunnel" within the ribosome.
A quick note on polymixins
These didn't really fit into the lecture outcomes, but since they were mentioned during the lecture, I might as well mention them here. Polymixins are antibiotics used pretty much of last resort as they are quite toxic. They are plasma membrane permeabilising agents that work by acting as a "detergent," disrupting the cell membrane by interacting with membrane phospholipids. Despite their toxic profile, they are making a bit of a comeback due to the rise of antibiotic-resistant bacteria.
Explain key concepts relevant to the action of antibacterial drugs, including spectrum of action, mechanism of action, individuation of dosing, drug resistance, etc.
Uhh, I'm fairly sure I've done this already...
Show awareness of major drugs such as the penicillins and related drug classes that interfere with the synthesis of the bacterial cell wall.
Firstly, a few points on what the bacterial cell wall looks like and how it is synthesised. Bacterial cell walls are made up of peptidoglycans, which are essentially lattices of glycan (sugar) chains. These chains are crosslinked by short peptide chains. The peptidoglycan layer varies in thickness depending on the type of bacteria- gram-positive bacteria have thicker peptidoglycans than gram-negative. (Gram-positive/negative simply refers to whether or not they are stained by Gram's stain.)
The actual cell wall production process is kinda complex, but the main gist of it is that the cross-linking peptide chains are synthesised on the inside of the cell and are dragged across the cell membrane by binding to a 55-carbon lipid. While it's being dragged across, some other stuff gets added, including the crosslinking peptide chains. After being towed across the membrane, the crosslinking chains can do their job and crosslink stuff, helped along by enzymes like transpeptidase.
The important part for you to know is that transpeptidase catalyses crosslinkages, and without crosslinkages a stable cell wall does not form. β-lactam drugs, so called because they have a 4-carbon β-lactam ring, form covalent bonds with transpeptidases, irreversibly stopping them from forming crosslinks. β-lactam drugs include the penicillins, the cephalosporins and the carbapenems.
There are several problems emerging with the use of penicillins. Firstly, people are developing allergies to penicillin. This is because the β-lactam ring of penicillin can open up to form penicilloic acid, which can then bind to other stuff, creating new and interesting antigens that some people's immune systems don't seem to like. Penicillin can result in some nasty allergic responses in these people.
Another problem with the use of penicillins (or with antibiotics in general) is the emergence of resistance. Bacteria can build up resistance to penicillin via the formation of β-lactamases (enzymes that break down the all-important β-lactam ring), stopping the drug from reaching the target area of the cell or by creating new proteins that penicillin can't bind to. The first mechanism of resistance that I mentioned, formation of β-lactamases, can be circumvented by the use of clavulanic acid. This is an irreversible inhibitor of β-lactamases. This is why amoxicillin (one of the penicillins) is often co-administered with clavulanic acid.
Show an understanding of the basic features of bacterial protein synthesis, including an appreciation for how specific antibacterial drugs block specific steps.
Protein synthesis inhibitors can inhibit pretty much every stage of the process of protein synthesis. (I have a post here specifically on prokaryotic protein synthesis, though it might be too much detail for the purposes of this post.) The four main types that I am going to talk about are the aminoglycasides, tetracyclines, amphenicols and macrolides.
Aminoglycosides are amino sugars (amino = amino, glyco = sugar). As both amino groups and sugars tend to be quite polar and hydrophilic, they don't cross cell membranes so well and thus they tend to have poor absorption. Their mechanism of action is by binding to the decoding site on the 30S ribosome, leading to misreading of the mRNA template and the insertion of the wrong amino acid. This, in turn, leads to non-functional proteins, making things quite difficult for the bacteria. Aminoglycosides also happen to be bactericidal- they are the only bactericidal protein synthesis inhibitors (other protein synthesis inhibitors are bacteriostatic).
Tetracyclines have four rings (tetra = four, cycl = ring). They prevent tRNAs from binding to the mRNA-ribosome complex, halting protein synthesis.
Chloramphenicol (a major player in the amphenicols) binds to the 50S ribosomal subunit, preventing the formation of peptide bonds between adjacent amino acids.
Macrolides are fairly large molecules that bind to the 50S subunit, preventing the tRNA from moving from the A site to the P site (these sites are mentioned in a previous post). They do this by plugging up the "tunnel" within the ribosome.
A quick note on polymixins
These didn't really fit into the lecture outcomes, but since they were mentioned during the lecture, I might as well mention them here. Polymixins are antibiotics used pretty much of last resort as they are quite toxic. They are plasma membrane permeabilising agents that work by acting as a "detergent," disrupting the cell membrane by interacting with membrane phospholipids. Despite their toxic profile, they are making a bit of a comeback due to the rise of antibiotic-resistant bacteria.
Wednesday, May 25, 2016
Chemotherapy I: Basic Concepts
If you saw the title of this post and thought, "Cool, we're going to be learning how to cure cancer!" then you should prepare to be sorely disappointed. Chemotherapy is actually a broad term that refers to treating diseases with drugs that are toxic towards whatever pathogen is making you sick. That means that even stuff like antibiotics and antivirals count as chemotherapy agents. In fact, over the next few posts, I will only be covering antibiotics, antivirals and antifungals. (Apparently anticancer drugs are in later pharmacology units, but I'm currently not intending to specialise in pharmacology.)
Demonstrate awareness of the profound impact antibiotics have had on human health throughout the past century
In a nutshell, antibiotics stopped us from dying of lots of infectious diseases. Now that we don't have as many of these diseases to worry about, we're living long enough to get heart disease and cancer instead. w00t w00t. Oh, and antibiotic resistance is giving us a lot of new challenges. The war on microbes isn't over yet.
Show awareness of key concepts relevant to the action of antibacterial drugs
Spectrum of action
Spectrum of action refers to the range of bacteria that an antibiotic can kill. A narrow spectrum antibiotic will only be able to kill one or two bacteria well (e.g. isoniazid is good against mycobacteria, which causes tuberculosis), whereas a broad spectrum antibiotic (e.g. tetracycline) can act against a range of bacteria.
Mechanism of action (show understanding of difference between bacteriostatic and bactericidal drug action)
Okay, first a quick note on bacteriostatic and bacteriocidal drugs. Bacteriostatic drugs slow down the growth and/or replication of the bacteria, whereas bacteriocidal drugs outright kill them ("-cidal" means kill, as in "suicidal" or "homicidal"), possibly by the generation of free radicals.
As for the specific mechanisms, antibiotics target differences in key pathways between bacteria and human cells so that the bacteria die and we don't. Specifically, antibiotics tend to target cell wall synthesis, folate synthesis, protein synthesis, nucleic acid synthesis and so forth. Most antibiotics target cell wall synthesis (these tend to be bacteriocidal) or protein synthesis (these tend to be bacteriostatic).
Individualization of dosing
One way that drug therapy can be individualised is by taking a microbe sample from the patient and testing to see which antibiotics kill the microbe. This is done by culturing the microbe on an agar gel and putting on some discs that contain different antibiotics. After incubation, you can observe where the organism has grown. Discs that are surrounded by spaces with little or no growth ("zones of inhibition") are likely to contain the effective antibiotics that you need. The larger the "zone of inhibition," the more effective that particular drug. If there is no zone of inhibition, you're probably looking at a resistant organism- eep!
Obviously, this process takes time. Hence, a patient might be started off on a broad-spectrum antibiotic until a more effective drug for that particular organism is found.
Another thing to take into consideration when individualising dosing regimens is pharmacokinetics: y'know, stuff like clearance, volume of distribution and so forth. These may differ from person to person due to differing liver function, kidney function etc.
Drug resistance
As I'm sure you know, the use of antibiotics has gradually led to the emergence of antibiotic-resistant strains of bacteria. There are a variety of ways in which bacteria can acquire resistance to antibiotics:
Demonstrate awareness of the profound impact antibiotics have had on human health throughout the past century
In a nutshell, antibiotics stopped us from dying of lots of infectious diseases. Now that we don't have as many of these diseases to worry about, we're living long enough to get heart disease and cancer instead. w00t w00t. Oh, and antibiotic resistance is giving us a lot of new challenges. The war on microbes isn't over yet.
Show awareness of key concepts relevant to the action of antibacterial drugs
Spectrum of action
Spectrum of action refers to the range of bacteria that an antibiotic can kill. A narrow spectrum antibiotic will only be able to kill one or two bacteria well (e.g. isoniazid is good against mycobacteria, which causes tuberculosis), whereas a broad spectrum antibiotic (e.g. tetracycline) can act against a range of bacteria.
Mechanism of action (show understanding of difference between bacteriostatic and bactericidal drug action)
Okay, first a quick note on bacteriostatic and bacteriocidal drugs. Bacteriostatic drugs slow down the growth and/or replication of the bacteria, whereas bacteriocidal drugs outright kill them ("-cidal" means kill, as in "suicidal" or "homicidal"), possibly by the generation of free radicals.
As for the specific mechanisms, antibiotics target differences in key pathways between bacteria and human cells so that the bacteria die and we don't. Specifically, antibiotics tend to target cell wall synthesis, folate synthesis, protein synthesis, nucleic acid synthesis and so forth. Most antibiotics target cell wall synthesis (these tend to be bacteriocidal) or protein synthesis (these tend to be bacteriostatic).
Individualization of dosing
One way that drug therapy can be individualised is by taking a microbe sample from the patient and testing to see which antibiotics kill the microbe. This is done by culturing the microbe on an agar gel and putting on some discs that contain different antibiotics. After incubation, you can observe where the organism has grown. Discs that are surrounded by spaces with little or no growth ("zones of inhibition") are likely to contain the effective antibiotics that you need. The larger the "zone of inhibition," the more effective that particular drug. If there is no zone of inhibition, you're probably looking at a resistant organism- eep!
Obviously, this process takes time. Hence, a patient might be started off on a broad-spectrum antibiotic until a more effective drug for that particular organism is found.
Another thing to take into consideration when individualising dosing regimens is pharmacokinetics: y'know, stuff like clearance, volume of distribution and so forth. These may differ from person to person due to differing liver function, kidney function etc.
Drug resistance
As I'm sure you know, the use of antibiotics has gradually led to the emergence of antibiotic-resistant strains of bacteria. There are a variety of ways in which bacteria can acquire resistance to antibiotics:
- They can coat themselves with a "protective slime" or a protective membrane, preventing the drug from reaching the target
- They can inactivate the drug, for example by using β-lactamases (which destroy the β-lactam rings present in some antibiotics- more on this in a later post)
- They can alter the drug target so that the drug can no longer bind
- They can pump the drug out via an efflux pump
- They can develop bypass pathways so that it doesn't matter if the main pathway gets blocked by a drug
Wednesday, May 11, 2016
Nuclear Receptors
Fourth and final type of receptors! (If you've completely forgotten everything about receptors, please take a look at my post on Receptors and Other Drug Targets.)
Recall classification of nuclear receptors and provide examples of ligands
Nuclear receptors can be classified into four categories, imaginatively named Class I, Class II, Class III and Class IV. I'm going to present them in a table, because y'know, madddd HTML skillz:
*REs are recognition sites that the nuclear receptors recognise and bind to.
**Not in the lecture or the textbook, so I assume it's not important.
Describe the general structure of nuclear receptors
Nuclear receptors have four main domains, which I'm going to illustrate using my maddddd HTML skillz (since the excitement of finding out that I still remember how to use HTML still hasn't worn off...)
The N-terminal domain is the least conserved (i.e. most prone to variation in length and sequence). It contains Activation Function 1 (AF1), which binds to co-regulators independent of ligand binding (i.e. it doesn't care whether ligands are bound or not, it'll bind to co-regulators no matter what).
The DNA-binding domain is highly conserved. As the name suggests, it is where the nuclear receptor binds to DNA by recognising response elements (REs, or if you want to be more technical, HREs- hormone response elements). It has two zinc fingers formed by cysteine-rich loops. (For more information about zinc fingers, please see my earlier post on transcription factors.)
The hinge region is quite flexible, and may be responsible for dimerisation and conformation changes of the receptor.
The ligand-binding domain, as the name suggests, is where ligands bind. It is highly conserved and has 12 α-helices which form a hydrophobic "pocket." It also contains Activation Function 2 (AF2), which is kinda like AF1, but unlike AF1, AF2 actually cares if ligands are bound or not (i.e. it binds co-regulators in a ligand-dependent manner).
Describe the mechanism of activation of glucocorticoid receptor and peroxisome proliferator-activated receptor, and the effects on gene transcription
Glucocorticoid Receptor
Glucocorticoid receptors are class I receptors that is synthesised from the splicing together of 9 exons (the ninth of which has an α and β isoform, but I don't know how important it is for us to remember that). From the table above, you should have already figured out that the glucorticoid receptor's Class I categorisation means that it is activated by hormones (glucocorticoids in this case, hence the name), hangs around in the cytoplasm but translocates to the nucleus when bound, forms homodimers and recognises inverted repeat recognition sequences. Now I'm going to expand on that a bit more!
In the cytoplasm, the glucocorticoid receptor usually associates with heat-shock proteins. When bound, the glucocorticoid receptor can dissociate, which allows it to dimerise and translocate to the nucleus. Once in the nucleus, the glucocorticoid receptor dimers can have effects in three ways. Firstly, the homodimers can bind to GRE (glucocorticoid response element), which activates the transcription of its genes. Secondly, the homodimers can bind to nGRE (negative glucocorticoid receptor), which, as the name suggests, has the opposite effect of GRE: it suppresses gene transcription. The third pathway is a little bit different. In this pathway, a monomer of the glucocorticoid receptor binds to a transcription factor called NFκB (that little K is the Greek letter "kappa" btw), which in turn is bound to NRE (which I presume is NFκB response element). This can increase or decrease transcription.
Peroxisome proliferator-activated receptor (PPAR)
PPAR is a class II nuclear receptor that comes in four isoforms: α, β, γ and δ. α, β and δ are all involved in fatty acid oxidation whereas γ is involved in adipogenesis, lipid metabolism and glucose homeostasis. Over a long time, PPARγ can also lead to insulin sensitisation. Their locations are also somewhat different: while β and δ are expressed ubiquitously, α is expressed predominantly in the liver, heart and brown adipose tissue, whereas γ is found in white and brown adipose tissue. The isoform that we're going to focus on is PPARγ.
So, back to the nitty-gritty how it works stuff. As you can guess from PPARγ being a class II receptor, it exists in the nucleus, forms heterodimers with the retinoid receptor (RXR) and binds to response elements with direct repeats. Like the insulin receptor, PPARγ doesn't dimerise after binding as it already exists in a dimerised state. PPARγ is also usually already bound to its response elements before ligand binding, but it's prevented from having an effect by a corepressor. When the ligand comes in, it kicks out the corepressor and brings in a coactivator. In this way, the ligand activates PPARγ and allows it to have its effects.
Now for some other random facts about PPARγ that I don't know where to put! Firstly, Ser112 (that's the 112th residue of the amino acid chain making up PPARγ that happens to be a serine) can be phosphorylated by different enzymes, including the cyclin-dependent kinases Cdk9 and Cdk7 and good ol' MAP Kinase of the Ras -> Raf -> Mek -> MAP Kinase pathway fame. The trick here is that when Ser112 is phosphorylated by Cdk9 and Cdk7, PPARγ's activity increases; when phosphorylated by MAP Kinase, its activity decreases. Why? Well, I have no idea, but it was kinda interesting.
Summarise the similarities and differences between four receptor superfamilies in terms of their location, activation and signal transduction
I've done this before, but I thought I'll put it in a nice neat table now that I know how to make them on Blogspot. (Hopefully Blogspot doesn't have the bright idea of putting in heaps of line breaks before the table after I hit "update." And yup, I'm updating because our ever-vigilant unit coordinator pointed out that I didn't actually address this point last time :) )
Recall classification of nuclear receptors and provide examples of ligands
Nuclear receptors can be classified into four categories, imaginatively named Class I, Class II, Class III and Class IV. I'm going to present them in a table, because y'know, madddd HTML skillz:
| Ligands | Location | Structure | Response Elements (RE)* | |
| Class I | Hormones | Cytoplasm- translocates to nucleus | Homodimers | Inverted repeats |
| Class II | Lipids | Nucleus | Heterodimer (one of the monomers is always RXR, the retinoid receptor) | Direct repeats |
| Class III | ** | ** | Homodimers | No inverted repeats |
| Class IV | ** | ** | Monomers or dimers | Only to one RE half-site |
*REs are recognition sites that the nuclear receptors recognise and bind to.
**Not in the lecture or the textbook, so I assume it's not important.
Describe the general structure of nuclear receptors
Nuclear receptors have four main domains, which I'm going to illustrate using my maddddd HTML skillz (since the excitement of finding out that I still remember how to use HTML still hasn't worn off...)
| N- | N-terminal domain | DNA Binding Domain | Hinge region | Ligand-binding domain | -C |
The N-terminal domain is the least conserved (i.e. most prone to variation in length and sequence). It contains Activation Function 1 (AF1), which binds to co-regulators independent of ligand binding (i.e. it doesn't care whether ligands are bound or not, it'll bind to co-regulators no matter what).
The DNA-binding domain is highly conserved. As the name suggests, it is where the nuclear receptor binds to DNA by recognising response elements (REs, or if you want to be more technical, HREs- hormone response elements). It has two zinc fingers formed by cysteine-rich loops. (For more information about zinc fingers, please see my earlier post on transcription factors.)
The hinge region is quite flexible, and may be responsible for dimerisation and conformation changes of the receptor.
The ligand-binding domain, as the name suggests, is where ligands bind. It is highly conserved and has 12 α-helices which form a hydrophobic "pocket." It also contains Activation Function 2 (AF2), which is kinda like AF1, but unlike AF1, AF2 actually cares if ligands are bound or not (i.e. it binds co-regulators in a ligand-dependent manner).
Describe the mechanism of activation of glucocorticoid receptor and peroxisome proliferator-activated receptor, and the effects on gene transcription
Glucocorticoid Receptor
Glucocorticoid receptors are class I receptors that is synthesised from the splicing together of 9 exons (the ninth of which has an α and β isoform, but I don't know how important it is for us to remember that). From the table above, you should have already figured out that the glucorticoid receptor's Class I categorisation means that it is activated by hormones (glucocorticoids in this case, hence the name), hangs around in the cytoplasm but translocates to the nucleus when bound, forms homodimers and recognises inverted repeat recognition sequences. Now I'm going to expand on that a bit more!
In the cytoplasm, the glucocorticoid receptor usually associates with heat-shock proteins. When bound, the glucocorticoid receptor can dissociate, which allows it to dimerise and translocate to the nucleus. Once in the nucleus, the glucocorticoid receptor dimers can have effects in three ways. Firstly, the homodimers can bind to GRE (glucocorticoid response element), which activates the transcription of its genes. Secondly, the homodimers can bind to nGRE (negative glucocorticoid receptor), which, as the name suggests, has the opposite effect of GRE: it suppresses gene transcription. The third pathway is a little bit different. In this pathway, a monomer of the glucocorticoid receptor binds to a transcription factor called NFκB (that little K is the Greek letter "kappa" btw), which in turn is bound to NRE (which I presume is NFκB response element). This can increase or decrease transcription.
Peroxisome proliferator-activated receptor (PPAR)
PPAR is a class II nuclear receptor that comes in four isoforms: α, β, γ and δ. α, β and δ are all involved in fatty acid oxidation whereas γ is involved in adipogenesis, lipid metabolism and glucose homeostasis. Over a long time, PPARγ can also lead to insulin sensitisation. Their locations are also somewhat different: while β and δ are expressed ubiquitously, α is expressed predominantly in the liver, heart and brown adipose tissue, whereas γ is found in white and brown adipose tissue. The isoform that we're going to focus on is PPARγ.
So, back to the nitty-gritty how it works stuff. As you can guess from PPARγ being a class II receptor, it exists in the nucleus, forms heterodimers with the retinoid receptor (RXR) and binds to response elements with direct repeats. Like the insulin receptor, PPARγ doesn't dimerise after binding as it already exists in a dimerised state. PPARγ is also usually already bound to its response elements before ligand binding, but it's prevented from having an effect by a corepressor. When the ligand comes in, it kicks out the corepressor and brings in a coactivator. In this way, the ligand activates PPARγ and allows it to have its effects.
Now for some other random facts about PPARγ that I don't know where to put! Firstly, Ser112 (that's the 112th residue of the amino acid chain making up PPARγ that happens to be a serine) can be phosphorylated by different enzymes, including the cyclin-dependent kinases Cdk9 and Cdk7 and good ol' MAP Kinase of the Ras -> Raf -> Mek -> MAP Kinase pathway fame. The trick here is that when Ser112 is phosphorylated by Cdk9 and Cdk7, PPARγ's activity increases; when phosphorylated by MAP Kinase, its activity decreases. Why? Well, I have no idea, but it was kinda interesting.
Summarise the similarities and differences between four receptor superfamilies in terms of their location, activation and signal transduction
I've done this before, but I thought I'll put it in a nice neat table now that I know how to make them on Blogspot. (Hopefully Blogspot doesn't have the bright idea of putting in heaps of line breaks before the table after I hit "update." And yup, I'm updating because our ever-vigilant unit coordinator pointed out that I didn't actually address this point last time :) )
| Receptor superfamily | Location | Activation (Ligands, time scale) | Signal transduction |
| Ion-channel coupled receptor | Cell membrane: N-extracellular, C-intracellular | Fast neurotransmitters, very fast (milliseconds) | Channel opens, allowing ion flow |
| G-protein coupled receptors | Cell membrane: N-extracellular, C-intracellular | Hormones or slow neurotransmitters, fast (seconds) | Activate G-proteins, which go on to activate other pathways |
| Enzyme-linked receptors | Cell membrane: N-extracellular, C-intracellular | Hormones, slow (minutes) | Act as enzymes or recruit enzymes such as JAK (Janus Kinase) |
| Nuclear receptors | Intracellular: cytoplasm and/or nucleus | Steroid hormones etc., very slow (hours) | Bind to response elements, induce transcription of genes |
Tuesday, May 10, 2016
Enzyme-Linked Receptors
Recognize the significance of protein phosphorylation in regulating protein function and therefore cellular processes
Protein phosphorylation is, as its name suggests, the addition of a phosphate group to a protein. This often serves to turn the protein "on" or "off" as the addition of the phosphate group adds bulk and a negative charge, which might help other proteins and substances within the cell to interact with it.
Recall major groups of enzyme-linked receptors and their ligands
Probably the most common groups of enzyme-linked receptors that you will encounter are kinases (enzymes that phosphorylate other proteins). They come in two types: receptor serine/threonine kinases and receptor tyrosine kinases (RTKs), all named after the residues that they add phosphate groups to. They are both bound by various growth factors. RTKs tend to be bound by other ligands such as insulin. (The only example given as a ligand for receptor serine/threonine kinases is transforming growth factor TGF-β, but I'm sure there are more than just that one.)
Three other types of receptors that you need to know are cytokine receptors, receptor guanylate cyclases and receptor tyrosine phosphatases. Cytokine receptors bind molecules known as cytokines, which according to http://www.news-medical.net/health/What-are-Cytokines.aspx aid in cell-to-cell communication and cell movement during immune responses. (From what I've heard, I'm going to hear about cytokines way too much during Immunology next semester.) As for the other two? Receptor guanylate cyclase is like the guanine version of adenylate cyclase: it converts GTP into cyclic GMP (cGMP). They are activated in response to natriuretic peptides, which help get rid of excess Na+ and H2O. Receptor tyrosine phosphatases remove phosphate from tyrosine residues and tend to exhibit constitutive activity, which means that they don't have to be bound to have an effect.
Describe the general structure and signalling pathways of kinase-linked receptors
Kinase-linked receptors are enzyme-linked receptors that come in two varieties: those with intrinsic kinase activity (i.e. they can add phosphate groups onto stuff all by themselves) and those without intrinsic kinase activity. Like many other receptors, they have three main parts: an extracellular N-terminal domain, a transmembrane domain and an intracellular C-terminal domain. Most enzyme-linked receptors (and kinase-linked receptors by extension) dimerise when bound, and this dimerisation is required for them to have an effect. Receptor tyrosine kinases (RTKs) are kinase-linked receptors with intrinsic kinase activity, whereas many cytokine receptors are kinase-linked receptors without this activity. I'm going to go through both of these cases separately.
Kinase-linked receptors with intrinsic kinase activity
When RTKs (which have intrinsic kinase activity) dimerise, the two monomers can phosphorylate each other in a process known as autophosphorylation or transphosphorylation. This allows recruitment of Grb2 (growth factor receptor bound protein 2). Grb2 then acts as GEF (guanine exchange factor) that removes GDP from the G-protein Ras, allowing GTP to bind. This activates Ras, which activates Raf, which is also known as MAPKK Kinase, or MAPKKK. This then phosphorylates Mek (MAPK Kinase/ MAPK), which phosphorylates MAP Kinase (MAPK), which then phosphorylates various transcription factors, which ultimately leads to gene transcription. (Raf, Mek and MAP Kinase are all serine/threonine kinases. Don't know how important that bit of info is, but it was in the slides.)
So, in short: Ligand binding -> Dimerisation -> Autophosphorylation -> Grb2 recruitment -> Ras activation -> Raf/MAPKKK -> Mek/MAPKK -> MAPK -> transcription factors -> ??? -> PROFIT!!!
Kinase-linked receptors without intrinsic kinase activity
Cytokine receptors lack intrinsic kinase activity, so they recruit another protein called Janus Kinase (JAK) to do their dirty work for them. JAK binds and phosphorylates another protein called STAT, which is a short way of saying Signal Transducer and Activator of Transcription. When STAT is phosphorylated, it can dimerise. STAT dimers can then stimulate gene transcription in the nucleus.
Describe the mechanisms of activation of receptor tyrosine kinases
In order to carry out their catalytic function, the active site of the receptor tyrosine kinases must be accessible. Sometimes, this active site must also be phosphorylated. Conversely, when a receptor tyrosine kinase is inactive, that is because the active site has been rendered inaccessible in some way.
The first mechanism in which this can happen is through the formation of an "activation loop," or "activation loop inhibition." The insulin receptor is an example of a receptor that does this. In its inactive form, a tyrosine residue projects into the active site as if it's meant to be the target of phosphorylation. This stabilises the configuration of the "active loop." When this activation loop inhibition is removed (by autophosphorylation of the tyrosine residues on each monomer), the receptor is activated.
The second mechanism is known as "juxtamembrane inhibition." In this mechanism, residues near the membrane ("juxta"- next to, "membrane"- membrane) interact with parts of the tyrosine kinase domain, including the active loop. Once again, when this inhibition is removed, the receptor is activated. KIT is an example of a receptor that uses this mechanism. (According to Google, KIT is another name for the mast/stem cell growth factor receptor.)
The third mechanism is known as "C-terminal tail inhibition." To my understanding, this is kinda similar to juxtamembrane inhibition, but this time it's residues near the C-terminal that are doing the trick, not the juxtamembrane residues. Tie2 is an example of a receptor that uses this mechanism. There seem to be a shitload of functions for this receptor, so I'm not even going to try and list all of them- hop over to http://www.phosphosite.org/proteinAction.action?id=1246 if you're really interested.
The fourth and final mechanism is allosteric activation. This mechanism requires two tyrosine kinases to work together: one as the activator and the other as the receiver. As the names suggest, the activator activates the receiver. EGF (epithelial growth factor) uses allosteric activation.
Describe the structure of insulin receptor, the signal transduction and effects associated with the activation of insulin receptor
Unlike other enzyme-linked receptors that I've written about so far, the insulin receptor does not dimerise when bound. This is because it naturally exists as a homodimer. Each monomer has an α subunit which is located entirely extracellularly, and a β subunit which pokes into the cell. These two monomers are linked by disulfide bonds (I'll write up a bit more about insulin in a later post for BIOC2001). Therefore, the overall structure of insulin can be represented by (αβ)2.
Insulin exerts its effects in several different ways. Firstly, I'll have a look at the way in which it exerts its long-term effects. Insulin interacts with a protein called IRS, or insulin receptor substrate. This is an "adaptor" protein that interacts with Grb2, which interacts with Ras, Raf, Mek, MAP Kinase and finally with transcription factors. If you think that this looks familiar, you're right: as I mentioned before, this is a general pathway for many kinase-linked receptors with intrinsic kinase activity.
There are two pathways in which insulin exerts its short-term effects. Fortunately we don't have to know the details- just the names and what they do. Unfortunately, even the names are pretty long. The first pathway is the APS/c-CBL/CAP-mediated pathway, which creates targeting sites for GLUT4 storage vesicles (GSVs) on cell membrane. (Some quick definitions: targeting sites tell the vesicles where to go, and the vesicles here are carrying GLUT4 which is a glucose transporter.) The other pathway is the IRS/PI3K/Akt-mediated pathway, which causes the translocation and exocytosis of GSVs. This results in transporters being inserted into the membrane.
Phew! That was a long haul. There are still some things I'm not 100% sure on, such as the activation processes. But that's something I'll need to check up on another day.
Protein phosphorylation is, as its name suggests, the addition of a phosphate group to a protein. This often serves to turn the protein "on" or "off" as the addition of the phosphate group adds bulk and a negative charge, which might help other proteins and substances within the cell to interact with it.
Recall major groups of enzyme-linked receptors and their ligands
Probably the most common groups of enzyme-linked receptors that you will encounter are kinases (enzymes that phosphorylate other proteins). They come in two types: receptor serine/threonine kinases and receptor tyrosine kinases (RTKs), all named after the residues that they add phosphate groups to. They are both bound by various growth factors. RTKs tend to be bound by other ligands such as insulin. (The only example given as a ligand for receptor serine/threonine kinases is transforming growth factor TGF-β, but I'm sure there are more than just that one.)
Three other types of receptors that you need to know are cytokine receptors, receptor guanylate cyclases and receptor tyrosine phosphatases. Cytokine receptors bind molecules known as cytokines, which according to http://www.news-medical.net/health/What-are-Cytokines.aspx aid in cell-to-cell communication and cell movement during immune responses. (From what I've heard, I'm going to hear about cytokines way too much during Immunology next semester.) As for the other two? Receptor guanylate cyclase is like the guanine version of adenylate cyclase: it converts GTP into cyclic GMP (cGMP). They are activated in response to natriuretic peptides, which help get rid of excess Na+ and H2O. Receptor tyrosine phosphatases remove phosphate from tyrosine residues and tend to exhibit constitutive activity, which means that they don't have to be bound to have an effect.
Describe the general structure and signalling pathways of kinase-linked receptors
Kinase-linked receptors are enzyme-linked receptors that come in two varieties: those with intrinsic kinase activity (i.e. they can add phosphate groups onto stuff all by themselves) and those without intrinsic kinase activity. Like many other receptors, they have three main parts: an extracellular N-terminal domain, a transmembrane domain and an intracellular C-terminal domain. Most enzyme-linked receptors (and kinase-linked receptors by extension) dimerise when bound, and this dimerisation is required for them to have an effect. Receptor tyrosine kinases (RTKs) are kinase-linked receptors with intrinsic kinase activity, whereas many cytokine receptors are kinase-linked receptors without this activity. I'm going to go through both of these cases separately.
Kinase-linked receptors with intrinsic kinase activity
When RTKs (which have intrinsic kinase activity) dimerise, the two monomers can phosphorylate each other in a process known as autophosphorylation or transphosphorylation. This allows recruitment of Grb2 (growth factor receptor bound protein 2). Grb2 then acts as GEF (guanine exchange factor) that removes GDP from the G-protein Ras, allowing GTP to bind. This activates Ras, which activates Raf, which is also known as MAPKK Kinase, or MAPKKK. This then phosphorylates Mek (MAPK Kinase/ MAPK), which phosphorylates MAP Kinase (MAPK), which then phosphorylates various transcription factors, which ultimately leads to gene transcription. (Raf, Mek and MAP Kinase are all serine/threonine kinases. Don't know how important that bit of info is, but it was in the slides.)
So, in short: Ligand binding -> Dimerisation -> Autophosphorylation -> Grb2 recruitment -> Ras activation -> Raf/MAPKKK -> Mek/MAPKK -> MAPK -> transcription factors -> ??? -> PROFIT!!!
Kinase-linked receptors without intrinsic kinase activity
Cytokine receptors lack intrinsic kinase activity, so they recruit another protein called Janus Kinase (JAK) to do their dirty work for them. JAK binds and phosphorylates another protein called STAT, which is a short way of saying Signal Transducer and Activator of Transcription. When STAT is phosphorylated, it can dimerise. STAT dimers can then stimulate gene transcription in the nucleus.
Describe the mechanisms of activation of receptor tyrosine kinases
In order to carry out their catalytic function, the active site of the receptor tyrosine kinases must be accessible. Sometimes, this active site must also be phosphorylated. Conversely, when a receptor tyrosine kinase is inactive, that is because the active site has been rendered inaccessible in some way.
The first mechanism in which this can happen is through the formation of an "activation loop," or "activation loop inhibition." The insulin receptor is an example of a receptor that does this. In its inactive form, a tyrosine residue projects into the active site as if it's meant to be the target of phosphorylation. This stabilises the configuration of the "active loop." When this activation loop inhibition is removed (by autophosphorylation of the tyrosine residues on each monomer), the receptor is activated.
The second mechanism is known as "juxtamembrane inhibition." In this mechanism, residues near the membrane ("juxta"- next to, "membrane"- membrane) interact with parts of the tyrosine kinase domain, including the active loop. Once again, when this inhibition is removed, the receptor is activated. KIT is an example of a receptor that uses this mechanism. (According to Google, KIT is another name for the mast/stem cell growth factor receptor.)
The third mechanism is known as "C-terminal tail inhibition." To my understanding, this is kinda similar to juxtamembrane inhibition, but this time it's residues near the C-terminal that are doing the trick, not the juxtamembrane residues. Tie2 is an example of a receptor that uses this mechanism. There seem to be a shitload of functions for this receptor, so I'm not even going to try and list all of them- hop over to http://www.phosphosite.org/proteinAction.action?id=1246 if you're really interested.
The fourth and final mechanism is allosteric activation. This mechanism requires two tyrosine kinases to work together: one as the activator and the other as the receiver. As the names suggest, the activator activates the receiver. EGF (epithelial growth factor) uses allosteric activation.
Describe the structure of insulin receptor, the signal transduction and effects associated with the activation of insulin receptor
Unlike other enzyme-linked receptors that I've written about so far, the insulin receptor does not dimerise when bound. This is because it naturally exists as a homodimer. Each monomer has an α subunit which is located entirely extracellularly, and a β subunit which pokes into the cell. These two monomers are linked by disulfide bonds (I'll write up a bit more about insulin in a later post for BIOC2001). Therefore, the overall structure of insulin can be represented by (αβ)2.
Insulin exerts its effects in several different ways. Firstly, I'll have a look at the way in which it exerts its long-term effects. Insulin interacts with a protein called IRS, or insulin receptor substrate. This is an "adaptor" protein that interacts with Grb2, which interacts with Ras, Raf, Mek, MAP Kinase and finally with transcription factors. If you think that this looks familiar, you're right: as I mentioned before, this is a general pathway for many kinase-linked receptors with intrinsic kinase activity.
There are two pathways in which insulin exerts its short-term effects. Fortunately we don't have to know the details- just the names and what they do. Unfortunately, even the names are pretty long. The first pathway is the APS/c-CBL/CAP-mediated pathway, which creates targeting sites for GLUT4 storage vesicles (GSVs) on cell membrane. (Some quick definitions: targeting sites tell the vesicles where to go, and the vesicles here are carrying GLUT4 which is a glucose transporter.) The other pathway is the IRS/PI3K/Akt-mediated pathway, which causes the translocation and exocytosis of GSVs. This results in transporters being inserted into the membrane.
Phew! That was a long haul. There are still some things I'm not 100% sure on, such as the activation processes. But that's something I'll need to check up on another day.
Monday, May 9, 2016
G-Protein Coupled Receptors
Describe the structure of G protein-coupled receptors
As I've mentioned in an earlier post, G-protein coupled receptors have an extracellular N-terminal domain and an intracellular C-terminal domain. They have 7 transmembrane helices connecting the two, with three extracellular loops and three intracellular loops.
Describe the role of G protein-coupled receptors in physiological processes
Discuss the common themes of G protein-coupled receptor signal transduction
Describe the mechanisms by which G protein-coupled receptors cause a change in cell function
I feel like I've already pretty much done this in my previous post, but here's a refresher: when G-protein coupled receptors are bound, they remove GDP from a G-protein, allowing it to be replaced with GTP. This causes the G-protein to break into alpha and beta/gamma parts, which go on to activate other stuff.
Of importance might be the different types of G-proteins. There are three different types of alpha subunits. These are Gαs, Gαi and Gαq. Gαs work by activating adenylate cyclase, which is the enzyme that converts ATP into cyclic AMP (cAMP). This is a second messenger molecule that activates cAMP-dependent protein kinase A, which leads to effects such as bronchodilation in the airways. (β-adrenoceptors use this pathway.) Gαi has the opposite effect: it inhibits adenylate cyclase. (α2-adrenoreceptors use this pathway.) Gαq is kinda special, in that it actually activates two second messengers. It cleaves phosphatidylinositol 4,5-bisphosphate to form inositol 1,4,5-trisphosphate and diacylglycerol. Inositol 1,4,5-trisphosphate goes on to release Ca2+ from the endoplasmic reticulum, which goes on to form a Ca2+-calmodulin complex, while diacylglycerol activates protein kinase C. Both of these pathways go on to produce an overall effect, such as bronchoconstriction in the airways. (α1-adrenoreceptors and muscarinic ACh receptors use this pathway. It is the latter type that results in bronchoconstriction.)
Aaaaand believe it or not, I think I'm done on this topic! That was a short post... I guess the next two will make up for it though! D:
As I've mentioned in an earlier post, G-protein coupled receptors have an extracellular N-terminal domain and an intracellular C-terminal domain. They have 7 transmembrane helices connecting the two, with three extracellular loops and three intracellular loops.
Describe the role of G protein-coupled receptors in physiological processes
Discuss the common themes of G protein-coupled receptor signal transduction
Describe the mechanisms by which G protein-coupled receptors cause a change in cell function
I feel like I've already pretty much done this in my previous post, but here's a refresher: when G-protein coupled receptors are bound, they remove GDP from a G-protein, allowing it to be replaced with GTP. This causes the G-protein to break into alpha and beta/gamma parts, which go on to activate other stuff.
Of importance might be the different types of G-proteins. There are three different types of alpha subunits. These are Gαs, Gαi and Gαq. Gαs work by activating adenylate cyclase, which is the enzyme that converts ATP into cyclic AMP (cAMP). This is a second messenger molecule that activates cAMP-dependent protein kinase A, which leads to effects such as bronchodilation in the airways. (β-adrenoceptors use this pathway.) Gαi has the opposite effect: it inhibits adenylate cyclase. (α2-adrenoreceptors use this pathway.) Gαq is kinda special, in that it actually activates two second messengers. It cleaves phosphatidylinositol 4,5-bisphosphate to form inositol 1,4,5-trisphosphate and diacylglycerol. Inositol 1,4,5-trisphosphate goes on to release Ca2+ from the endoplasmic reticulum, which goes on to form a Ca2+-calmodulin complex, while diacylglycerol activates protein kinase C. Both of these pathways go on to produce an overall effect, such as bronchoconstriction in the airways. (α1-adrenoreceptors and muscarinic ACh receptors use this pathway. It is the latter type that results in bronchoconstriction.)
Aaaaand believe it or not, I think I'm done on this topic! That was a short post... I guess the next two will make up for it though! D:
Tuesday, April 26, 2016
Ligand gated ion channels
In my last pharmacology post, I spoke about the four main classes of receptors: ion channels, G-protein coupled, enzyme-linked and nuclear/DNA-linked. In these next few posts, I'll be expanding on each of these four categories.
Provide examples of ligand-gated ion channels.
My previous post already provided one: nicotinic ACh receptors, which are also Na+ channels. (These also got a mention in my post on the autonomic nervous system. If you're wondering about muscarinic receptors, they're actually G-protein coupled.)
A second example of a ligand-gated ion channel is GABAA receptors. (GABAC receptors do the same thing, but we're just going to ignore them for now. Oh, and there's also GABAB receptors, but they're G-protein coupled.) GABA receptors are Cl- channels which hyperpolarise the cell when open.
Both of the above receptors (nicotinic and GABAA) have similar structures. I'll discuss the structures in a bit...
Be able to describe the basic structure and transduction mechanisms of ligand-gated ion channels.
Ligand-gated ion channels usually have around 4-5 subunits that make up the "channel." These subunits are imaginatively called alpha, beta, gamma, delta etc. Each subunit has both the N- and C-terminals located extracellularly, with four alpha helices crossing the membrane. In the case of the two previous receptors mentioned (nicotinic and GABAA), there are five subunits present: alpha, alpha, beta, gamma and delta. Each alpha subunit must have a molecule of agonist bound for the subunits to turn slightly, opening the channel and allowing ions to pass through.
The movement of ions through the membrane may make the inside of the cell more positive (depolarisation) or more negative (hyperpolarisation). The cell is usually negatively charged compared to its environment, with a "resting potential" of around -70mV. In some cells (particularly neurons and skeletal muscle), once the cell reaches around -50mV, an "action potential" (i.e. a rapid shift from negative to positive charge inside the cell) is initiated. This results in neurons firing, skeletal muscle contraction, and so on.
Another interesting point to mention is that the lining of the channel (i.e. the amino acid groups) might influence which ions pass through a channel once it opens. For positively-charged ions such as Na+, the lining will have more negatively-charged amino acids such as aspartic acid and glutamic acid, whereas the opposite will be true for negatively-charged ions.
Be able to explain how interaction of a ligand with an ion channel is coupled to a biological effect within the cell.
I feel like I've already explained this in my last few paragraphs, so I'm going to expand on this by talking about antagonists and long-acting agonists.
As mentioned in my previous post, pancuronium is a competitive antagonist of the nicotinic ACh receptor (i.e. it binds to the receptor without having an effect, also preventing ACh from binding and having an effect). When the ACh receptor is blocked, the Na+ channel is prevented from opening. This prevents depolarisation of the cell, which stops skeletal muscle from having an action potential and contracting. Hence, pancuronium is used as a local anaesthetic as it stops muscles from twitching during surgery.
Another kind of drug is a long-acting agonist. An example of a long-acting agonist is suxamethonium. Long-acting agonists also bind to the receptor, but they keep the channel open. This prevents the cell from depolarising over and over again. Since such repetitive depolarisations are necessary for sustained contraction, suxamethonium is good for short-term muscle relaxation in short surgical procedures.
Provide examples of ligand-gated ion channels.
My previous post already provided one: nicotinic ACh receptors, which are also Na+ channels. (These also got a mention in my post on the autonomic nervous system. If you're wondering about muscarinic receptors, they're actually G-protein coupled.)
A second example of a ligand-gated ion channel is GABAA receptors. (GABAC receptors do the same thing, but we're just going to ignore them for now. Oh, and there's also GABAB receptors, but they're G-protein coupled.) GABA receptors are Cl- channels which hyperpolarise the cell when open.
Both of the above receptors (nicotinic and GABAA) have similar structures. I'll discuss the structures in a bit...
Be able to describe the basic structure and transduction mechanisms of ligand-gated ion channels.
Ligand-gated ion channels usually have around 4-5 subunits that make up the "channel." These subunits are imaginatively called alpha, beta, gamma, delta etc. Each subunit has both the N- and C-terminals located extracellularly, with four alpha helices crossing the membrane. In the case of the two previous receptors mentioned (nicotinic and GABAA), there are five subunits present: alpha, alpha, beta, gamma and delta. Each alpha subunit must have a molecule of agonist bound for the subunits to turn slightly, opening the channel and allowing ions to pass through.
The movement of ions through the membrane may make the inside of the cell more positive (depolarisation) or more negative (hyperpolarisation). The cell is usually negatively charged compared to its environment, with a "resting potential" of around -70mV. In some cells (particularly neurons and skeletal muscle), once the cell reaches around -50mV, an "action potential" (i.e. a rapid shift from negative to positive charge inside the cell) is initiated. This results in neurons firing, skeletal muscle contraction, and so on.
Another interesting point to mention is that the lining of the channel (i.e. the amino acid groups) might influence which ions pass through a channel once it opens. For positively-charged ions such as Na+, the lining will have more negatively-charged amino acids such as aspartic acid and glutamic acid, whereas the opposite will be true for negatively-charged ions.
Be able to explain how interaction of a ligand with an ion channel is coupled to a biological effect within the cell.
I feel like I've already explained this in my last few paragraphs, so I'm going to expand on this by talking about antagonists and long-acting agonists.
As mentioned in my previous post, pancuronium is a competitive antagonist of the nicotinic ACh receptor (i.e. it binds to the receptor without having an effect, also preventing ACh from binding and having an effect). When the ACh receptor is blocked, the Na+ channel is prevented from opening. This prevents depolarisation of the cell, which stops skeletal muscle from having an action potential and contracting. Hence, pancuronium is used as a local anaesthetic as it stops muscles from twitching during surgery.
Another kind of drug is a long-acting agonist. An example of a long-acting agonist is suxamethonium. Long-acting agonists also bind to the receptor, but they keep the channel open. This prevents the cell from depolarising over and over again. Since such repetitive depolarisations are necessary for sustained contraction, suxamethonium is good for short-term muscle relaxation in short surgical procedures.
Thursday, April 14, 2016
Receptors and Other Drug Targets
Last post before the test! Joy :(
For each of the 4 superfamilies of receptor, be able to describe major characteristic features including
– mechanism of signal transduction
– receptor location
– effector protein(s)
– time scale of action
Provide at least one detailed example of a drug that acts via each of the 4 receptor superfamilies
Ion-channel receptors
Ion-channel receptors, as their name suggests, are receptors that are ion channels. When activated by an agonist (usually a fast neurotransmitter), they open, allowing ions to flow through them. (Ions generally do not cross the cell membrane as they are charged particles.) They are located in the cell membrane, with both their C and N-termini located extracellularly. As ion flow occurs rapidly, their mechanism of action likewise occurs rapidly.
One example of a drug that acts on ion-channel receptors is Pancuronium. It antagonises nicotinic ACh receptors, which are also Na+ channels. Since it is an antagonist, it prevents the Na+ channels from opening and allowing Na+ from crossing the cell membrane. This stops neurons from producing their action potentials, and thus results in local anaesthesia.
G-protein coupled receptors
G-protein coupled receptors have their effects by interacting with G-proteins, which in turn react with other second messengers in the cell. I've spoken about G-proteins before, but just a quick recap: G-proteins are proteins that bind GTP (guanosine triphosphate). When this GTP is hydrolysed, only GDP (guanosine diphosphate) remains. G-protein coupled receptors act as GEFs (guanosine exchange factors), which, when bound by an agonist, get rid of the GDP on the G-protein so that it can be replaced with a fresh GTP molecule, thus activating the G-protein. G-proteins have three subunits (alpha, beta and gamma) which can interact with other effector proteins that release second messengers (for example Gαs activates adenylate cyclase, which produces cAMP).
G-protein coupled receptors are also located in the cell membrane. However, while their N-terminus is also located extracellularly, their C-terminus is located intracellularly. The intracellular region reacts with the G-protein.
Most G-protein coupled receptors respond to hormones or slow neurotransmitters. As more steps have to take place for them to have their effect (they have to activate the G-protein, which in turn has to react with other stuff), their method of action is relatively slow compared to ion-channel receptors. However, in the whole scheme of things, they are considered to be fast-acting (within seconds).
Now for an example! Salbutamol is a beta 2-adrenoceptor agonist that relieves bronchospasm in asthma. Beta 2-adrenoceptors are G-coupled receptors that, when activated, release a stimulatory G protein (Gs) which activates adenylate cyclase. cAMP is then produced, resulting in relaxation of the airways.
Enzyme-linked receptors
Enzyme-linked receptors pretty much are enzymes. Like G-protein coupled receptors, they are located in the cell membrane, with their N-terminus extracellular and their C-terminus intracellular. Usually the N-terminus is where the signalling molecule binds, whereas the C-terminus is the enzyme part. They usually respond to hormones for growth and differentiation, and as such their effects are slower, normally in the time scale of minutes.
An example of a molecule that binds to an enzyme-linked receptor is insulin. Insulin binds to insulin receptors, which act as tyrosine kinases. The overall effect of these kinases is to translocate the glucose transporter GLUT4 to the cell membrane, so that glucose can enter the cell where it is needed for metabolism.
DNA-linked receptors
DNA-linked receptors act directly on DNA (again, as their name suggests... methinks these receptors won't be difficult to remember). They are the only receptors that are located intracellularly. DNA-linked receptors normally respond to hormones, particularly steroids. Their mechanism of action can take hours as genes have to be transcribed and proteins have to be produced.
An example here is glucocorticoid drugs such as cortisone. They are anti-inflammatory agents that have their effects by binding to DNA-linked receptors, which in turn bind to the DNA. (As for which genes they transcribe... I guess I'll have to find that out.)
Explain, with examples, how ion channels, enzymes and transporters are important drug targets.
I feel like I've already done this by talking about the receptors above, but they did provide some more examples in the lecture so let's go over those.
Ion channels
Some of the drugs that react with ion channels include blockers and modulators. Blockers physically plug the channel, preventing stuff from passing through. I've already given an example here with local anaesthetics blocking nicotinic ACh receptors (which are also Na+ channels). Modulators bind to other accessory sites on the channels, modulating their activity. An example here is benzodiazepines (e.g. Valium) which are sometimes prescribed for anxiety. They enhance the opening of GABA-activated Cl- channels.
Enzymes
Once again, I feel like I've pretty much covered this in the enzyme-linked receptor section, so I'm just going to provide a few more interesting tidbits instead. Interesting tidbit number 1 involves substrate analogues, which are basically drugs that act as competitive inhibitors. Sometimes the enzyme might actually break them down, but an abnormal metabolite is produced. For example, fluorouracil, an anti-cancer drug, replaces uracil. It cannot be broken down to thymidylate (a component of DNA), so DNA synthesis is inhibited.
Transporters
Sometimes transporters are also targets for drugs. For example, SSRIs (selective serotonin reuptake inhibitors) selectively prevent serotonin from being transported back into a neuron, allowing it to hang around in the synapse for longer and continue to stimulate the postsynaptic neuron. False substrates can be an issue here as well- amphetamines can hijack the noradrenaline transporter and replace or release noradrenaline and serotonin.
For each of the 4 superfamilies of receptor, be able to describe major characteristic features including
– mechanism of signal transduction
– receptor location
– effector protein(s)
– time scale of action
Provide at least one detailed example of a drug that acts via each of the 4 receptor superfamilies
Ion-channel receptors
Ion-channel receptors, as their name suggests, are receptors that are ion channels. When activated by an agonist (usually a fast neurotransmitter), they open, allowing ions to flow through them. (Ions generally do not cross the cell membrane as they are charged particles.) They are located in the cell membrane, with both their C and N-termini located extracellularly. As ion flow occurs rapidly, their mechanism of action likewise occurs rapidly.
One example of a drug that acts on ion-channel receptors is Pancuronium. It antagonises nicotinic ACh receptors, which are also Na+ channels. Since it is an antagonist, it prevents the Na+ channels from opening and allowing Na+ from crossing the cell membrane. This stops neurons from producing their action potentials, and thus results in local anaesthesia.
G-protein coupled receptors
G-protein coupled receptors have their effects by interacting with G-proteins, which in turn react with other second messengers in the cell. I've spoken about G-proteins before, but just a quick recap: G-proteins are proteins that bind GTP (guanosine triphosphate). When this GTP is hydrolysed, only GDP (guanosine diphosphate) remains. G-protein coupled receptors act as GEFs (guanosine exchange factors), which, when bound by an agonist, get rid of the GDP on the G-protein so that it can be replaced with a fresh GTP molecule, thus activating the G-protein. G-proteins have three subunits (alpha, beta and gamma) which can interact with other effector proteins that release second messengers (for example Gαs activates adenylate cyclase, which produces cAMP).
G-protein coupled receptors are also located in the cell membrane. However, while their N-terminus is also located extracellularly, their C-terminus is located intracellularly. The intracellular region reacts with the G-protein.
Most G-protein coupled receptors respond to hormones or slow neurotransmitters. As more steps have to take place for them to have their effect (they have to activate the G-protein, which in turn has to react with other stuff), their method of action is relatively slow compared to ion-channel receptors. However, in the whole scheme of things, they are considered to be fast-acting (within seconds).
Now for an example! Salbutamol is a beta 2-adrenoceptor agonist that relieves bronchospasm in asthma. Beta 2-adrenoceptors are G-coupled receptors that, when activated, release a stimulatory G protein (Gs) which activates adenylate cyclase. cAMP is then produced, resulting in relaxation of the airways.
Enzyme-linked receptors
Enzyme-linked receptors pretty much are enzymes. Like G-protein coupled receptors, they are located in the cell membrane, with their N-terminus extracellular and their C-terminus intracellular. Usually the N-terminus is where the signalling molecule binds, whereas the C-terminus is the enzyme part. They usually respond to hormones for growth and differentiation, and as such their effects are slower, normally in the time scale of minutes.
An example of a molecule that binds to an enzyme-linked receptor is insulin. Insulin binds to insulin receptors, which act as tyrosine kinases. The overall effect of these kinases is to translocate the glucose transporter GLUT4 to the cell membrane, so that glucose can enter the cell where it is needed for metabolism.
DNA-linked receptors
DNA-linked receptors act directly on DNA (again, as their name suggests... methinks these receptors won't be difficult to remember). They are the only receptors that are located intracellularly. DNA-linked receptors normally respond to hormones, particularly steroids. Their mechanism of action can take hours as genes have to be transcribed and proteins have to be produced.
An example here is glucocorticoid drugs such as cortisone. They are anti-inflammatory agents that have their effects by binding to DNA-linked receptors, which in turn bind to the DNA. (As for which genes they transcribe... I guess I'll have to find that out.)
Explain, with examples, how ion channels, enzymes and transporters are important drug targets.
I feel like I've already done this by talking about the receptors above, but they did provide some more examples in the lecture so let's go over those.
Ion channels
Some of the drugs that react with ion channels include blockers and modulators. Blockers physically plug the channel, preventing stuff from passing through. I've already given an example here with local anaesthetics blocking nicotinic ACh receptors (which are also Na+ channels). Modulators bind to other accessory sites on the channels, modulating their activity. An example here is benzodiazepines (e.g. Valium) which are sometimes prescribed for anxiety. They enhance the opening of GABA-activated Cl- channels.
Enzymes
Once again, I feel like I've pretty much covered this in the enzyme-linked receptor section, so I'm just going to provide a few more interesting tidbits instead. Interesting tidbit number 1 involves substrate analogues, which are basically drugs that act as competitive inhibitors. Sometimes the enzyme might actually break them down, but an abnormal metabolite is produced. For example, fluorouracil, an anti-cancer drug, replaces uracil. It cannot be broken down to thymidylate (a component of DNA), so DNA synthesis is inhibited.
Transporters
Sometimes transporters are also targets for drugs. For example, SSRIs (selective serotonin reuptake inhibitors) selectively prevent serotonin from being transported back into a neuron, allowing it to hang around in the synapse for longer and continue to stimulate the postsynaptic neuron. False substrates can be an issue here as well- amphetamines can hijack the noradrenaline transporter and replace or release noradrenaline and serotonin.
Friday, April 8, 2016
Introduction to Drug Action
Yet another pharmacology post! I've decided that I really don't get the order in which these lectures have been in... but oh well.
Be able to answer the question “What is a receptor?”
A receptor is basically something that a drug or another signalling molecule can bind to. When it is bound (or, in some cases, when it isn't bound), it induces some kind of effect. See my earlier post on Biochemical Messengers for more information.
Be able to explain concepts such as agonism, partial agonism and antagonism in terms of drug affinity and efficacy
Firstly I'll explain what "affinity" and "efficacy" mean. "Affinity" refers to a drug's ability to bind to a target. "Efficacy" refers to the degree of receptor activation- a high efficacy means a high activation of receptors.
An "agonist" is a drug that binds to a receptor to induce an effect. Agonists have affinity and efficacy. An antagonist is the opposite: it also binds to a receptor, but it does not cause an effect: instead, it blocks other agonists from having a potential effect. (Rude.) They have affinity, as they do bind to the receptors, but they have no efficacy. A partial agonist is somewhere in between: it causes some effect, but not as much as a full agonist. They also have affinity, but low efficacy.
Be able to explain what is meant by drug selectivity
Drug selectivity, as the term suggests, refers to a drug binding to certain targets but not to others. This is often to do with the structure of the drug. See my previous post on structure-activity relationships for more details.
Be able to describe the relationship between agonist concentration and receptor occupancy or response
Generally, as the concentration of an agonist increases, the number of receptors that are occupied by said agonist also increases. Because of this, the response also increases. The concentration at which half the maximal response is achieved is known as EC50. If a drug has a low EC50, it is said to be more potent; if a drug has a high EC50, it is not as potent. Potency depends largely on affinity and efficacy.
Be able to explain the differences between reversible and irreversible receptor antagonism
Antagonists come in two main types: those that bind temporarily and dissociate (known as "reversible antagonists") and those that get stuck there forever (also known as "irreversible antagonists").
When a reversible antagonist binds, the concentration-effect curve is "shifted" to the right. This means that EC50 is higher, which in turn means that more agonist is required in order to "compete successfully" for receptors. The size of this "curve shift" depends on several factors such as the affinity and concentration of the antagonist.
When an irreversible antagonist binds, there are fewer spaces in which an agonist can occupy. The overall effect is just like having fewer receptors: the maximum effect is decreased, and the slope of the curve is also reduced. Once again, this "curve shift" depends on factors such as concentration.
Be able to answer the question “What is a receptor?”
A receptor is basically something that a drug or another signalling molecule can bind to. When it is bound (or, in some cases, when it isn't bound), it induces some kind of effect. See my earlier post on Biochemical Messengers for more information.
Be able to explain concepts such as agonism, partial agonism and antagonism in terms of drug affinity and efficacy
Firstly I'll explain what "affinity" and "efficacy" mean. "Affinity" refers to a drug's ability to bind to a target. "Efficacy" refers to the degree of receptor activation- a high efficacy means a high activation of receptors.
An "agonist" is a drug that binds to a receptor to induce an effect. Agonists have affinity and efficacy. An antagonist is the opposite: it also binds to a receptor, but it does not cause an effect: instead, it blocks other agonists from having a potential effect. (Rude.) They have affinity, as they do bind to the receptors, but they have no efficacy. A partial agonist is somewhere in between: it causes some effect, but not as much as a full agonist. They also have affinity, but low efficacy.
Be able to explain what is meant by drug selectivity
Drug selectivity, as the term suggests, refers to a drug binding to certain targets but not to others. This is often to do with the structure of the drug. See my previous post on structure-activity relationships for more details.
Be able to describe the relationship between agonist concentration and receptor occupancy or response
Generally, as the concentration of an agonist increases, the number of receptors that are occupied by said agonist also increases. Because of this, the response also increases. The concentration at which half the maximal response is achieved is known as EC50. If a drug has a low EC50, it is said to be more potent; if a drug has a high EC50, it is not as potent. Potency depends largely on affinity and efficacy.
Be able to explain the differences between reversible and irreversible receptor antagonism
Antagonists come in two main types: those that bind temporarily and dissociate (known as "reversible antagonists") and those that get stuck there forever (also known as "irreversible antagonists").
When a reversible antagonist binds, the concentration-effect curve is "shifted" to the right. This means that EC50 is higher, which in turn means that more agonist is required in order to "compete successfully" for receptors. The size of this "curve shift" depends on several factors such as the affinity and concentration of the antagonist.
When an irreversible antagonist binds, there are fewer spaces in which an agonist can occupy. The overall effect is just like having fewer receptors: the maximum effect is decreased, and the slope of the curve is also reduced. Once again, this "curve shift" depends on factors such as concentration.
Thursday, April 7, 2016
Stereochemistry
This *should* be a real quickie, given that it draws on concepts I've talked about before (namely chirality and structure-activity relationships). (Strangely, I don't have a post entirely on chirality- probably a good post would involve diagrams which I'm too lazy to draw- but that post I just linked to, as well as this post on carbohydrates, cover a fair bit.)
1) Define the terms “stereoisomer” & “chiral centre"
Ehhhh can't be bothered typing, read this post on carbohydrates instead.
2) Demonstrate a basic appreciation of the drawing conventions used to denote the presence of a chiral carbon within drug structures
In diagrams, you might have seen dashed lines and/or wedges between atoms. Dashed lines indicate that the atom in question is going "into" the page (i.e. away from you) while wedges indicate the opposite- that the atom in question is coming "out of" the page. You can use these to help you work out how the atoms are oriented, and from that you can work out which enantiomer they are. In racemic mixtures (i.e. mixtures with both enantiomers in equal quantities), a squiggly line might be drawn instead.
3) Show an appreciation of the pharmacodynamic implications of stereoisomerism, using the “3 contact point model” to explain such phenomena
Stereoisomers have different shapes, which means that they have different affinities for different targets. Can't be bothered drawing models or anything, unless anyone really wants me to.
4) Show an appreciation of the pharmacokinetic implications of stereoisomerism in drugs, especially during drug metabolism.
Once again, different stereoisomers have different affinities for different targets. Not only does this affect how well a drug binds to its target, but it may also affect absorption because certain stereoisomers may have greater affinities for certain transport proteins. (Note that chirality only affects "active" processes such as active transport- it doesn't affect a drug's ability to diffuse across the membrane.)
Another important point of note is that sometimes metabolism affects stereoisomerism. Metabolism can change a drug from one stereoisomer to another, abolish chirality or establish new sites of chirality. These all have implications for how a drug acts in the body.
5) Be able to define the term “chiral switching” and give examples of the use of this strategy.
"Chiral switching" is basically a company marketing a pure enantiomer of a drug, rather than the racemic mixture. For example, citalopram, an antidepressant drug, is a racemic mixture; escitalopram contains the S-enantiomer of citalopram only (hence escitalopram. Very funny, pharmacists). This is often done if one enantiomer is known to be much more effective than the other. Selling a drug containing only the effective stereoisomer means that only half the dose needs to be taken, while eliminating any negative effects that metabolism of the ineffective stereoisomer may have had. Going back to the escitalopram example, apparently escitalopram may be more effective than citalopram. (On an unrelated note, escitalopram is a massive pain in the rear end to go off. I'm currently tempted to stick a meme on here with my psychiatrist's face and the words "'Go off your escitalopram,' she said. 'It will be easy,' she said." But I'm nice, and I won't do that to her.)
A few quick definitions: if one enantiomer is better, then the better one is known as the eutomer while the worse one is the distomer (from the Greek "eu" meaning "good" and "dis" meaning "bad"). The ratio between the two is known as the eudismic ratio.
6) Show an appreciation of the toxicological implications of stereoisomerism
Of course, if stereoisomerism can have implications as to which targets drugs bind to, it can also have toxicological implications if the binding of one stereoisomer leads to negative effects. The no-brainer solution then is to use "chiral switching" to give patients only the stereoisomer that isn't toxic; however, this isn't always so simple as chirality may change within the body due to metabolism or otherwise.
One possible example of so-called "chiral toxicity" is ketamine. It's an anaesthetic, but it's also used as a recreational drug. It's thought that S-Ketamine has better anaesthetic activity and fewer side effects, while R-Ketamine is more likely to cause psychosis, agitation and amnesia.
1) Define the terms “stereoisomer” & “chiral centre"
Ehhhh can't be bothered typing, read this post on carbohydrates instead.
2) Demonstrate a basic appreciation of the drawing conventions used to denote the presence of a chiral carbon within drug structures
In diagrams, you might have seen dashed lines and/or wedges between atoms. Dashed lines indicate that the atom in question is going "into" the page (i.e. away from you) while wedges indicate the opposite- that the atom in question is coming "out of" the page. You can use these to help you work out how the atoms are oriented, and from that you can work out which enantiomer they are. In racemic mixtures (i.e. mixtures with both enantiomers in equal quantities), a squiggly line might be drawn instead.
3) Show an appreciation of the pharmacodynamic implications of stereoisomerism, using the “3 contact point model” to explain such phenomena
Stereoisomers have different shapes, which means that they have different affinities for different targets. Can't be bothered drawing models or anything, unless anyone really wants me to.
4) Show an appreciation of the pharmacokinetic implications of stereoisomerism in drugs, especially during drug metabolism.
Once again, different stereoisomers have different affinities for different targets. Not only does this affect how well a drug binds to its target, but it may also affect absorption because certain stereoisomers may have greater affinities for certain transport proteins. (Note that chirality only affects "active" processes such as active transport- it doesn't affect a drug's ability to diffuse across the membrane.)
Another important point of note is that sometimes metabolism affects stereoisomerism. Metabolism can change a drug from one stereoisomer to another, abolish chirality or establish new sites of chirality. These all have implications for how a drug acts in the body.
5) Be able to define the term “chiral switching” and give examples of the use of this strategy.
"Chiral switching" is basically a company marketing a pure enantiomer of a drug, rather than the racemic mixture. For example, citalopram, an antidepressant drug, is a racemic mixture; escitalopram contains the S-enantiomer of citalopram only (hence escitalopram. Very funny, pharmacists). This is often done if one enantiomer is known to be much more effective than the other. Selling a drug containing only the effective stereoisomer means that only half the dose needs to be taken, while eliminating any negative effects that metabolism of the ineffective stereoisomer may have had. Going back to the escitalopram example, apparently escitalopram may be more effective than citalopram. (On an unrelated note, escitalopram is a massive pain in the rear end to go off. I'm currently tempted to stick a meme on here with my psychiatrist's face and the words "'Go off your escitalopram,' she said. 'It will be easy,' she said." But I'm nice, and I won't do that to her.)
A few quick definitions: if one enantiomer is better, then the better one is known as the eutomer while the worse one is the distomer (from the Greek "eu" meaning "good" and "dis" meaning "bad"). The ratio between the two is known as the eudismic ratio.
6) Show an appreciation of the toxicological implications of stereoisomerism
Of course, if stereoisomerism can have implications as to which targets drugs bind to, it can also have toxicological implications if the binding of one stereoisomer leads to negative effects. The no-brainer solution then is to use "chiral switching" to give patients only the stereoisomer that isn't toxic; however, this isn't always so simple as chirality may change within the body due to metabolism or otherwise.
One possible example of so-called "chiral toxicity" is ketamine. It's an anaesthetic, but it's also used as a recreational drug. It's thought that S-Ketamine has better anaesthetic activity and fewer side effects, while R-Ketamine is more likely to cause psychosis, agitation and amnesia.
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