Catalytic Power and Biological Efficiency

Back to the love saga of enzymes and substrates! Unfortunately there probably won't be anywhere near as much steamy action in this instalment :(

Firstly, a reminder about some of the basics to do with free energy diagrams. (I've also covered this in a post that was actually written in year 11, but I felt like that diagram wasn't the best as it used different symbols to those used in these BIOC2001 lectures.)

Anyway, on one side you have your reactants, and on the other side you have your products. Joining the two is a curve thing. As the y-axis suggests, the higher up the reactants, products or transition state are, the higher the energy of those particular substances.

The difference between the free energy of the products and reactants, or ΔG, is the free energy change in the overall reaction. If this is negative, the reaction will proceed spontaneously. This does not mean that the reaction will proceed quickly. It could take 2 milliseconds or 2 googol years for all you know. All "the reaction will proceed spontaneously" means is that the reaction can take place without any help from anybody.

The main factor determining the speed of the reaction (out of the stuff on the diagram, at least), is the activation energy. From the diagram, the activation energy of the forward reaction is ΔG^‡_f (the diagram doesn't have the double dagger because I couldn't be bothered drawing one in) while the activation energy of the reverse reaction is ΔG^‡_r. The lower the activation energy, the higher the rate constant, and vice versa.

In fact, there's a nice fancy equation for this, because as you know, nerds would be nothing without their equations:

k_f = (k/h)Te^-ΔG‡_f/RT
where k and h are constants (Boltzmann's constant and Planck's constant, respectively), T is temperature (presumably in Kelvin?) and R is the gas constant. So basically, there's a shitload of constants. Also, same goes for the reverse reaction, but swap the fs for rs.

As you can see from the equation, k_f increases as T increases (since the whole thing is multiplied by T), but decreases as ΔG^‡_f increases (as e is raised to a negative power).

Now let's look at what happens when catalysts are involved! Catalysts are like matchmakers- they help the reactants get together and do wonderful stuff! Here is a diagram for a general catalysed reaction. The black curve shows the normal pathway, whereas the red line shows the catalysed pathway.

From the graph, you should be able to see that ΔG^‡_f and ΔG^‡_r have changed, but ΔG is still the same. Hence a catalyst cannot change whether or not a reaction proceeds spontaneously, but it can change the activation energies and hence the rate constant. It has these effects on both forward and reverse reactions.

Now let's go back to looking at how this applies to our good friends Enzymes and Substrates! Enzymes and substrates are a bit wacky, as even the most basic enzyme-catalysed reaction has at least two intermediates that the reaction has to go through. The first intermediate is ES, when the enzyme and substrate are together, and the second is EP, in which the pair are still together after the substrate has been transformed into a product. Here's the graph:

When calculating the activation energy, you need to use the highest of the three "transition state" peaks (or however many peaks there are if you're looking at a more complex reaction). From there, you can work out the effect on reaction rate and so forth.

Sometimes there may be even more intermediates in the reaction. For example, enolase catalyses the transformation of 2-phosphoglycerate (2-PG) into phosphoenolpyruvate (PEP). It does this by first converting 2-phosphoglycerate into an enolic intermediate. Hence there are even more transition state "peaks" in this reaction:

1. Binding of enzyme to 2-PG
2. Conversion of 2-PG into enolate (enolate is still bound to the enzyme)
3. Conversion of enolate into PEP (PEP is still bound to the enzyme)
4. Release of PEP from the enzyme

The highest peak (conversion of enolate into PEP in this case) is used to calculate overall activation energy for the reaction.

Now, you might have wondered why the activation energy is always positive. Or maybe you haven't wondered that, but I don't care, I'm telling you anyway. It all comes down to a simple equation:

ΔG^‡ = ΔH^‡ - TΔS^‡
where ΔG^‡ is the activation energy, ΔH^‡ is enthalpy (internal energy of a reaction), T is temperature and ΔS^‡ is entropy (which is kinda like the "degree of randomness" of a reaction, but I'm sure shitloads of physicists would slap me for such a simplistic explanation).

Anyway, what you have to know is this: ΔH^‡ is positive during a reaction as some bonds are broken and that leads to greater enthalpy and ΔS^‡ is negative as molecules are being positioned precisely for the reaction to occur so there is a decrease in random motion. As a positive subtracting a negative is positive, ΔG^‡ is always positive.

Arrhenius Plots

I'm sorry, I just felt like I'd gone way too long without breaking things up with a heading. So there's a heading. Hope you appreciate it.

Anyway, let's go back to that equation I mentioned earlier, the k_f = (k/h)Te^-ΔG‡_f/RT one. Yeah, that one. It looks a lot less scary when you simplify all those constants at the beginning into one letter, I promise!

k_f = Ae^-ΔG‡_f/RT

See?! I told you so! You can also tidy it up by making k_f just a generic k, and ΔG^‡_f into E_a (which I'm guessing is short for "energy of activation" or something):

k = Ae^-E_a/RT

Now, nerds like graphs. So let's turn this into a graph! Exponentials suck though... hmm... let's make this linear by using logs!

ln k = ln(Ae^-E_a/RT)
ln k = ln A + ln(e^-E_a/RT)
ln k = ln A - (E_a/RT)

Rearrange this very slightly and you get:

ln k = ln A + (1/T)(-E_a/R)

You can then use this to make a nice, neat linear graph of ln k against 1/T. The slope of this plot is (-E_a/R) and the intercept is ln A. Aren't linear equations great? (And to think that I used to hate them so much when I was in year 8 or 9...)

Let's see what we can do with our knowledge of this plot!

First we'll start with another equation, because equations are great! (Yup, I'm a nerd. I freely admit it.)

E_a = ΔH^‡ + RT

This can be rearranged to give ΔH^‡ = E_a - RT, which is useful if you already know what E_a is. (And you can work out E_a by finding out the slope of the Arrhenius plot, which is equal to (-E_a/R), and multiplying that by the negative of the gas constant.)

You can also get an expression for ΔG^‡ as well, though it's not quite as nice looking. Near the beginning, I mentioned the equation k_f = (k/h)Te^-ΔG‡_f/RT which can eventually be rearranged to give ΔG^‡ = -RT ln (hk_f/kT). (Yes, there's two ks in there. I kept the first one as k_f to separate them out. In the equation I gave, k_f is the rate constant whereas the other k is Boltzmann's constant. Why they couldn't have picked a different letter is beyond me. Some places, like the lecture slides, may italicise one of the ks to separate them out.)

Anyway, now you have ΔH^‡ and ΔG^‡, you can substitute those into the ΔG^‡ = ΔH^‡ - TΔS^‡ equation to find ΔS^‡. Because I'm sure you were dying to know that.

So overall there are a fair few things you can find out from the Arrhenius plot and a bit of manipulation: activation energy, change in enthalpy, change in free energy and change in entropy. Before you go too crazy though, only calculate these parameters at the temperatures that are on the Arrhenius plot. Calculating at temperatures before and after is extrapolating, which might not be appropriate. Oh, and just so you know, all of these parameters are for the transition state with the largest activation energy.

Catalytic Power

Yay, last bit! Catalytic power is an easy way to compare how good enzymes are. It's very easy to calculate- you simply divide the rate constant of the catalysed reaction by the rate constant of the uncatalysed reaction. For example, the hydrolysis of urea is pretty slow when uncatalysed, with a rate constant of 3*(10^(-10)) s^-1. When catalysed by jack bean urease, however, the rate constant is 3*(10^4) s^-1. This gives a catalytic power of (3*(10^4))/(3*(10^(-10))) = 3*(10^14).

Tubes

I feel like my ANHB2212 posts are dragging out too long, and I need to be more concise. Sorry about that! Will try to be more concise here...

Describe the pattern of layers in bodily tubes

Hmm, I know how I can get through this concisely! Here's a list of the layers, from outside in.

• Adventitia/serosa- Adventitias are anchoring layers that help anchor the tube down to something else. Serosas are serous membranes, as mentioned in my post about the coelomic cavity.
• Muscularis externa- This is the muscle layer, which is usually two muscle layers: an outer longitudinal layer and an inner circular layer.
• Submucosa- Loose connective tissue with blood vessels and nerves.
• Mucosa- Consists of the muscularis mucosa (a thin layer of muscle tissue), the lamina propria (which supports the epithelium) and the epithelium itself (derived from endoderm if it's the gut, or from mesoderm if it's a blood vessel).
• Lumen- The cavity in the middle of the blood vessel.

Some tubes have glands- these can be found in the mucosa, submucosa or external layers.

Understand how this pattern is adapted to suit different functions in the gut

I'm going to whip out a table here. For conciseness. Or something. I'll only write stuff in the cells if there's something interesting or unique to write about. (I omitted "adventitia/serosa" because I didn't have anything special to talk about for either of them.)

Section	Main function(s)	Muscularis externa	Submucosa	Mucosa
Oesophagus	Transfer food to stomach- peristalsis		Mucous glands	Mucous glands, stratified squamous epithelium to protect from abrasion
Stomach	Some storage of food, control of food progression	Sphincters- pyloric and cardiac sphincters. Additional oblique muscle layer to mix food.		Thick and glandular, protects stomach. Glands secrete acids that kill pathogens and alkaline mucus that protects from acid. Columnar epithelium for absorption.
Small intestine	Absorption of nutrients, chemical digestion	Duodenal glands secrete alkaline mucus via intestinal pits	Spiral "plicae circulares" to extend contact time, villi, microvilli. Simple columnar epithelium, highly vascularised. Ileum has lymphoid tissue ("Peyer's Patches" to protect against colon bacteria.
Large intestine	Reabsorption of water, propulsion to rectum	3 "taeniae coli" that bunch intestine into sacculations/haustra		Increasing number of mucous cells. Smooth (no villi). Columnar epithelium.

Understand how this pattern is adapted to suit other bodily tubes

Blood Vessels

Blood vessels have three main layers:

• Tunica adventitia- similar to adventitia. Made of loose connective tissue that anchors the blood vessel in place. Sometimes it contains vasa vasorum, or "blood vessels of the blood vessels," which are especially important in ensuring that large, thick vessels are supplied with blood.
• Tunica media- similar to mucsularis mucosa. This is the main structural layer, made up of smooth muscle, elastin and/or connective tissue.
• Tunica intima- similar to mucosa. Made of simple squamous endothelium and supporting connective tissue. Nourished by diffusion from the blood in the lumen.

Now let's look at arteries in particular:

• Elastic arteries (e.g. carotid artery)- have a thick internal elastic lamina to keep the blood pressure constant. They have a vasa vasorum but very little muscle. They may also have baroreceptors and/or chemoreceptors.
• Muscular arteries (e.g. splenic artery)- have a thick wall of muscle (75% of wall thickness is muscle). This allows them to control blood flow to organs.
• Arterioles- there's lots of them, so their collective large cross-sectional area reduces the blood pressure before the blood gets to the capillaries (more on this when I write up on physiology).

And veins, which tend to have larger lumens than arteries:

• Large veins (e.g. IVC): no valves, but some longitudinal smooth muscle
• Medium veins (e.g. femoral vein)- have valves, which are just flaps of connective tissue lined with endothelium
• Venules- the veins' answer to arterioles
• Sinuses (e.g. coronary sinus)- super dilated veins

And then there's the capillaries between them. They are made of a single layer of endothelial cells, which makes it easy for stuff to diffuse through. They also have pericytes that serve as little muscular sphincters for the capillaries. Capillaries differ in the size of the pores between cells. Fenestrated capillaries allow large molecules like hormones to pass through, whereas sinusoids have massive gaps that allow entire cells to pass through.

Respiratory Tubes

Trachea, bronchi and bronchioles all have adventitia to anchor them. The trachea and bronchi also have cartilage "horseshoes" and respiratory epithelium consisting of mucous glands and cilia. The bronchioles do not have cartilage, but they are elastic and have smooth muscle that can adjust airway size. Bronchioles do not have glands in their mucosa.

Urinary Tubes

Urinary tubes (ureters and urethra) are retroperitoneal and have an adventitia. The urethra has an additional inner longitudinal muscular layer as well as a skeletal muscle sphincter. Ureters and urethra all have mucous glands to protect against corrosive urine, as well as transitional epithelium that becomes stratified squamous near the external opening of the urethra.

Reproductive Tubes

The ductus deferens/ vas deferens has an adventitia and 3 muscular layers. Its mucosa is elastic and has stereocilia, which are cilia that don't "waft" things but instead absorb things.

The uterine/fallopian tubes have a serosa rather than an adventitia, but they also have 3 muscular layers. Their mucosa has cilia that help waft the egg along, as well as glands that secrete nourishing fluid.

...Okay, that post was still quite long. Damn.

Gastrointestinal Tract: Mesenteries, Blood and Nerve Supply

Following on from my post on the patterns of blood vessels, here's a more specific post about the gastrointestinal tract!

Describe the arterial supply of the GIT (gastrointestinal tract)

The gastrointestinal tract is supplied by three main arteries, as mentioned in my previous post about coelomic cavities. The coeliac artery supplies the foregut (which is the digestive tract up to where the pancreatic and hepatic ducts drain into the duodenum), the superior mesenteric artery supplies the midgut (which runs up to around 2/3 of the transverse colon of the large intestine) and the inferior mesenteric artery supplies the hindgut (which is the rest of the GI tract). From there, the arteries branch out a bit. Let's look at each of the arteries in turn, and their branches.

The coeliac artery pokes out from the aorta just under the crura of the diaphragm. It has three main branches, each of which have three branches. Some of those branches have three branches too:

1. Hepatic artery
1. Hepatic artery proper
1. Right hepatic artery
2. Left hepatic artery
3. Cystic artery (supplies gallbladder)
2. Right gastric artery
3. Gastroduodenal artery
   1. Pancreaticoduodenal artery
   2. Right gastro-epiploic/ gastro-omental artery
   3. Supraduodenal artery
2. Left gastric artery
1. Left gastric proper
2. Accessory hepatic
3. Oesophageal branches
3. Splenic artery
1. Splenic artery proper
2. Short gastric artery
3. Left gastro-epiploic/ gastro-omental artery

There are also some anastamoses here- the right and left gastric arteries join up, as do the right and left gastro-epiploic/ gastro-omental arteries.

The superior mesenteric artery emerges just above the duodenum and has five main branches:

1. Jejunal branches
2. Ileal branches
3. Ileocolic artery
4. Right colic artery
5. Middle colic artery

Essentially, the ileocolic artery supplies the area around the ileocecal (i.e. ileum - cecum) junction, the right colic artery supplies the ascending colon and the middle colic artery supplies the transverse colon. (That's just a very rough guide, it's not 100% neatly defined like that.)

The inferior mesenteric artery emerges further down the aorta and only has three main branches:

1. Left colic artery
2. Sigmoidal artery
3. Superior rectal artery

Describe pattern of organs and vessels that lie in the mesenteries of the abdomen

As I mentioned in my post about coelomic cavities, some organs are retroperitoneal (lie against the body wall and are covered by peritoneum) while others are intraperitoneal (in the middle of the abdominal cavity, anchored by mesenteries).

Arteries supplying intraperitoneal organs lie in mesenteries. Let's have a look at the mesenteries and what blood vessels lie in each one:

Mesentery	Location	Vessels
Greater omentum	Hangs over the intestines like an apron. ("Omentum" is Latin for "apron.")	Short gastric artery Right and left gastro-epiploic/ gastro-omental arteries
Lesser omentum	Between the liver and the lesser curvature of the stomach.	Hepatic artery Right and left gastric arteries
The mesentery	Anchors small intestine	Jejunal branches Ileal branches
Transverse mesocolon	Anchors transverse colon	Middle colic artery Marginal artery (connects the colic arteries)
Sigmoid mesocolon	Anchors sigmoid colon	Sigmoid arteries

Pretty much any organ that I haven't mentioned is retroperitoneal. An easy way to remember it is that pretty much every second organ is retroperitoneal: duodenum is retroperitoneal, small intestine is intraperitoneal, ascending colon is retroperitoneal, transverse colon is intraperitoneal, descending colon is retroperitoneal, sigmoid colon is intraperitoneal, rectum is retroperitoneal etc.

Understand the importance of the portal venous drainage of the GIT

As mentioned in my post about the patterns of blood vessels, a portal system goes from capillaries to veins to even more capillaries, rather than going straight back to the heart. Veins draining the GI tract go through the liver to go back to the heart.

The main veins draining the GI tract are the splenic, superior mesenteric and inferior mesenteric veins. The splenic and superior mesenteric drain straight into the hepatic vein, whereas the inferior mesenteric usually joins the splenic before draining into the hepatic vein. This allows the liver to process all the stuff digested by the GI tract before allowing it into the general circulation.

Understand the role of anastomoses in blood supply of the GIT

As I've mentioned in a previous post, anastamoses are important as they give alternative routes for blood to flow in case of blockages or whatever.

Aside from arterial anastomoses, there are plenty of places where venous anastamoses can form in the GI tract. These allow blood to bypass the liver if the liver is diseased or the blood vessels supplying it are occluded or whatever. These anastomoses are known as portocaval anastomoses, and usually form around the beginning and end of the GI tract. Here are some examples:

• Oesophageal blood can drain into the inferior vena cava via the azygos system
• Middle and inferior rectal veins drain into the inferior vena cava
• Blood from retroperitoneal organs can drain via body wall or renal veins

Understand the distribution and function of autonomic nerves in the abdomen

Nerves are important in controlling the release of stuff that helps us digest our food, and so on. I've already written about how the autonomic nervous system works- now let's have a look at how this applies to the GI tract!

The parasympathetic nerves that supply the GI tract include the vagus nerve and the pelvic splanchnic (splanchnic = visceral) nerves. The vagus nerve (which I've mentioned before) supplies the thoracic, foregut and midgut organs, by following the oesophagus into the abdomen. It also follows blood vessels to reach the target organs of the gut. The pelvic splanchnics, S234, supply the hindgut and pelvic organs. They emerge from the sacrum, join the inferior hypogastric plexus and run up to join the superior hypogastric plexus to join the inferior mesenteric artery. (As for how the parasympathetics do their job, that's a topic for a Physiology post.)

I've already spoken about the general structure of sympathetic nerves before, back when I wrote about embryology. Essentially, the preganglionic neuron goes out to the sympathetic ganglion via the white rami. In the sympathetic ganglion, it can synapse with postganglionic grey rami neurons, which innervate the body wall. However, the preganglionic neuron can also keep going out and synapse in preaortic (i.e. in front of the aorta) ganglia. These supply the viscera and include cardiac nerves, coeliac nerves, superior mesenteric nerves and so on. From here, the nerves follow the blood supply to reach target organs:

• T6-9 supplies the foregut via the coeliac plexus
• T9-11 supplies the midgut via the superior mesenteric plexus
• T11-L2 supplies the hindgut with the inferior mesenteric plexus
• T8-L1 supply the adrenals- coeliac plexus again
• T10-T12 supply the kidneys and ureters via the aorticorenal plexus
• T12-L2 supply the pelvic organs via the hypogastric plexus

Now a quick note on sensory nerves, which I don't think I've covered before! The autonomic nerves themselves are not sensory, but sensory nerves do run alongside the autonomic nerves. Hence visceral pain tends to arise from wherever the sympathetic nerve supplying that region came from, and are felt all over that entire dermatome. For example, if you have foregut pain, you might feel dull, achy pain all over T6-T9.

Somatic pain is a bit different- it's a sharp pain close to where the pain actually is.

Phrenic referred pain is a similar phenomenon to visceral pain, in which the pain is felt somewhere else. ("Referred pain" is basically pain that is felt somewhere other than the spot that's actually damaged.) Since the phrenic nerves arise from the neck and supply the diaphragm, pain around the diaphragm may be felt in the neck. As if working out what's wrong with you when you're sick had to be made even more difficult...!

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.)

Drug Name	Lecture(s)	Important Information
Acetylcholine	12, 13	Neurotransmitter mainly involved in pre-postganglionic cell junctions and in the parasympathetic nervous system. Activates nicotinic receptors (Na+ channels) Activates muscarinic receptors (G-protein-coupled) Muscarinic receptors activate Gαq, which cleaves phosphatidylinositol 4,5-bisphosphate into inositol 1,4,5-trisphosphate and diacylglycerol Inositol 1,4,5-trisphosphate stimulates release of Ca2+ and its binding to calmodulin; diacylglycerol activates protein kinase C. These pathways lead to bronchoconstriction
Acrolein	7	Found in cigarette smoke Very electrophilic and very toxic
Acyclovir	19	Purine (deoxyguanosine) analogue inhibitor of DNA synthesis Prodrug- must be activated by viral thymidine kinase and other cellular kinases Active against herpes viruses
Adrenaline	10	Naturally-occurring R-adrenaline interacts strongly with β-adrenoceptor Synthetic-only S-adrenaline interacts only weakly with β-adrenoceptor Activates β-adrenoceptors on cardiac cells to increase contractility
Aminoglycaside (class)	18	Bacterial protein synthesis inhibitor Examples: streptomycin, gentamicin Poor oral absorption due to positive charges, serious side effects including ototoxicity, hearing loss, nephrotoxicity Bind to decoding site on 30S ribosome, causing misreading of mRNA and insertion of the wrong amino acid
Amphetamine	12	CNS stimulant Uses noradrenaline transporter to enter nerve terminals Replaces noradrenaline and serotonin
Amphotericin	20	Polyene antifungal Poor oral absorption Renal toxicity with IV dosing Binds to ergosterol and creates pores through which essential ions can leak out
Artemisinin	1	340AD: Ge Hong used Sweet Annie to alleviate fever 1596: Li Shizhen used it to treat malarial fever 1970s: Artemisinin isolated 1980s/90s: Soluble derivatives synthesised 2015: Tu Youyou won a Nobel Prize for helping to isolate artemisinin Used to treat malaria
Aspirin	1, 9	Nonsteroidal anti-inflammatory drug (NSAID) ~200AD: Willow bark used as an analgesic 1763: Antipyretic effects of willow bark reported by Rev E Stone 1828: Büchner isolated pure salicin 1879: Bayer started industrial synthesis of aspirin Serious problems with gastric injury with long-term use Inhibits COX-1 and COX-2. COX-1 necessary for formation of prostaglandins via arachidonic acid
Atropine	1	1831: Mein isolated atropine from belladonna 1867: Atropine shown to block cardiac vagal stimulation Atropine blocks muscarinic acetylcholine receptors
Azathioprine	22-1	Used to treat leukaemia and inflammation Metabolises to form 6-mercaptopurine (6-MP), a guanine analogue that inhibits DNA synthesis and thus reduces production of leukocytes Can also be metabolised by xanthine oxidase to form thiouric acid (inactive) Can also be metabolised by thiopurine methyl transferase (TPMT) to form an inactive metabolite. TPMT activity may differ from individual to individual
Azidothymidine (AZT)	19	First anti-HIV drug NRTI Good oral bioavailability and well-distributed Toxic effects: granulocytopenia, anaemia etc.
Azoles (class)	20	Synthetic antifungals Imidazoles (older) and triazoles (newer) Examples: miconazole, voriconazole, itraconazole Block 14-α-sterol demethylase, which converts lanosterol to ergosterol, causing ergosterol depletion and membrane dysfunction
Benzene	6	Chronic exposure: leukaemia, bone marrow toxicity Acute exposure: CNS depression
Benzodiazepine (class)	12	Some members, such as diazepam (Valium), are anxiolytics Modulators- enhance the opening of GABA-activated Cl- channels
Carbamazepine	3	Induces its own metabolism Half-life drops from 36h to 2h after 2 weeks This is due to induction of CYP3A4
Carbapenem (class)	18	β-lactam that blocks cell wall synthesis Names all end with -penem
Celecoxib	9	COX-2 blocker Causes fewer gastric ulcers than nonselective NSAIDs
Cephalosporin (class)	18	β-lactam that blocks cell wall synthesis Names all begin with cef- 1948- Bacterial source (Cephalosporium acremonium) found in Sardinian sewer There are first-, second-, third- and fourth-gen cephalosporins, differing in activity toward Gram-positive and Gram-negative bacteria
Chloramphenicol	18	Bacterial protein synthesis inhibitor useful against typhus and other severe infections Bind to 50S ribosomal subunit and prevent peptide bond formation Toxic to mitochondria, severely toxic to bone marrow, fatal to neonates due to limited glucuronidation capacity
Cimetidine	4	Suppresses active secretion of organic cations
Clavulanic acid	18	Irreversibly inhibits β-lactamases
Clozapine	3	Half-life decreased in smokers due to CYP450 induction
Cobicistat	3	"Pharmacokinetic enhancer"- inhibits CYP450, slows down metabolism of other drugs Found in Stribild
Codeine	3, (4)	Used to treat mild to moderate pain Codeine is non-analgesic, but it is metabolised to the analgesics morphine and morphine-6-glucuronide Metabolised by CYP2D6 (blocked by fluoxetine) (Morphine may undergo enterohepatic recirculation)
Colistin	17, 18	Polymixin "drug of last resort" used to treat antibiotic-resistant bacteria Poor oral absorption, toxicity issues Acts as a cationic detergent to disrupt cell membranes Best antibiotic against carbapenem-resistant NDM-1 Resistance to colistin by Mcr-1 reported in China in 2015 Mcr-1 encodes a phosphoethanolamine transferase which may alter the lipopolysaccharides in bacterial cell membranes, decreasing affinity for polymixins
Cortisone	12	Powerful anti-inflammatory glucacorticoid
Debrisoquine	3	A heart drug that is a CYP2D6 substrate
Deutetrabenazine	3	Used to treat Huntington's Chorea Replaced H-atoms on tetrabenzine's methyl groups with deuterium (D) Deuterium slows down hepatic metabolism (as opposed to tetrabenzine)
Dexamethasone	12	Powerful anti-inflammatory glucacorticoid
Didanosine	19	Purine analogue inhibitor of DNA synthesis
Diethylstilbesterol (DES)	6	Used to prevent miscarriages Caused vaginal and uterine tumours in female offspring
Diphenhydramine	11	Decongestant Binds to histamine receptors on vascular smooth muscle cells, causing decongestion Also binds to muscarinic acetylcholine receptors on salivary glands, causing dry mouth
Echinocandins (class)	20	Antifungal agents. Safer than amphotericin, but narrower spectrum Example: capsofungin Inhibit 1,3-β-D-glucan synthase, blocking synthesis of 1,3-β-D-glucan and thus altering membrane permeability
Enalapril	2	Lowers blood pressure Ethyl group cleaved off by hepatic esterases Becomes its active form: enalaprilat
Enflurane	10	Volatile inhalational anaesthetic agent used in surgery Racemic R-enantiomer undergoes oxidation by CYP2E1, eventually converted to an achiral metabolite
Esomeprazole	10	Pure S-enantiomer of omeprazole
Ethanol	7, 8	Inhibits CYP2E1 Inhibits bioactivation of paracetamol into NAPQI Can cause fetal alcohol syndrome if used in pregancy. FAS Triad: growth retardation, low IQ, craniofacial abnormalities Meconium markers of foetal alcohol exposure include ethyl-glucuronide, ethyl-sulfate and fatty acid ethyl esters Can also cause ARND (Alcohol-Related Neurodevelopmental Disorders)- CNS impairment in offspring not displaying structural FAS symptoms
Fluorouracil	12	Anti-cancer drug that blocks DNA synthesis Acts as a false substrate, replacing uracil in purine biosynthesis Cannot be converted to thymidylate, blocking DNA synthesis
Fluoxetine	1, 3, 12	Antidepressant (SSRI) Inhibits serotonin reuptake Inhibits CYP2D6 Blocks formation of morphine and morphine-6-G from codeine
GABA (γ-aminobutyric acid)	13	Formed from glutamate in the brain Inhibitory in many CNS pathways Receptors: GABAA and GABAC are ion-channels; GABAB is G-protein-coupled
Glucocorticoid (family)	16	Binds to glucocorticoid receptor, causing dimerisation, dissociation from heat-shock proteins and translocation into the nucleus Once in nucleus: increases gene expression by binding to GRE (glucocorticoid response element), decreases gene expression by binding to nGRE (negative glucocorticoid response element) or has other effects through binding to NFκB
Glutamate	13	Fast-acting. Excitatory in CNS
Glycine	13	Inhibitory in spinal cord Antagonised by strychnine
HIV Protease Inhibitors (class)	19	Inhibit HIV Protease Examples: saquinavir, ritonavir, atazanavir Very effective at reducing viral loads Side effects: gastrointestinal problems, drug interactions, increased risk of heart disease
Ibuprofen	10	NSAID S-ibuprofen is the active stereoisomer: marketed as dexibuprofen in Europe
Imipramine	5	High volume of distribution
Insulin	12, 15, 21	Reduces blood glucose levels Causes glucose channels (GLUT4) to become embedded in the cell membrane, allowing the cells to take up more glucose 3 main pathways, all via IRS (insulin receptor substrate). APS/c-CBL/CAP-mediated pathway creates a targeting site for GLUT4 in cell membrane. IRS/PI3K/Akt-mediated pathway exocytoses GLUT4 storage vesicles (GSVs) and thus embeds GLUT4 into the cell membrane. There is also a long-term pathway: Ras/Raf/Mek/MAP Kinase -> Transcription Factors Excessive insulin can cause hypoglycaemia (Type A ADR)
Isoniazid	17	Narrow-spectrum antibiotic used to treat tuberculosis
Ketamine	10	S-enantiomer has strongest anaesthesia activity and fewer side-effects R-enantiomer associated with psychosis, agitation and amnesia
Lumiracoxib	7, 9	COX-2 selective NSAID Banned in Australia due to 8 serious cases of hepatotoxicity It is simply Diclofenac (a drug that nonselectively inhibits COX-1 and COX-2) with an extra methyl group
Lenalidomide	8	Thalidomide analogue used to treat multiple myeloma
Macrolide (class)	18	Broad-spectrum bacteriostatic drug Examples: erythromycin Binds to 50S ribosomal subunit, plugging the "ribosomal tunnel" and preventing transfer of tRNA from A site to P site Gastrointestinal side effects common, risk of cardiotoxicity
Meperidine (MPPP)	7	Street drug Can cause Parkinson's-like symptoms if contaminated by MPTP MPTP crosses blood-brain barrier and is activated by MAO-B in astrocytes to form MPP+ MPP+ accumulates in dopaminergic neurons, promotes redistribution of dopamine, forms reactive oxygen species, inhibits electron transport chain
Methoxychlor	10	Organic pesticide Originally achiral, but undergoes CYP-catalysed oxidative O-demethylation (i.e. oxygen loses -CH3 group). Results mainly in S-hydroxy metabolite
Mianserin	10	Antidepressant Racemic- each stereoisomer undergoes a different metabolic pathway Chiral centre not altered by metabolism
NNRTI (class)	19	Non-Nucleotide Reverse Transcription Inhibitors Examples: efavirenz, delavirdine etc.
Nystatin	20	First antifungal Discovered by Hazen and Brown Polyene- similar mechanism to amphotericin B (amphotericin is better)
Omeprazole	10	Proton pump inhibitor Reduces gastric acidity and heartburn Racemic mixture
Pancuronium	12, 13	Produces paralysis during anaesthesia Antagonises nicotinic acetylcholine receptors (Na+ channels) Competitive antagonist
Paracetamol/ acetaminophen	7, 12	Over-the-counter analagesic Inhibits cyclooxygenase Intrinsic hepatotoxicant Undergoes sulfation, glucuronidation and bioactivation Bioactivated via CYP2E1, CYP3A4 and CYP1A2 to form NAPQI (toxic metabolite) NAPQI can be trapped by glutathione Glutathione can be replenished by N-acetylcysteine
Penicillin	17, 18	Antibiotic Fleming fround that Penicillum mould inhibited bacterial growth Florey pioneered clinical testing of penicillin β-lactam that blocks cell wall synthesis Can isomerise to form penicilloic acid, creating a novel antigen that triggers an antibody response in susceptible individuals
Phenylephrine	11	Binds to α-adrenoceptors in vascular smooth muscle cells Causes vasoconstriction, relieving nasocongestion Binds only weakly to other receptors, so unlikely to produce adverse effects
Phenytoin	8	Treats epilepsy Risk of "foetal hydantoin syndrome" if used in pregnancy- involves craniofacial and heart malformations, low birth weight, mental deficits Bioactivated by CYP450 to form a reactive epoxide- detoxified by epoxide hydrolase to form nontoxic diol metabolite Bioactivated by fetal peroxidases to form phenytoin hydroperoxide- blocked by antioxidants
Prednisolone	12	Powerful anti-inflammatory glucacorticoid
Probenecid	4	Suppresses active secretion of organic anions
Prontosil	17	Sulfonamide Found to be bacteriostatic by Domagk
Salbutamol	11, 14	Used to treat asthma Interacts with β2-adrenoceptors, releases Gαs and thus activates adenylate cyclase. This converts ATP into cAMP cAMP activates cAMP-dependent protein kinase A Causes bronchodilation
Salvarsan	17	Introduced by Ehrlich Used to treat syphilis
Serotonin (5-hydroxytryptamine)	13	Many actions- increase gastrointestinal mobility, vasodilation, platelet aggregation
Sparteine	3	A heart drug that is a CYP2D6 substrate
Stribild	3	A cocktail of anti-HIV drugs Contains "pharmacokinetic enhaner" cobicistat (CYP inhibitor that slows down the metabolism of other HIV drugs) Only needs to be taken once daily
Strychnine	13	Antagonises glycine Causes convulsions and muscle contraction
Suxamethonium	13	Short-acting muscle relaxant used in short surgical procedures Binds to a ligand-gated ion channel, keeping it open Prevents repetitive depolarisations and thus sustained muscle contraction from occurring
Tetracycline (class)	17, 18	Broad-spectrum antibiotics Prevent aminoacyl-tRNA from binding to A site Poor oral absorption, toxicity includes irritation of the GI tract, hyperpigmentation, staining of teeth etc.
Thalidomide	8, 10	1956: Used to treat nausea and insomnia in pregnancy Well-tolerated in mothers, but led to a large increase in rare limb malformations (amelia: no limbs, phocomelia: shortened long bones) Many theories surrounding mechanism for teratogenicity One such theory is chiral toxicity- however, not much experimental data to support this
Theophylline	3	Half-life decreased in smokers due to CYP450 induction
Thiopental	2	Very lipophilic
Tri-o-cresylphosphate (TOCP)	6	Plasticiser used in varnishes, lubricants etc. Has anticholinesterase activity 1 week following exposure: delayed neuropathy syndrome
Valproate	9	Anticonvulsant Hepatotoxic and teratogenic Teratogenicity possibly through inhibition of histone deacetylase enzymes (HDACs) which regulate chromatin remodelling Antiepileptic properties maintained over a range of structural changes, hence valproate analogues are being investigated to reduce teratogenicity
Warfarin	2, 5	99% protein-bound Low volume of distribution

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."

0. Initial human pharmacokinetic and pharmacodynamic studies. Very low doses are used.
1. 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.
2. 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).
3. 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.)
4. 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*)

• 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.

• 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.

Identify basic features of Type A adverse drug reactions, identifying insulin hypoglycaemia as a classic example of these syndromes

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.

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.

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.

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:

• 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

In the next pharmacology post, I will talk a bit more about antibiotics.

Abdominal Wall and Hernia

Describe the surface anatomy of the abdominal wall

In textbooks and so forth, you might read about how the abdomen can be divided up into 9 regions, like a noughts and crosses board. The two vertical lines subdividing the abdomen are in line with the middle of the clavicle, and are called "midclavicular lines." There are two horizontal lines: one is in line with the bottom of the ribs (subcostal line) and the other is in line with the tubercles of the pelvic iliac bone (intertubercular line).

Here are the 9 regions of the abdomen, in a table, because tables are cool.

Right hypochondrium	Epigastric	Left hypochondrium
Right lumbar	Umbilical	Left lumbar
Right iliac/inguinal	Hypogastric	Left iliac/inguinal

Describe the layered structure of the abdominal wall and its function

The abdominal wall has several important functions. Aside from protecting the abdominal organs, the muscles of the abdominal wall can also contract and relax to change the pressure within the abdominal cavity. This, in turn, affects processes such as breathing, defecation and so on.

The abdominal wall is made up of many layers. The innermost layer is the parietal peritoneum, which surrounds the peritoneal cavity, as mentioned in my post about coelomic cavities. Surrounding this is a layer of extraperitoneal fat and then the transversalis fascia. Next up there are the three main muscles of the abdomen: the transversus abdominus (which has fibres running transversely), internal oblique (which has fibres running medially and superior- i.e. from bottom up) and external oblique (which has fibres running medially and inferior). The aponeuroses (tendinous parts) of these three muscles combine to form the rectus sheath, which surrounds the rectus abdominus which lies at the front of the abdomen.

A quick note on the rectus sheath: it has two layers, anterior and posterior. The anterior layer is made up of the aponeurosis from the external oblique as well as part of the aponeurosis from the internal oblique. The posterior layer is made up of the rest of the aponeurosis from the internal oblique as well as the aponeurosis of the transversus abdominus. Towards the bottom, however, all three layers pass in front of the rectus abdominus. The point at which this happens is the "arcuate line." Because of this, the abdominal wall is weaker below the arcuate line.

Another quick note: the medial edge of the rectus sheath (i.e. the part of the sheath between the two "halves" of the rectus abdominus) is called the linea alba, and the lateral edge is called the linea semilunaris.

Anyway, back to the layers. Outside of the muscles there is another fascia, called the superficial layer. It has a fatty layer, as well as a membranous layer. This membranous layer is called Scarpa's fascia and continues down into the scrotum or labia to become Colle's fascia. Finally, outside this fascia there is skin.

Understand how and why the wall structure differs between the sexes

Obviously, one of the main anatomical differences between the sexes is that males have penises while women have vaginas. Males also have testes, and when they descend, they drag some of the body wall into the scrotum with them. Some of the peritoneal cavity gets dragged along too, so each testis has a mini cavity called the tunica vaginalis.

The scrotum has pretty much all the same layers as the rest of the abdominal wall, just renamed or with some slight differences. The skin of the scrotum also has a layer of muscle called dartos muscle, which contracts the scrotum when it gets cold. Scarpa's fascia is now called Colle's fascia, as mentioned above. The external oblique is now the external spermatic fascia, and the transversalis fascia now the internal spermatic fascia. The main difference is what lies in between them: the internal oblique and transverse abdominus combine to develop the cremaster muscle and fascia. (That's the muscle that draws the testes closer to the body when it gets cold.)

Understand how sex determines risk of hernia

To talk about this, first I need to describe what a hernia is. A hernia is a weakness in the body wall that allows stuff to poke through when the abdominal pressure is increased.

The most common type of hernia is the inguinal hernia. The inguinal canal is a canal that passes through the aponeuroses of the three main abdominal muscles. It runs obliquely (i.e. diagonally), from the deep inguinal ring above the halfway point of the inguinal ligament, to the superficial inguinal ring at the medial end of the inguinal ligament. The spermatic cord (in males) or round ligament/ ligamentum teres (in females) pass through here, but in a hernia other things can pass through too.

There are two main types of inguinal hernia. The first, indirect inguinal hernia, is usually congenital. It occurs when the processus vaginalis (the peritoneum that descends into the scrotum) doesn't fuse shut, creating what is known as a "patent processus vaginalis." Abdominal organs can then herniate through the spermatic cord (males) or round ligament (females). The second type of inguinal hernia, direct inguinal hernia, is usually acquired as a result of weak musculature. This hernia occurs through the superficial inguinal ring.

As males have more stuff going through the inguinal canal (spermatic cord vs. round ligament), and they also have the whole issue of the processus vaginalis descending into the scrotum which may or may not close, they are much more at risk of getting inguinal hernias than females.

Females are, however, more at risk of femoral hernias. This is because their femoral canals (where femoral arteries and veins pass through) are much larger than males.

Hernias are bad because they can result in blood vessels or organs getting squished. There may be some ways of preventing them though. Aside from indirect inguinal hernias which are usually congenital, most hernias are acquired through strenuous activities such as childbearing, heavy coughing, weightlifting etc. This is why weightlifters often wear a belt to support their abdominal wall.

Fundamental Enzyme Kinetics part 3: Attack of the Inhibitors

Not all remained good in EnzymeLand. Soon the inhibitors arrived, and they were here to steal those enzymes. They came in all different varieties...

Competitive Inhibitors

The competitive inhibitors had come, and they had come to steal the enzymes. Not long after their arrival, the enzymes had gotten themselves preoccupied in new relationships with those hot new inhibitors:

This meant that now, fewer enzyme-substrate complexes were forming, and even fewer products were forming. This was a sad day for all.

At the Relationships Institute of EnzymeLand, they began conducting some research to find out which competitive inhibitors were most successful at competing with substrates for enzymes. Once again, they came up with a shiny new equation, this time for the dissociation of the enzyme-inhibitor complex:

EI -> E + I

They gave this equation a rate constant of K_I. Since this is the rate constant for the dissociation of EI to form free E and I, the lower the K_I, the better the inhibitor was at competing with the substrate and hanging on to those relationships with the enzymes.

Another curious phenomenon that those Relationships Institute researchers found was that, if the substrate concentration was much higher than the inhibitor concentration, the substrates could easily outcompete the inhibitors and hence the V_max returned to normal. Strength in numbers, as they say!

Of course, those nerds at the Relationship Institute wouldn't be nerds if they didn't like graphs. Here's the graph that they prepared earlier:

The y-intercept, which from my previous post was 1/V_max, was the same regardless of whether the competitive inhibitor was present or not. The x-intercept (-1/K_m), however, did change. This shouldn't be surprising, as those pesky inhibitors were preventing those enzymes and substrates from getting together for a while. Anyway, from the graph, 1/K_m decreased, which incidentally meant that K_m increased. (You do have to play around with opposites a bit when reciprocals are involved.)

Uncompetitive Inhibitors

The next lot of inhibitors decided that they would not be quite as selfish when they sought out the enzymes. Instead of taking the enzymes for themselves, they thought that they'd be polite and slot themselves in to the enzyme-substrate complex, like so:

(See? I told you that there'd be steamy bed action!)

Anyway, the result of this was that the enzymes and substrates could get together, but there'd always be an inhibitor waiting to join in the action. This decreased the substrates' morale, and prevented their transformation into a product. Strength in numbers didn't seem to overcome this problem either: no matter how much more substrate there was, as soon as that substrate got together with an enzyme, the inhibitor would be there too. Watching. Waiting.

What did the Relationships Institute think? Well, they started off with an equation again:

EIS -> ES + I

This time, they called the rate constant K_IS. Once again, the lower the K_IS, the more effective the inhibitor at preventing the formation of product.

Here's the graph that they made:

As you can see, the 1/V_max increased, and so did 1/K_m, which means that V_max and K_m both decreased. (Remember, reciprocals = opposite day). The V_max decreased because as I said earlier, no matter how much substrate there was, an inhibitor would always be there to prevent its final transformation into a product. The change in K_m is a bit less intuitive to understand though. You see, when the enzyme-substrate-inhibitor complex forms, that means that there is less ES floating around. That shifts the E + S <--> ES equilibrium in favour of ES formation. This means that eventually pretty much all the substrate you put in is going to end up as ES or EIS, so that the apparent K_m ends up being lower than what you'd get without an uncompetitive inhibitor.

Noncompetitive (Mixed) Inhibitors

Noncompetitive (mixed) inhibitors decided that they'd be extra fancy and use both tricks: they'd steal free enzyme, just like the competitive inhibitors, and they'd insert themselves into an existing enzyme-substrate relationship, just like the uncompetitive inhibitors.

The characteristics of mixed inhibitors essentially combined the characteristics of competitive and uncompetitive inhibitors. The final outcome depended on how much competitive vs. uncompetitive inhibition that the noncompetitive inhibitor liked to engage in. If it was better at using competitive inhibition (i.e. K_I < K_IS - remember that a lower K_I or K_IS = better inhibition), then its characteristics would be closer to that of competitive inhibition, or vice versa.

No matter what kind of inhibition prevailed, noncompetitive inhibitors would always result in a lower V_max. This is because competitive inhibition didn't have any effect on V_max, while uncompetitive inhibition caused it to decrease.

K_m got a bit more interesting, however. Noncompetitive inhibitors more adept at using competitive inhibition (i.e. K_I < K_IS) had a higher K_m. The opposite was true for those more adept at uncompetitive inhibition: they had a lower K_m. There were, however, some special noncompetitive inhibitors that were equally adept at using both: they didn't have any effect on the K_m.

Irreversible Inhibitors

Many inhibitors were reversible, which meant that they, too, could suffer break-ups with enzymes. But there were a few that could not. These were the irreversible inhibitors.

They, masters of black magic, could bind irreversibly to the enzymes. Thus ended the romantic lives of those poor enzymes: no substrate would ever want to be with them now that they would have to put up with an inhibitor too. As time passed, more enzymes would be bound to inhibitors, and so their romantic activity would decrease.

Allosteric Enzymes

Forget the inhibitors- sometimes enzymes could be their own worst enemies. Enzymes could exist in either a tense state (T state) or a relaxed state (R state). When tense, their romantic lives suffered, and they couldn't form enzyme-substrate complexes quite so readily. They were much better partners in their relaxed state, however.

Most enzymes existed in an equilibrium between being tense and relaxed, which could be represented as T <--> R. Without substrate available, the tense state would prevail- I suppose because the enzymes were worried that they'd die lonely. Once some substrate came along and bound to that small percentage in the R state, however, the equilibrium would begin to shift. The enzymes would increase their morale as they saw their friends finding partners and settling down. As more substrate arrived, the more relaxed the enzymes became.

There were, however, other influences on whether enzymes were tense or relaxed. Allosteric inhibitors tended to make enzymes more tense, so that fewer enzyme-substrate complexes would occur. Allosteric activators, however, were the opposite: they would relax the enzymes so that they could find love more quickly.

And thus concludes this, er... lovely series of blog posts! Until next time!