Lecture 1

Question 0

What is a receptor?

Answer 0

A biological molecule whose function is modulated (changed in some way) by a chemical combination with the drug. Another term for a drug receptor is target.

Question 1

What is a drug?

Answer 1

An agent that alters the rate or extent of a biological process. Often referred to as a ligand. In modern pharmacology, drugs and receptors can be switched in that the receptor can be administered to change a response. But for the most part drugs refer to the ligand acting on the receptor. In some cases, receptors can have multiple binding sites.

Question 2

What is pharmacokinetics?

Answer 2

Study of drug distribution, metabolism, and elimination in a living system. Adsorption, Distribution, Metabolism, Excretion.

Question 3

What is pharmacodynamics?

Answer 3

Study of the interaction between drugs and receptors at the molecular level. Study of the binding of drugs to receptors.

Question 4

What are some examples of drug receptors?

Answer 4

GPCR: Hormones: Neurotransmitters

Ligand-Gated Ion Channels: Neurotransmitters

Enzymes (COX)

Cotransporters

DNA (Nucleic Acids)

Signal Transduction Proteins (e.g. Kinases: potential target for cancer)

Structural Proteins (Microtubules)

Membranes and Bacterial Cell Walls

Ribosomes

Proteosomes: large macromolecular complex that degrades proteins
that are either misfolded or not meant to be expressed/ active at
certain points of the cell cycle. Proteosome inhibitors could be useful
in cancer therapeutics. Proteosome stimulators could be useful
in neurodegenerative diseases that involve aggregations of insoluble
misfolded proteins.

Question 5

What are important parameters used to characterize a drug?

Answer 5

Potency, affinity, and efficacy.

Question 6

What is drug potency?

Answer 6

The amount of drug required to elicit a certain biological response. Often described as the dose of a drug given. Can be quantified by the concentration of drug in some body fluid e.g. blood plasma. The lower the dose or concentration required to solicit a biological effect, the greater the potency.

Question 7

What were Paul Ehrlichs important conceptual contributions to pharmacology?

Answer 7

1. ) affinity: the attractive force between substances that causes them to enter into and remain in chemical combination.
2. ) chemotherapy: using chemicals not created in the body to fight diseases.
3. ) magic bullet: a drug that can kill microbes but not the person with the disease.
4. ) lock and key: the idea that a drug molecule has a shape that fits the shape of the receptor site.

Question 8

What’s a requirement for a magic bullet?

Answer 8

In order for a therapeutic agent to work, it has to be somewhat selective for the microbe so as to not kill the host too.

Question 9

What is the idea of template action?

Answer 9

Idea of template action: A drug can modify the receptor by changing its structure in whats
called induced fit. Probably not the best explanation for how the immune system works, but
prions may utilize template action to convert folded proteins into misfolded aggregates.

Question 10

How do you know that there is enough drug getting to where it needs to be?

Answer 10

Could take a biopsy,
grind it up, and measure the amount of drug present. An easier way would be to take a sample
of bodily fluid such as blood plasma. Can then measure the drug concentration and calculate
the concentration of drug present in the area of interest.

Question 11

Methylene blue as a dye

Answer 11

Stain is not retained in the cytoplasm. Methylene blue intercalates the DNA and also binds to the phosphate backbone.

Question 12

Methylene blue as a drug

Answer 12

Useful in the treatment of malaria. Was an early treatment. Could be used when the malaria is resistant to all of the other drugs. . When its exposed to light it undergoes a conformational change that makes it useful as an antibacterial agent. There are many other options for antibacterial agents, but as drug resistance becomes
more prevalent, it can be used more and more. Its backbone was used
to develop tricyclic antidepressants. Tricycs are also used to treat
anxiety and other psychiatric disorders.

Question 13

Why do most drugs interact through non-covalent interactions? Give an example of a drug that acts through a covalent interaction.

Answer 13

You want a reaction that is reversible because most of the target molecules are essential to normal function. If using a covalent interacting drug, you may inhibit these target molecules too much. The exception is if the body can easily churn out more of the target relatively fast so as to minimize the negative impact the
covalent drug would have on the bodies normal function. Aspirin is an example of an acetylating acting drug that works to “kill” COX enzymes. The body can make more COX back to normal levels in about 4 hours so its ok. Platelets cant make new COX though, so they are permanently inhibited so there is still potential for overdose.

Question 14

What is the lock and key model?

Answer 14

Interacting groups in the structure of a drug and it’s receptor must be positioned very close to each other in order for the chemical bonds that hold the two together to form. This requires that the drug must fit tightly into the 3-D structure of the receptor.

Question 15

What are the weak forces involved in drug receptor binding?

Answer 15

1. ) ionic bonds ~5-10kcal/mol
2. ) hydrogen bonds ~3-7kcal/mol
3. ) van der walls forces 1kcal/mol
4. ) hydrophobic ~1-2kcal/mol

The weaker interactions require a specific architecture for them to be strong enough to hold the drug and receptor together.

Question 16

If the weak bonds that contribute to drug receptor binding are weak, how can they lead to successful interaction? Give the example with methylene blue.

Answer 16

These weak bonds have low binding energy, but they make up for it with a high number of these interactions. With the case of methylene blue, it interacts by stacking in between DNA bases and also binds along the phosphate backbone. The strongest interaction is the intercalating action which
occurs through hydrophobic interactions between the bases and the methylene blue molecule. Theweaker interaction is the electrostatic interaction occurring through charge interactions between the phosphate and the positively charged group on methylene blue. So even though, alone, the ionic bonds are stronger,
there are fewer of them present in this case and so the hydrophobic interactions contribute more. If you have a limited amount of methylene blue, you’ll have more of it bound as the intercalating agent than bound through ionic interactions.

Question 17

What does equilibrium mean?

Answer 17

Equilibrium: Equal, unchanging amounts of drug and drug-receptor
complex. These amounts do not have to be equal, but the relative amounts
should be unchanging. Some reactions reach equilibrium fast, some take a while. There is an inverse relationship between Kd and time required to reach equilibrium, so a drug with a low affinity will reach equilibrium faster than a drug with a high affinity.

Question 18

why is Kd used

Answer 18

KD is used primarily because of the units which are moles/liter aka molarity. How to tell between KD and Ka? If the exponent is positive: 5E6, then
we are most likely dealing with the association constant. If its a negative value on the exponent then its most likely the dissociation constant.

Question 19

What does it mean if Kd is much higher than Ka

Answer 19

If KD is much higher than Ka, then thats telling you that you have much more free receptor present than
drug-receptor complex. So this drug would have a low affinity for the receptor.

Question 20

What graph would you look at if you don’t have a purified receptor?

Answer 20

If you dont have isolated receptor, then youll most likely be looking at percent biological response versus the drug concentration. Biological response is not always positive (blood pressure lowering drugs).

Question 21

Kd and Kd app

Answer 21

Kd and Kd Apparent (Kd app)

• Kd is the equilibrium dissociation constant = [drug] at which 50% of the drug receptors are occupied. KD apparent is a measure of drug potency.
• Kd can be determined from arithmetic plot (fraction bound vs. [drug], log plot, inverse plot (Lineweaver Burke), or Scatchard plot
• Rang uses KA in place of Kd
• Kd apparent is determined by plots of % biological response vs. [drug]; also known as EC50
• Scatchard plot cannot be used to calculate KD apparent, because it looks at binding.

Question 22

What is the general assumption of potency and affinity?

Answer 22

The assumption is that potency is due to the binding affinity. If possible, its useful to plot both the KD apparent and the
KD on the same plot using two different Y-axises, the two lines should coincide perfectly at where the KD/KD apparent
is if the KD apparent is indeed due solely to binding affinity. This is not always the case. When using the log dose response curve, the KD apparent is less likely to be affected by scattered data due to experimental error because
it lies in the linear region of the graph and scattered data wont affect
the slop too much.

Question 23

What is an agonist

Answer 23

Agonist:
Favors the biological response thats necessary for changing
the normal biological response. Sometimes are naturally endogenous
molecules that aren’t present in adequate amounts to get a desired response, so they’re added exogenously but are the same molecules as the endogenous ones. Sometimes they’re exogenous compounds that can give a stronger response than the normal compound. .

Question 24

What is an antagonist?

Answer 24

The vast majority of drugs are antagonists. They compete with natural agonists
and block its binding and therefor its ability to stimulate a biological response.
Key distinguishing feature is that antagonists have absolutely no activity
in the biological response for the particular interaction. They may act as
agonists at other responses for other receptors though.
They do not stimulate the activity of the receptor in any way.

Question 25

Competitive antagonism

Answer 25

Binds reversibly to the same site as the agonist. Shifts a log dose response curve to the right.

Question 26

Efficacy and affinity

Answer 26

Affinity: how tightly the drug is binding to the receptor.
Efficacy: Once bound, how effective the drug can alter the structure or function of the receptor in order to get a biological response. Antagonists have no efficacy. Some drugs can cause a small change,
but not enough to elicit the maximal response seen with an agonist, so these are classified as partial agonists, they can never
achieve the maximal response. .

Question 27

Noncompetitive antagonist

Answer 27

Binds reversible to an allosteric site or irreversibly to the same site as agonist (rare). Lowers the maximum response in a log dose response curve, does not shift the curve to the right. Binding to the allosteric site is able to indirectly effect the binding of the agonist to it’s normal active site. Never able to outcompete the noncompetitive antagonist. Looks like a partial agonist, but since there’s multiple drugs being used you know a noncompetitive antagonist is present.

Question 28

Occupancy assumptions

Answer 28

1) The drug acts by binding to a receptor, AND the number of receptors is fixed. Physiological adaptations can occur where the number of receptors may be upregulated or
downregulated by the organisms cells. This will lead to violation in the first rule.
2) Biological response is proportional to occupancy of the receptors; e.g. 50% occupancy causes 50% response.

3) The concentration of free drug at the receptor site is the same as what is typically measured (e.g., drug concentration in the blood, culture medium, etc). If you have a receptor that is present at a distant location than the site of administration, there may be barriers blocking some of the drug from reaching the site of action. Alternatively the drug can be pumped excessively into a cell. In both cases if the drug concentration is measured somewhere besides its site of action, the measurement may be flawed.
If there are violations in these rules then the KD may not be equal to the
KD apparent.

Question 29

What are extra receptors. Give an example of a system with them.

Answer 29

You need to bind extra receptors in order to elicit 50% of a response. There is so muchcarbonic anhydrase that its present in excess than whats necessary to regulate this balance, so you need to inhibit almost all of it in order to get a significant biological response.

Question 30

What are spare receptors. Give an example of them. How can you tell if you are dealing with spare receptors?

Answer 30

You bind less receptors in order to elicit 50% of response. Hence, spares left over. G protein coupled receptors. Easy way to determine if you are dealing with spare receptors is to use an irreversible inhibitor for the particular drug-receptor interaction. This is a noncompetitive situation. Assuming that the first curve shown is just with the agonist itself while the second and third curve represent the addition of an irreversible inhibitor at two different doses. However the curves shown are not what is expected with an
irreversible inhibitor: we would expect the Emax to lower but the Kd would stay the same so we wouldnt see the shift to the right. An explanation is that all of the receptors are saturated (even the ones not needed),
so now you need more drug in order to get the same effect that you were previously with all of the receptors. Once you are effecting the actual number of receptors needed to see the effect, then you get the
expected shift down. The right shift may be observed depending on the concentration of drug being used: at higher concentrations you should start to only see shift downs, so subsequent curves will only
have this feature and less of the shift. This trend is a particular indication of the spare receptor effect taking place, without knowing the specific molecular interactions taking place.Diurectics can treatgluacoma by
decreasing the retention of sodium ions. the amount of bicarbonate ion affects the action of the sodium cotransporters.

Question 31

What is QSAR

Answer 31

Quantitative Structure Activity Relationship

1) Identifying the desired receptor (target) (example the glucocorticoid receptor): if all you have is the biological response and don’t know what the receptor is.
2) Mapping the active site on a receptor (example, nicotinic acetylcholine receptor): could use to determine the types of interactins necessary to elicit a biological response and could see how far apart the groups are interacting with the receptor.
3) Distinguishing mechanistically between toxic (undesired effects) and desired drug effects: could use to make a more potent version of the drug.
4) Searching for a more potent drug

Question 32

How can QSAR be used

Answer 32

QSAR can be used to identify features of the receptor prior to
isolation. Different conjimers are used. In a normal log dose response curve
there is a nice plateua at higher drug concentrations. An explanation for why the graph below
doesn’t have this is because of nonspecific binding of the drug to something else in the body
in addition to what may be specific binding. To show that there is some specific binding occurring, you can label the
agonist you are using, and then take an unlabeled conjimer and look at binding of the labeled agonist without and with
the additional unlabeled drug. The unlabeled drug is able to displace the binding but only in a certain region.

Question 33

What is nonspecific binding. How can you prove nonspecific binding is occurring where a plateu is absent.

Answer 33

Nonspecific binding: Could be binding to a lipid, or maybe a hydrophobic region
of a protein. Possiblyserum albumin (a sinc for many drugs that get into circulation). Taking the difference between the curves gives you the more
typical curve you would expect. Soyou use structurally similar conjimers that
have changes in side groups to gain evidence that there is specific binding occurring in that portion of the curve.Because you dont see similar displacement in the higher drug concentrations, it proves that there is
nonspecific binding occurring in thehigher concentration
regions.

Question 34

Nuclear receptors

Answer 34

Ligand (Red) is initially outside of the cell while the receptor starts out inside of the
cytoplasm. Ligand (lipid soluble) binds to the cytoplasmic receptor. This complex
can then translocate into the nucleus and bind to a specific sequence of DNA
upstream to one or more genes and regulate (usually in a positive manner) the
transcription of the genes. Regulation of P450 is an example of an enzyme that is
regulated by a nuclear receptor. The example shown is dealing with the glucocorticoid
receptor (responds to hormones), which is involved in regulating glucose metabolism.

Question 35

What are two things to consider when developing a drug?

Answer 35

When developing a drug, the focus is on two things. 1. How effective is the drug at producing a desired response. This deals with potency and looks at the ED50. 2. The toxic effects are analyzed and ideally there is a good distance between the effective dose and toxic dose (high therapeutic index). The bigger the TI, the safer the drug. Goal is to modify the drug to lessen the toxic effect while keeping the desired effect. Modifications can also increase potency and reduce the chance of the drug hitting off-site targets.

Question 36

How was QSAR used to measure the binding site of ACh on the nACHR?

Answer 36

QSAR can be used to try to define the distances within binding sites within the nicotinic acetylcholine receptor. Receptor has two binding sites for ACh. 5 subunits. Multiple genes for the subunits so theres many different types of receptors. This can be done prior to isolation of the receptor. Different conjimers with modifications to the side chain of ACh were used.

Question 37

What is the therapeutic index?

Answer 37

TI= TD50/ED50 , where TD50 dose that produces a toxic effect in 50% of individuals in a population and ED50 is the effective 34 dose for 50% of individuals in the population