Preclinical Drug Research – INSERT
Norvir®
Molecular weight = 720.95
lipinsky’s rule of five MW: 200-500 = too big = bad absorption
— a very large compound will not pass the membranes = less absorption
Freely soluble in methanol and ethanol, soluble in isopropanol and practically insoluble in
water
— no solubility in water
— but we don’t have information about the absolute bioavailability why?
o it is insoluble in water you cannot administer it IV
o IV with methanol is also not possible = massive irritation
you cannot determine the absolute bioavailability
Route of administration
gelatin capsules
oral solution
2 formulations: for the pedriatic population
baby cannot get infected during the pregnancy (when the mother is infected)
during labor there is a risk that the baby can get infected get in contact with the blood of
the mother
— solution: caesarean section
a lot of of pedriatic patients
— dose is too high + pill itself its too large
— oral solution with a flavour: compound is too bitter + babies/kids will like this more
they will not spill it out
comparison of bioavailability in both solution
— relative bioavailibility
bio-equivalence:
— comparison with a reference drug (with the same mode of action) To know if your
new drug is better than the one already on the market
— mechanism of action: inhibitor of both the HIV-1 and HIV-2 proteases
Clinical pharmacology
Potency low affinity for the target is not good
exogenous targets no off-target effects
very high therapeutic index concentration where we expect an efficacy compared to the
concentration where you see adverse effects (cytotoxicity) difference of 1000-fold
— The IC50 of viral replication ranged from 3.8 to 153 nM
— cytotoxicity studies on several cell lines showed that >20 M was required to inhibit
cellular growth by 50% resulting in an in vivo therapeutic index of at least 1000.
Cross-resistance to other anti-retroviral
among protease inhibitors, variable cross-resistance has been recognized
cross-resistance between ritonavir and reverse transcriptase inhibitors is unlikely
Safety pharmacology
no data
, exogenous compound + 1000 fold difference (therapeutic index) + no drugs for an effective
HIV treatment you can deviate from the guidelines
Examen: what should you normally perform CORE BATTERY
— name of tests + explanation
Pharmacokinetics
ritonavir has been studied in healthy volunteers and HIV-infected patients
The absolute bioavailability of ritonavir has not been determined
peak concentrations of ritonavir were achieved approximately 2 hours and 4 hours after
dosing under fasting and non-fasting conditions, respectively.
— non-fasting: will start interfering with the absorption (plasma concentration will be
lower)
after 4 hours you achieve plasma peak concentrations effect will appear later
effect of food on oral absorption
— no difference between the oral solution and the capsule
metabolism
— isopropylthiazole oxidation metabolite is the major metabolite and has antiviral
activity similar to that of the parent drug !! concentrations of this metabolite in
plasma are low
— liver microsomes have demonstrated that CYP3A is the major isoform + CYP2D6
contributes
— lot of metabolisms drug will be cleared less compound + less effect
o but one of the metabolites is active
contribution will be low to the efficacy, because the concentration of the
active metabolite is low
— what other in vitro tests can be performed?
o S9 fractions (entire cytosolic fraction present) or primary hepatocytes (the
cells itself) or liver slices
liver microsomes are very valuable for inhibition and affinity studies
S9 fraction is good for phase 2 enzymes
you need the complete cells (hepatocytes) for induction studies
— how can you asses that ritonavir affinity CYP3A and CYP2D6
o recombinant enzymes
o liver microsomes how could you know this? specific inhibitors for
CYP3A
first you add the inhibitors, then the drug if the concentration
doesn’t change, it will be metabolized by this enzyme
inhibitor is already bounded to the CYPs and that’s why the
drug cannot bind anymore
indirect way for assessing this
not always specific can bind with a different affinity to other
enzymes
CYP2D6 = polymorphic you need to do sequencing + check what
the polymorphic gene is maybe change the dose
slow reduce dose to avoid toxicity
high increase dose to see an effect
elimination
only a small proportion is cleared by the kidneys faeces = route of elimination
— what type of study was used? mass balance study
Norvir®
Molecular weight = 720.95
lipinsky’s rule of five MW: 200-500 = too big = bad absorption
— a very large compound will not pass the membranes = less absorption
Freely soluble in methanol and ethanol, soluble in isopropanol and practically insoluble in
water
— no solubility in water
— but we don’t have information about the absolute bioavailability why?
o it is insoluble in water you cannot administer it IV
o IV with methanol is also not possible = massive irritation
you cannot determine the absolute bioavailability
Route of administration
gelatin capsules
oral solution
2 formulations: for the pedriatic population
baby cannot get infected during the pregnancy (when the mother is infected)
during labor there is a risk that the baby can get infected get in contact with the blood of
the mother
— solution: caesarean section
a lot of of pedriatic patients
— dose is too high + pill itself its too large
— oral solution with a flavour: compound is too bitter + babies/kids will like this more
they will not spill it out
comparison of bioavailability in both solution
— relative bioavailibility
bio-equivalence:
— comparison with a reference drug (with the same mode of action) To know if your
new drug is better than the one already on the market
— mechanism of action: inhibitor of both the HIV-1 and HIV-2 proteases
Clinical pharmacology
Potency low affinity for the target is not good
exogenous targets no off-target effects
very high therapeutic index concentration where we expect an efficacy compared to the
concentration where you see adverse effects (cytotoxicity) difference of 1000-fold
— The IC50 of viral replication ranged from 3.8 to 153 nM
— cytotoxicity studies on several cell lines showed that >20 M was required to inhibit
cellular growth by 50% resulting in an in vivo therapeutic index of at least 1000.
Cross-resistance to other anti-retroviral
among protease inhibitors, variable cross-resistance has been recognized
cross-resistance between ritonavir and reverse transcriptase inhibitors is unlikely
Safety pharmacology
no data
, exogenous compound + 1000 fold difference (therapeutic index) + no drugs for an effective
HIV treatment you can deviate from the guidelines
Examen: what should you normally perform CORE BATTERY
— name of tests + explanation
Pharmacokinetics
ritonavir has been studied in healthy volunteers and HIV-infected patients
The absolute bioavailability of ritonavir has not been determined
peak concentrations of ritonavir were achieved approximately 2 hours and 4 hours after
dosing under fasting and non-fasting conditions, respectively.
— non-fasting: will start interfering with the absorption (plasma concentration will be
lower)
after 4 hours you achieve plasma peak concentrations effect will appear later
effect of food on oral absorption
— no difference between the oral solution and the capsule
metabolism
— isopropylthiazole oxidation metabolite is the major metabolite and has antiviral
activity similar to that of the parent drug !! concentrations of this metabolite in
plasma are low
— liver microsomes have demonstrated that CYP3A is the major isoform + CYP2D6
contributes
— lot of metabolisms drug will be cleared less compound + less effect
o but one of the metabolites is active
contribution will be low to the efficacy, because the concentration of the
active metabolite is low
— what other in vitro tests can be performed?
o S9 fractions (entire cytosolic fraction present) or primary hepatocytes (the
cells itself) or liver slices
liver microsomes are very valuable for inhibition and affinity studies
S9 fraction is good for phase 2 enzymes
you need the complete cells (hepatocytes) for induction studies
— how can you asses that ritonavir affinity CYP3A and CYP2D6
o recombinant enzymes
o liver microsomes how could you know this? specific inhibitors for
CYP3A
first you add the inhibitors, then the drug if the concentration
doesn’t change, it will be metabolized by this enzyme
inhibitor is already bounded to the CYPs and that’s why the
drug cannot bind anymore
indirect way for assessing this
not always specific can bind with a different affinity to other
enzymes
CYP2D6 = polymorphic you need to do sequencing + check what
the polymorphic gene is maybe change the dose
slow reduce dose to avoid toxicity
high increase dose to see an effect
elimination
only a small proportion is cleared by the kidneys faeces = route of elimination
— what type of study was used? mass balance study
