Transcript cf. J.Med.Chem. 36: 4131, 1993.

Natural Product Drug Discovery and
Development - an NCI Perspective
John A. Beutler, Ph.D.
The funnel of drug discovery and
development
Total Cost
Number of Compounds
0
2
2
ADME
4
Discovery
Pharmacology
4
6
6
Years
Toxicology
Years
• The task: find the “needle in
the haystack”
• Estimated 10,000 compounds
tested to find one which has
clinical activity in humans
• Cost of process increases as
one nears the goal
• Estimated $300-600 million
average development cost for a
single drug
0
8
8
Phase I Clinical
10
10
Phase II Clinical
12
14
16
1e-1
Phase III Clinical
1e+0
1e+1
1e+2
1e+3
Number of Compounds
1e+4
12
14
16
1e+5 1e-1 1e+0 1e+1 1e+2 1e+3 1e+4 1e+5 1e+6 1e+7 1e+8
$$$
Primary screening
• This is the filter that is supposed to separate
the “wheat” from the “chaff”
• Cost, speed, relevance all important
• Whole animals precluded due to cost
• Tissue assays too slow
• Cellular or biochemical assays best – using
a validated molecular target
But….
What does one lose by throwing out the
context in which the target operates?
• Penetration to target inside cell
• Metabolism, excretion
• Toxicity through other mechanism
Secondary screening
• Attempts to put back some of the factors
which the primary screen ignored
• Biochemical mechanisms, tissue assay, or
whole animal
• Reduce the number of active samples to be
considered by eliminating the least desirable
ones
Animal testing
• Toxicology – what dose kills or injures animal, in
what organ system? (e.g.,liver, brain)
• Can therapeutic blood levels of drug be obtained?
• Extrapolation from model animal to human
imprecise but necessary
• Cost requires small animals: mouse, then dog
• Computer models limited usefulness, but cellular
toxicology increasingly used
Clinical trials
•
•
•
•
Phase 0 – sub-pharmacologic doses
Phase I – first time in humans - safety
Phase II – efficacy in small numbers
Phase III – larger populations
Postmarket processes
• Newly introduced drugs are closely
monitored to detect lower frequency
adverse effects
• Unacceptable toxicity may lead to drug
withdrawal
• Broader indications may be explored
without FDA approval (“off-label uses”)
Plants for medicines
• Direct application of ethnomedical
use
• Source of chemical diversity for
screening
• Herbal products under DSHEA
The ethnomedical approach
• Problem: It may be hard to correlate folk medical
concepts to Western medicine
• Shaman Pharmaceuticals: antifungals,
antidiabetics
• Property rights asserted by countries over
ethnomedical knowledge, genetic heritage
Herbal products under DSHEA
• Dietary Supplement Health and Education
Act of 1994
• No premarket evaluation for efficacy or
toxicity
• Requires correct labeling, GMPs
• Established Office of Dietary Supplements
within NIH
Chemical diversity of plants
• Attractive for screening in bioassays – each
extract can contain many novel compounds
[x number of plant species]
• Plant defensive compounds may be tailored to
interfere with mechanisms common to human
diseases
• Antibiotic/antifungal compounds may be useful
both to the plant and to human medicine
• Many drugs originally obtained from nature
A Case History
NCI collectors
collect tree
Calophyllum
lanigerum from
Malaysian
rainforest
Screening for HIV inhibitors
• Extracts of many plants screened for the
ability to protect host cells against the cellkilling action of HIV-1
• Protection vs. cellular toxicity (TI)
Extract Compound
Bioassay-guided fractionation
• Chromatographic separation of crude extract into
fractions
• Test each fraction, further separate most active
fractions until pure compound is obtained
• NCI chemists isolate calanolide A as anti-HIV
principle of Calophyllum lanigerum extract
cf. J.Med.Chem.35: 2735, 1992.
Calanolide A
O
Structure determined by NMR:
O
O
OH
Other related compounds
isolated with lesser activity
O
Mechanism of action
• Calanolide A found to inhibit HIV reverse
transcriptase in unique fashion (NNRTI)
• Binds to enzyme at site distinct from TIBO
and other NNRTIs
• Development of drug resistance – mutant
enzyme resistant to AZT is more sensitive
to calanolide A cf. J.Virol. 67: 2412, 1993.
• T139I mutant
resistant to
calanolide
Supply of compound problematic
• Recollection attempt – tree had been logged
• Other collections had only <0.1 % calanolide A
• Taxonomy of Calophyllum difficult – new
species & varieties recognized
• Related compounds considered as drug
development leads
Lead compound synthesized
• SmithKline scientists isolate
inophyllums from giant African
snail as HIV RT inhibitors
cf. J.Med.Chem. 36: 4131, 1993.
• traced to Calophyllum in diet
• Calanolide superior to inophyllums
• Total synthesis of calanolide A
achieved by SmithKline
O
O
O
OH
O
Plant sources identified
• Latex of Calophyllum species contains large
quantities of calanolides ~5%
• Tap like rubber tree – renewable resource
• Different species of Calophyllum surveyed for
calanolide content
cf. J.Nat.Prod.61: 1252, 1998.
Calanolide licensed
• Medichem Co.
• Partner with Sarawak (Malaysia) gov’t.
“Sarawak Medichem Pharmaceuticals”
• access to Malaysian resources
Preclinical development
• formulation – how to administer orally
• pharmacokinetics – how fast do blood
levels rise and fall in body (HPLC: rat, dog)
• distribution – to what organs
– crosses blood-brain barrier
• toxicology – negative effects
Hollow fiber HIV model
• Hollow fibers containing HIV-infected cells
implanted i.p. or s.c. in mice
• Calanolide A administered oral or parenteral
• Fibers removed and cells assayed for HIV
cytopathicity
• Calanolide blocked HIV cytopathicity and was
synergistic with AZT
cf. Bioorg.Med.Chem.Lett. 9: 133, 1999.
Calanolide A Phase I trials
safety in humans
Phase IA (1998):
47 healthy adults, US
single oral doses up to 600 mg
higher blood levels than animal studies
predicted
side effects – dizziness, oily aftertaste,
headache, belching, nausea
Phase I trials, cont’d.
Phase IB (1999):
43 patients, HIV-infected asymptomatic
randomized, placebo-controlled, double- blind
adverse effects – mild to moderate, transient
nausea, dyspepsia and headache
no drug resistance
modest viral load reduction –0.8 log
Asian patients had drug-related toxicity – fever &
rash
Further trials
Clinical trial evaluated the therapy's safety and
pharmacokinetics (n=48)
Complete March 2003
Calanolide A in combination therapy for HIV
Evaluated the therapy's effect on pharmacokinetic
enhancement and safety. Results confirmed that the
combination therapy was effective in increasing the
blood levels of calanolide in human volunteers.
Additionally, no serious adverse events were noted in
any subjects and the small number of adverse events
observed were similar to those previously associated
with the drug.
A new wrinkle
• Calanolide A also possesses activity against
Mycobacterium tuberculosis.
• No other anti-HIV agent, either in development or
approved, possesses this dual therapeutic
capability.
• Since patients with HIV/AIDS have weakened
immune systems and are more susceptible to
contracting TB compared to healthy individuals,
the dual therapeutic properties of (+)-Calanolide A
make it a valuable therapeutic agent to this patient
group and will help to distinguish it from other
agents in its class.
Timeline
• Discovery - 1991
• Patenting & publishing - 1992
• Resupply compound – synthesis 1993
– Natural source 1994
•
•
•
•
Mechanism of action - 1993
Licensing - 1995
Phase I - 1997
Phase II – 2002 ??