Non-Clinical Drug Development Chris H. Takimoto, MD, PhD Institute for Drug Development San Antonio Cancer Institute University of Texas Health Science

Non-Clinical Drug Development

Chris H. Takimoto, MD, PhD

Institute for Drug DevelopmentSan Antonio Cancer Institute

University of Texas Health Science CenterSan Antonio, TX

Drug Development

• Drug discovery & screening

• Non-clinical development

• Animal scale up

• Phase I studies

• Phase II studies

• Phase III studies

Specific examples from anticancer drug development

Overview of Anticancer Drug Development

IND NDA

Chemical Synthesis and Formulation Development

Animal Models for Efficacy

Assay Development

Animal PK and PD

Dose Escalation and Initial PK

Proof of Concept and Dose Finding

Large Efficacy Trials with PK Screen

PHASE I PHASE II PHASE III

Pre-Clinical Development Clinical Development

PK/PD Studies in Special Populations

Goals of Non-Clinical Testing of Small Molecule Drugs and Biologicals

• To characterize potential adverse drug effects– Define end organ toxicities– Define reversibility of toxicity

• To characterize pharmacokinetic profile• To characterize beneficial pharmacodynamic

effects– Proof of principle

• To guide safe use in human clinical studies– To determine a safe & reasonable starting dose– Provide monitoring guidelines for the clinical study

• Provide sufficient data to conclude that patients are not exposed to unreasonable risks– Potential for benefit must also exist

Oncology drug development is changing in the new era of targeted cancer therapies

Conventional Wisdom in the New Age of Modern Drug Development

• Targeted therapies are unique and distinct from classic cytotoxic chemotherapies– Classical chemotherapy = poisons

• All new agents entering the clinical have well defined molecular targets

• Conventional clinical study designs are outdated, outmoded, and poorly-suited to develop targeted therapies

• Biomarkers rule!

What are Targeted Therapies?The NCI’s Definition

• From the NCI’s internet fact sheet – Targeted cancer therapies use drugs that

block the growth and spread of cancer. – They interfere with specific molecules involved

in carcinogenesis … and tumor growth. – Because scientists call these molecules

“molecular targets,” these therapies are sometimes called “molecularly-targeted therapies.”

– By focusing on molecular and cellular changes that are specific to cancer, targeted cancer therapies may be more effective than current treatments and less harmful to normal cells.

(http://www.cancer.gov/cancertopics/factsheet/Therapy/targeted)(http://www.cancer.gov/cancertopics/factsheet/Therapy/targeted)

Poster Child for Targeted Therapies

• Prototypical targeted agent is imatinib• A selective inhibitor of BCR-Abl tyrosine

kinase in CML or c-Kit in GIST

Imatinib (Gleevec™)Imatinib (Gleevec™)Imatinib (Gleevec™)Imatinib (Gleevec™)N

N

NHN

HN

N

N

O CH3SO3H

Imatinib (Gleevec™)

Chronic Myelogenous Leukemia and the Ph Chromosome

• Abnormal Philadelphia (Ph) chromosome identified in most patients with chronic myelogenous leukemia (CML)– Identified in over 90% of CML, 20% of adult ALL and 5%

of pediatric ALL patients• Piece of chromosome 9 is abnormally linked to

chromosome 22– 9:22 translocation

• c-Abl, the cellular homologue of the transforming retrovirus oncogene (v-Abl), is located on chromosome 9– Activation of c-Abl signals the cell to proliferate and

grow

Targeted Therapy with Imatinib

• Imatinib is a potent inhibitor of the BCR-Abl and the c-Kit tyrosine kinases

• Generates marked growth inhibition of CML cells and Gastrointestinal Stromal Cell Tumors (GIST)

• Early Phase I data in CML– Hematological response in 98% and

cytogenetic remissions in 13% of patients treated in Phase I

• Substantial single agent activity in GIST tumors

Targeted Therapies Targeted Therapies & Preclinical Development& Preclinical Development

(adapted from Paoletti 2005)(adapted from Paoletti 2005)

Characteristic Cytotoxic Agents Targeted Agents

Discovery Cell based, empiricalReceptor based screen, rationale

Mechanism Often unknown Basis for screening

Pharmacological Effect Cytotoxic Cytostatic

Specificity Non-selective Selective

Dose and schedulePulsed, cyclical at MTD

Continuous, at tolerable dose

Targeted Therapies & Phase I Trials

(adapted from Paoletti 2005)

Characteristic Cytotoxic Agents Targeted Agents

Objectives PK, MTDOptimal biological dose (OBD), PK, PK-PD

Disease All typesAll types or target bearing

DoseToxicity-guided escalation

Biomarker-guided escalation

Endpoints Toxicity, MTD, PK Target inhibition, OBD, PK

DesignDose escalation in small cohorts

Dose escalation to target inhibition

Components of Non-Clinical Drug Development

1. In vitro studies: Cell lines, cell-free systems (drug screening)

2. Drug formulation 3. Chemistry, Manufacturing, and

Controls: Drug supply & quality4. In vivo efficacy studies: Animal

models and proof of principle5. Non-clinical safety studies

In Vitro Study Goals: Define the Drug’s Pharmacology

• Molecular mechanism of action and specific drug targets

• Molecular pharmacology

• Determinants of response

• Intracellular pharmacodynamics

• Mechanisms of drug resistance

In Vitro Study Systems

• Cell-free assay for specific molecular effects– Enzyme inhibition, receptor blockade,

etc.

• Yeast-based screening in genetically defined target

• Mammalian cell lines: (murine, human, etc.)

Preclinical PharmacologyIn Vitro Studies of Cancer Agents (1)

• Define anticancer effects– Growth inhibition, differentiation, apoptosis,

etc

• Impact on defined biochemical and molecular pathways– RNA, DNA and protein biosynthesis,

signaling kinases, etc

• Spectrum of antitumor activity– Human tumor cell lines

Preclinical PharmacologyIn Vitro Studies of Cancer Agents (2)

• Cellular uptake and membrane transport– MDR, MRP, etc

• Mechanisms of resistance• In vitro drug metabolism

– P450 isoenzymes

• Effects on hERG channels (prolonged QT interval risk)

• Preliminary protein binding studies






Drug Supply and Formulation

• Drug supply: bulk chemical synthesis, natural product isolation, etc.

• Good Manufacturing Practice (GMP) guidelines for pharmaceutical product manufacturing

• Formulation for clinical delivery of drug: vehicles for intravenous or other routes of administration

Drug Supply Issues

• Paclitaxel source from the bark and wood of the Pacific Yew tree

• Early drug supply limited the amount available for initial clinical trials

• Newer semisynthetic production from the needles of the Yew tree (renewable)

Drug Formulation Issues

• Poor water solubility of natural products

• Paclitaxel formulation in Cremophore EL™ (increased toxicity?)

• Camptothecin derivatives formulated in a dimethylacetamide, polyethylene glycol and phosphoric acid vehicle– Later formulated as a lipid colloidal dispersion






In Vivo Study Goals:Animal Models

• Efficacy: Proof of therapeutic principle

• Toxicology: Toxicity profile

• Practical Issues:– Animal pharmacokinetics and

pharmacodynamics– Starting dose and schedule for clinical

trials

Animal ModelsProof of Principle

• Animal screening is too expensive for routine use

• Efficacy in animal models of specific disease states occurs after in vitro studies

• Evaluation of therapeutic index– Toxicity versus efficacy

Ideal Animal Model

• Validity

• Selectivity

• Predictability

• Reproducibility

“There is no perfect tumor model”

Endostatin: An Endogenous Inhibitor of Angiogenesis and Tumor Growth

O'Reilly et al, Cell 88:277-285 (1997)

Animal Models in Cancer

• Spontaneous tumors– Idiopathic– Carcinogen-induced– Transgenic/gene knockout animals: p53, RB,

etc• Transplanted tumors

– Animal tumors: Lewis lung, S180 sarcoma, etc– Human tumor xenografts: human tumor lines

implanted in immunodeficient mice (current NCI standard in vivo efficacy testing system)

– Human tumors growing in vivo in implantable hollow fibers

Human Tumor Xenografts

• Athymic “nude”mice developed in 1960’s• Mutation in nu gene on chromosome 11• Phenotype: retarded growth, low fertility,

no fur, immunocompromised– Lack thymus gland, T-cell immunity

• First human tumor xenograft of colon adenocarcinoma by Rygaard & Poulson, 1969

Athymic Nude Mice

Murine Xenograft Sites

• Subcutaneous tumor (NCI method of choice) with IP drug administration

• Intraperitoneal• Intracranial• Intrasplenic• Renal subcapsule• Site-specific (orthotopic) organ

inoculation

Xenograft Study Endpoints

• Toxicity Endpoints:– Drug related death– Net animal weight loss

• Efficacy Endpoints– Clonogenic assay– Tumor growth assay (corrected for tumor

doubling time)– Treated/control survival ratio– Tumor weight change

Xenograft Tumor Weight Change

• Tumor weight change ratio (used by the NCI in xenograft evaluation)

• Defined as: treated/control x 100%• Tumor weight in mg = (a x b2)/2

– a = tumor length– b = tumor width

• T/C < 40-50% is considered significant

Xenograft Advantages

• Many different human tumor cell lines transplantable

• Wide representation of most human solid tumors

• Allows for evaluation of therapeutic index• Good correlation with drug regimens

active in human lung, colon, breast, and melanoma cancers

Xenograft Disadvantages

• Brain tumors difficult to model• Different biological behavior, metastases rare

– Survival not an ideal endpoint: death from bulk of tumor, not invasion

• Shorter doubling times than original growth in human

• Less necrosis, better blood supply• Difficult to maintain animals due to infection risks• Host directed therapies (angiogenesis, immune

modulation) may not be applicable– Human vs. murine effects

Other Animal Models

• Orthotopic animal models: Tumor cell implantation in target organ– Metastatic disease models

• Transgenic Animal Models– P53 or other tumor suppressor gene

knockout animals– Endogenous tumor cell development– May be of high value for mAb therapies

Non-Clinical Efficacy TestingThe FDA Perspective

(J. Leighton, FDA ODAC Meeting, March 13, 2006)

• Pharmacological activity assessed by models of disease are generally of low relevance to safety (IND) and efficacy (NDA) decisions– Efficacy in vivo and in vitro from non-clinical studies may

not dependably predict clinical efficacy• Heterogeneity of disease• Interspecies differences in ADME• Role of immune system

• Pharmacology studies are useful for:– Assessing an appropriate schedule (daily, weekly, q3wks)– Justification for a drug combination– Understanding effect at a molecular target

• Examine receptor specificity• Identifying and evaluating biomarkers






Non-Clinical Safety Studies

• Safety pharmacology

• Toxicokinetics & pharmacokinetic studies

• Single dose toxicity studies

• Repeated dose toxicity studies

Safety Pharmacology

• Assessment of drug on vital functions• Examples:

– Cardiovascular: heart rate, BP, ECG, QT interval

– Central nervous system: locomotor activity, coordination, proconvulsive effects, analgesic effects

– Respiratory system: respiratory rate, tidal and minute volumes

• Should complete prior to FIH studies• May be separate or a component of

toxicity studies

Pharmacokinetic & Toxicokinetic Studies

• Analytic assay development and testing

• Preclinical PK/PD efficacy and toxicity relationships

• Initial drug formulation testing• Testing of different schedules and

routes of administration• Animal ADME

Non-Clinical Toxicology Studies• GLP Toxicology is expected• Use the clinical schedule, route, and formulation• Single dose acute toxicity studies required in 2

mammalian species prior to FIH studies– Classically rat and dog for small molecules– Non-human primates for biologicals

• Repeat dose toxicology required for anticipated duration of clinical use for most non-oncology agents– 3 mo. toxicology for ≤ 3 mo. clinical study

• Recommendations for agents used in the treatment of advanced cancer differs

Expected Toxicology Testing for Phase I Oncology Drug Studies


Clinical Schedule Preclinical study schedule *

Every 21 d Single dose study

Every 14 d 2 doses, 14 d apart

Weekly x 3, week off Weekly x 3

Daily x 5, break Daily x 5

Continuous daily Daily for 28 days

* Study schedule does not include a a recovery period

-- 28 day toxicology is generally sufficient for DRUG trials extending beyond 28 days

Non-Clinical Toxicology Studies For Oncology Drug Combinations• May not be necessary for testing in

advanced cancer patients• May exclude if:

– No PK, PD, or metabolic interactions anticipated

– Drugs are not packaged as a combination

– All components well studied individually

Single Dose Toxicity Studies

• Dose escalation study may be an alternative to a single dose design– Dose range should include maximally

tolerated dose (MTD) and no adverse effect level (NOAEL)

• Standard design– Early sacrifice at 24 to 48 hr and after 14

days

Repeated Dose Toxicity Studies

• Duration of repeated dose studies related to duration of anticipated clinical use– Use same schedule and duration– Typically 14-28 days– Should include recovery group

• Use can support repeat dose clinical studies

Non-Clinical Toxicology Ongoing Endpoints

• Ongoing– Clinical signs, behavior– Body weights and food consumption– Clinical pathology (in larger species)

• Hematology• Chemistry panels

– Toxicokinetics• End of Study

– Macroscopic changes at necropsy– Organ weights– Histopathology of all organs

Other Toxicology Studies

• Local tolerance studies– If warranted by route of administration

• Genotoxicity studies

• Reproductive Toxicity studies

• Carcinogenicity studies

Genotoxicity studies

• General– Normally done prior to FIH studies, but not required

prior to phase I studies in oncology patients– Standard battery of genotoxicity tests required prior to

initiation of phase II• Specific genotoxicity studies

– In vitro bacterial reverse mutation assays: Ames test, point mutation test

– In vitro chromosome damage tests in mammalian cells: metaphase cell analysis, murine lymphoma gene mutation assays

– In vivo chromosomal damage assays: rodent micronucleus tests

Reproductive Toxicity Studies• Men

– May include in Phase I/II after relevant repeated dose toxicity studies

– Male fertility study should be completed prior to initiation of Phase III

• Women not of childbearing potential– May include in clinical trials after relevant repeated dose

toxicity studies• Women of childbearing potential

– May include in carefully monitored early studies with precautions

– Fertility and embryo-fetal toxicity studies should be completed prior to entry of women into phase III trials

• Pregnant women– All reproductive toxicity and genotoxicity studies must be

completed prior to entry of these women in trials

Carcinogenicity studies

• Usually not needed prior to clinical trial initiation

• Not needed in advanced cancer indications

Preclinical ToxicologyGoals

• Estimate a “safe” starting dose for phase I studies

• Determine the toxicity profile for acute and chronic administration

• NCI guidelines recommend single dose and multidose toxicity in two species (one non-rodent)

• Historical guidelines are 1/10 the LD10 in mice – Death, as an endpoint no longer required

Current FDA Approach to Starting Doses

• Starting dose of 1/10 the dose causing severe toxicity (or death) in 10% of rodents (STD10) on mg/m2 basis

• Provided the same dose causes no severe irreversible toxicity in a non-rodent species (usually dogs)

• If irreversible toxicities are seen, then 1/6 of the highest dose tested in non-rodents that does not cause severe, irreversible toxicity– Occasionally, species specific difference may mandate

the use of alternative species for selection of starting dose

Determine dose severely toxic to 10% of rodents (STD10)

Convert from mg/kg to mg/m2

Mouse x 3; Rat x 6; Guinea-pig x 7.7Hamster x 4.1

Determine non-rodent HighestNon-Severely Toxic Dose

(HNSTD)

Is rodent aninappropriate species?

(biochem, ADME, target, etc)

Is 1/10rodent STD10 (mg/m2)

severely toxic tonon-rodents?

Is non-rodent inappropriate?

Convert from mg/kg to mg/m2

Dog x 20; Monkey x 10Rabbit x 11.6

Start Dose =1/6Non-Rodent HNSTD

NO YES

YES

YESNO

Start Dose =1/10Rodent STD10

NO

Non-Clinical Drug Safety Testingfor Summary of the FDA Perspective


• Conduct 2 pivotal toxicology studies using the same schedule, formulation, and route as the proposed clinical trial– Conduct a rodent study that identifies life-threatening

doses– Conduct a non-rodent study that confirms non-life

threatening doses have been identified• Studies of 28 days should be provided for continuous

administration• Studies for one or several administrations, depending on

the schedule for intermittent schedules• Provide full histopathology in one of these studies

– Conduct other studies as needed

Non-Clinical Drug Safety TestingSummary of the FDA Perspective(J. Leighton, FDA ODAC Meeting, March 13, 2006)

• Multiple cycles/continuous treatment generally acceptable, assuming acceptable safety profile in the non-clinical setting

• Pre-IND meeting with sponsors are encouraged to discuss problem areas and provide alternative pathways to initiation of the phase I trial

• Most potential clinical holds resolved through discussion with sponsor

• Guidelines for biologicals (monoclonal antibodies, etc) are in preparation but may differ from small molecule recommendations

An Excellent Resource for Anticancer Drugs(DeGeorge et al Cancer Chemother Pharmacol 1998;41:173)

Plus numerous FDA guidances at http://www.fda.gov/cder/Guidance

Monoclonal Antibody (mAb) Therapeutics

• Targeted mAb are distinct from small molecule therapeutics– Explosion in popularity– Higher approval rates in oncology (~21% vs. <5%)

• High specificity, less off target risk• Long t1/2 (10-21 days)• Novel targets that are difficult or impossible to

modulate by small molecules• Flexible bioengineered design

– Modulation of functional domains

Non-Clinical Toxicology for mAb Therapies

• mAb present major safety challenges• Safety toxicology studies in primates

– Old world primates most common– May exceed primate toxicology resources– Chimpanzees in rare specialized cases

• Primate toxicology may still not predict human effects– TGN1412 anti CD28 super agonist causes non-specific broad

T-cell activation in humans with catastrophic consequences

• Transgenic rodents engineered to express human target may be selectively employed (knock out/knock in animals)

• Surrogate mAb (mouse equivalent) toxicity and efficacy studies to support clinical studies

Starting Doses for Biological Therapies

• Historically, some fraction of the no adverse event level (NOAEL)

• If species specific differences preclude precise dose calculations, then…

• Consider estimations of receptor occupancy, cellular dose response studies from best available models to estimate a Minimum Anticipated Biological Effect Level (MABEL)

• Recommendations for biological therapies are in evolution

An Example of a Phase I study of a Targeted Therapy that Incorporates Biomarkers Developed in Preclinical

Development

AEE788, A Dual EGFR & AEE788, A Dual EGFR & VEGFR Targeting AgentVEGFR Targeting AgentAEE788, A Dual EGFR & AEE788, A Dual EGFR & VEGFR Targeting AgentVEGFR Targeting Agent

• Oral receptor tyrosine kinase inhibitor Oral receptor tyrosine kinase inhibitor – A “dirty” kinase inhibitorA “dirty” kinase inhibitor

• 7H-pyrrolo[2,3-d] pyrimidine derivative7H-pyrrolo[2,3-d] pyrimidine derivative• Inhibits EGFR and ErbB-2 receptor tyrosine kinases with Inhibits EGFR and ErbB-2 receptor tyrosine kinases with

ICIC5050’s of 2-6 nM’s of 2-6 nM• Also inhibits multiple other kinasesAlso inhibits multiple other kinases

• Oral receptor tyrosine kinase inhibitor Oral receptor tyrosine kinase inhibitor – A “dirty” kinase inhibitorA “dirty” kinase inhibitor

• 7H-pyrrolo[2,3-d] pyrimidine derivative7H-pyrrolo[2,3-d] pyrimidine derivative• Inhibits EGFR and ErbB-2 receptor tyrosine kinases with Inhibits EGFR and ErbB-2 receptor tyrosine kinases with

ICIC5050’s of 2-6 nM’s of 2-6 nM• Also inhibits multiple other kinasesAlso inhibits multiple other kinases

N

N

HN

NH

N

N

In Vitro AEE788 PharmacologyIn Vitro AEE788 PharmacologyIn Vitro AEE788 PharmacologyIn Vitro AEE788 Pharmacology

KinaseKinase ICIC5050 (uM) (uM)

EGFREGFR 0.0020.002

ErbB2ErbB2 0.0060.006

KDRKDR 0.0770.077

HER4HER4 0.0590.059

C-AblC-Abl 0.0520.052

C-SrcC-Src 0.0610.061

RETRET 0.740.74

KinaseKinase ICIC5050 (uM) (uM)

EGFREGFR 0.0020.002

ErbB2ErbB2 0.0060.006

KDRKDR 0.0770.077

HER4HER4 0.0590.059

C-AblC-Abl 0.0520.052

C-SrcC-Src 0.0610.061

RETRET 0.740.74

KinaseKinase IC IC5050 (uM) (uM)

C-KitC-Kit 0.790 0.790

C-MetC-Met 2.902.90

Flk, TekFlk, Tek >2>2

IGF-1RIGF-1R >2>2

PKC-aPKC-a >10>10

CDK1/2CDK1/2>10>10

KinaseKinase IC IC5050 (uM) (uM)

C-KitC-Kit 0.790 0.790

C-MetC-Met 2.902.90

Flk, TekFlk, Tek >2>2

IGF-1RIGF-1R >2>2

PKC-aPKC-a >10>10

CDK1/2CDK1/2>10>10

AEE788 Study DesignAEE788 Study DesignAEE788 Study DesignAEE788 Study Design

• Three center N. American/European studyThree center N. American/European study– C.H. Takimoto, IDD/CTRC, San AntonioC.H. Takimoto, IDD/CTRC, San Antonio– J. Baselga, Vall d’Hebron, Barcelona, Spain J. Baselga, Vall d’Hebron, Barcelona, Spain – A.T. van Oosterom, Catholic University Leuven, BelgiumA.T. van Oosterom, Catholic University Leuven, Belgium

• Dose escalation design of AEE788 orally on a daily Dose escalation design of AEE788 orally on a daily dose schedule in advanced cancer patientsdose schedule in advanced cancer patients– Standard adult phase I patient populationStandard adult phase I patient population– 3-6 patients per dose level allowed3-6 patients per dose level allowed

• Endpoints:Endpoints:– Determine the MTD and DLT of AEE orally on a daily dose Determine the MTD and DLT of AEE orally on a daily dose

scheduleschedule– Characterize drug pharmacokineticsCharacterize drug pharmacokinetics– Extensive pharmacodynamic assessmentsExtensive pharmacodynamic assessments

• Three center N. American/European studyThree center N. American/European study– C.H. Takimoto, IDD/CTRC, San AntonioC.H. Takimoto, IDD/CTRC, San Antonio– J. Baselga, Vall d’Hebron, Barcelona, Spain J. Baselga, Vall d’Hebron, Barcelona, Spain – A.T. van Oosterom, Catholic University Leuven, BelgiumA.T. van Oosterom, Catholic University Leuven, Belgium

• Dose escalation design of AEE788 orally on a daily Dose escalation design of AEE788 orally on a daily dose schedule in advanced cancer patientsdose schedule in advanced cancer patients– Standard adult phase I patient populationStandard adult phase I patient population– 3-6 patients per dose level allowed3-6 patients per dose level allowed

• Endpoints:Endpoints:– Determine the MTD and DLT of AEE orally on a daily dose Determine the MTD and DLT of AEE orally on a daily dose

scheduleschedule– Characterize drug pharmacokineticsCharacterize drug pharmacokinetics– Extensive pharmacodynamic assessmentsExtensive pharmacodynamic assessments

Dose Levels and DLTs During Cycle 1Dose Levels and DLTs During Cycle 1(Baselga ASCO 2005)(Baselga ASCO 2005)

Dose Levels and DLTs During Cycle 1Dose Levels and DLTs During Cycle 1(Baselga ASCO 2005)(Baselga ASCO 2005)

Dose Level, mgDose Level, mg EnrolledEnrolled Pts with DLT, nPts with DLT, n DLT (n)DLT (n)

2525 55 00 ----

5050 66 00 ----

100100 55 00 ----

150150 55 00 ----

225225 77 00 ----

300300 77 00 ----

400400 77 00 ----

450450 77 00 ----

500500 66 22 Gr 3 diarrhea (2)Gr 3 diarrhea (2)

550550 99 22 Gr 3 diarrhea (2)Gr 3 diarrhea (2)

TotalTotal 6464 44

AEE788 Skin RashAEE788 Skin RashAEE788 Skin RashAEE788 Skin Rash

• Skin rash Skin rash – Any grade = 42.7%Any grade = 42.7%– Gr 3 or 4 = 0%Gr 3 or 4 = 0%

• Dry skin, fine rash Dry skin, fine rash seen at lower dose seen at lower dose levelslevels

• Pustular macular Pustular macular papular skin rash papular skin rash seen at higher dose seen at higher dose levels (>225 mg/d)levels (>225 mg/d)

• Skin rash Skin rash – Any grade = 42.7%Any grade = 42.7%– Gr 3 or 4 = 0%Gr 3 or 4 = 0%

• Dry skin, fine rash Dry skin, fine rash seen at lower dose seen at lower dose levelslevels

• Pustular macular Pustular macular papular skin rash papular skin rash seen at higher dose seen at higher dose levels (>225 mg/d)levels (>225 mg/d)

AEE788 Other ToxicitiesAEE788 Other ToxicitiesAEE788 Other ToxicitiesAEE788 Other Toxicities

• Grade 3 diarrhea (10 pts)Grade 3 diarrhea (10 pts)

• Grade 3 fatigue (5 pts)Grade 3 fatigue (5 pts)

• Grade 3 anorexia (4 pts)Grade 3 anorexia (4 pts)

• Grade 3 hyperbilirubinemia (3 pts)Grade 3 hyperbilirubinemia (3 pts)

• No evidence of cardiac toxicity or QTc No evidence of cardiac toxicity or QTc changes in 2811 EKGs in 96 ptschanges in 2811 EKGs in 96 pts

• Grade 3 diarrhea (10 pts)Grade 3 diarrhea (10 pts)

• Grade 3 fatigue (5 pts)Grade 3 fatigue (5 pts)

• Grade 3 anorexia (4 pts)Grade 3 anorexia (4 pts)

• Grade 3 hyperbilirubinemia (3 pts)Grade 3 hyperbilirubinemia (3 pts)

• No evidence of cardiac toxicity or QTc No evidence of cardiac toxicity or QTc changes in 2811 EKGs in 96 ptschanges in 2811 EKGs in 96 pts

Delayed Onset Reversible Hepatic Delayed Onset Reversible Hepatic Transaminase ElevationTransaminase Elevation

Delayed Onset Reversible Hepatic Delayed Onset Reversible Hepatic Transaminase ElevationTransaminase Elevation

• Reversible grade 3 / 4 elevations of Reversible grade 3 / 4 elevations of AST/ALT at doses AST/ALT at doses ≥≥300 mg seen in 12 pts300 mg seen in 12 pts

• Median onset of 99 days (after 4 cycles)Median onset of 99 days (after 4 cycles)

• Observed in presence and absence of liver Observed in presence and absence of liver metastasesmetastases

• Total bilirubin generally unaffectedTotal bilirubin generally unaffected

• Dose and duration of treatment dependentDose and duration of treatment dependent– Dosing at Dosing at ≥≥300 mg/d for 4+ cycles is 300 mg/d for 4+ cycles is

problematicproblematic

• Reversible grade 3 / 4 elevations of Reversible grade 3 / 4 elevations of AST/ALT at doses AST/ALT at doses ≥≥300 mg seen in 12 pts300 mg seen in 12 pts

• Median onset of 99 days (after 4 cycles)Median onset of 99 days (after 4 cycles)

• Observed in presence and absence of liver Observed in presence and absence of liver metastasesmetastases

• Total bilirubin generally unaffectedTotal bilirubin generally unaffected

• Dose and duration of treatment dependentDose and duration of treatment dependent– Dosing at Dosing at ≥≥300 mg/d for 4+ cycles is 300 mg/d for 4+ cycles is

problematicproblematic

AEE788 Efficacy DataAEE788 Efficacy DataAEE788 Efficacy DataAEE788 Efficacy Data

• 83 Patients treated with doses up to 550 83 Patients treated with doses up to 550 mgmg– One PR in angiosarcoma at 400 mg now One PR in angiosarcoma at 400 mg now

completed 5 cyclescompleted 5 cycles– 36 of 83 pts (43%) with stable disease 36 of 83 pts (43%) with stable disease

beyond 2 cyclesbeyond 2 cycles– Median number of cycles that patients Median number of cycles that patients

remained on study is 2 (range 0.5-14)remained on study is 2 (range 0.5-14)

• 83 Patients treated with doses up to 550 83 Patients treated with doses up to 550 mgmg– One PR in angiosarcoma at 400 mg now One PR in angiosarcoma at 400 mg now

completed 5 cyclescompleted 5 cycles– 36 of 83 pts (43%) with stable disease 36 of 83 pts (43%) with stable disease

beyond 2 cyclesbeyond 2 cycles– Median number of cycles that patients Median number of cycles that patients

remained on study is 2 (range 0.5-14)remained on study is 2 (range 0.5-14)

Planned PD AssessmentsPlanned PD AssessmentsPlanned PD AssessmentsPlanned PD Assessments

• Skin biopsies in 53 ptsSkin biopsies in 53 pts• Vascular IHC analyses in 32 ptsVascular IHC analyses in 32 pts• Tumor biopsy IHC data from 15 ptsTumor biopsy IHC data from 15 pts

• Skin biopsies in 53 ptsSkin biopsies in 53 pts• Vascular IHC analyses in 32 ptsVascular IHC analyses in 32 pts• Tumor biopsy IHC data from 15 ptsTumor biopsy IHC data from 15 pts

DailyAEE788

Skin (wound) biopsy day -1

Skin biopsy day +22

w1 w2 w3 w4 w5

Skin (wound) biopsy day +29

Skin biopsy day-8

Cycle 1 Cycle 2

Tumor biopsy day -14 to -1

Tumor biopsy day +28

Biopsy samples were evaluated by immunohistochemistry (IHC) and scored by Hscore. Hscore = (% faint stained cells) + (% moderate stained cells)*2 + (% strong stained cells)*3. Ki67 was scored by % positive cells --Baselga et al, ASCO 2005

Results: Skin (basal epidermis) p-EGFRResults: Skin (basal epidermis) p-EGFR

25mg 50mg 100mg 150mg

90

60

88

59

75 7573 73

0

30

60

90

120

150

Hscore basal pEGFR day -7 Hscore basal pEGFR day +22

75

45

120

105105

60

48

30

0

30

60

90

120

150


105

15

75

15

7875

12

30

60

42

0

20

40

60

80

100

120


60

15

30

75

60

42

0

30

60

90

120

150


225mg 300mg 400mg 550mg

75

42

72

30

17

30

15

54

28

15

0

30

60

90

120

150


72

15

54

12

72

24

36

6

60

30

0

30

60

90

120

150


66

0

45

0

30

60

90

120

150


48

0

30

0

72

0

30

0

30

60

90

120

150


--Baselga et al, ASCO 2005

Pharmacodynamic ModelingPharmacodynamic ModelingPharmacodynamic ModelingPharmacodynamic Modeling


Tumor Biopsy IHC Results (n=15)Tumor Biopsy IHC Results (n=15)Tumor Biopsy IHC Results (n=15)Tumor Biopsy IHC Results (n=15)pEGFR pMAPK pAkt Ki67

25 mg25 mg

Pre-RxPre-Rx

During RxDuring Rx

25 mg25 mg

Pre-RxPre-Rx

During RxDuring Rx

550 mg550 mg

Pre-RxPre-Rx

During RxDuring Rx

550 mg550 mg

Pre-RxPre-Rx

During RxDuring Rx


PD Tumor Marker ChangesPD Tumor Marker ChangesPD Tumor Marker ChangesPD Tumor Marker Changes

Maximal pEGFR SuppressionMaximal pEGFR Suppression


Pharmacodynamic FindingsPharmacodynamic FindingsPharmacodynamic FindingsPharmacodynamic Findings

• Inhibition of molecular targets was dose and serum concentration Inhibition of molecular targets was dose and serum concentration dependent with significant variablitydependent with significant variablity– Active concentration = AEE788 (parent) + AQM674 (active metabolite)Active concentration = AEE788 (parent) + AQM674 (active metabolite)

• Tumor pEGFR inhibition (ICTumor pEGFR inhibition (IC5050 18 nM) agrees with A431 cell line data 18 nM) agrees with A431 cell line data (IC(IC5050 = 11 nM) = 11 nM)

• Skin PD PotencySkin PD Potency– TestTest ID80ID80

– pEGFR/pMAPKpEGFR/pMAPK 225-250 mg225-250 mg

– Ki67Ki67 50-100 mg50-100 mg

• Tumor PD Potency (greater than skin)Tumor PD Potency (greater than skin)– TestTest ID80ID80

– pEGFRpEGFR 150 mg150 mg

– pAktpAkt 100-150 mg100-150 mg

• Optimal biological dose may be ~250 mg(?)Optimal biological dose may be ~250 mg(?)

• Inhibition of molecular targets was dose and serum concentration Inhibition of molecular targets was dose and serum concentration dependent with significant variablitydependent with significant variablity– Active concentration = AEE788 (parent) + AQM674 (active metabolite)Active concentration = AEE788 (parent) + AQM674 (active metabolite)

• Tumor pEGFR inhibition (ICTumor pEGFR inhibition (IC5050 18 nM) agrees with A431 cell line data 18 nM) agrees with A431 cell line data (IC(IC5050 = 11 nM) = 11 nM)

• Skin PD PotencySkin PD Potency– TestTest ID80ID80

– pEGFR/pMAPKpEGFR/pMAPK 225-250 mg225-250 mg

– Ki67Ki67 50-100 mg50-100 mg

• Tumor PD Potency (greater than skin)Tumor PD Potency (greater than skin)– TestTest ID80ID80

– pEGFRpEGFR 150 mg150 mg

– pAktpAkt 100-150 mg100-150 mg

• Optimal biological dose may be ~250 mg(?)Optimal biological dose may be ~250 mg(?)

AEE788 Phase I Study AEE788 Phase I Study ConclusionsConclusions

AEE788 Phase I Study AEE788 Phase I Study ConclusionsConclusions

• Cycle 1 DLT was grade 3 diarrhea despite supportive Cycle 1 DLT was grade 3 diarrhea despite supportive care at 500-550 mg/dcare at 500-550 mg/d

– Other toxicities: fatigue, nausea, rash, anorexia, vomiting, Other toxicities: fatigue, nausea, rash, anorexia, vomiting, stomatitisstomatitis

• Chronic dosing revealed grade 3/4 hepatic transaminitis Chronic dosing revealed grade 3/4 hepatic transaminitis at doses >300 mg after 4 cyclesat doses >300 mg after 4 cycles

• PK/PD analysis suggests dose PK/PD analysis suggests dose ≥≥250 mg may optimally 250 mg may optimally modulate biological target(s)modulate biological target(s)

• Further exploration of daily 250 mg dosing and Further exploration of daily 250 mg dosing and alternative schedules is ongoingalternative schedules is ongoing

– PK/PD biomarker data highly useful in clinical decision making PK/PD biomarker data highly useful in clinical decision making processprocess

• Cycle 1 DLT was grade 3 diarrhea despite supportive Cycle 1 DLT was grade 3 diarrhea despite supportive care at 500-550 mg/dcare at 500-550 mg/d

– Other toxicities: fatigue, nausea, rash, anorexia, vomiting, Other toxicities: fatigue, nausea, rash, anorexia, vomiting, stomatitisstomatitis

• Chronic dosing revealed grade 3/4 hepatic transaminitis Chronic dosing revealed grade 3/4 hepatic transaminitis at doses >300 mg after 4 cyclesat doses >300 mg after 4 cycles

• PK/PD analysis suggests dose PK/PD analysis suggests dose ≥≥250 mg may optimally 250 mg may optimally modulate biological target(s)modulate biological target(s)

• Further exploration of daily 250 mg dosing and Further exploration of daily 250 mg dosing and alternative schedules is ongoingalternative schedules is ongoing

– PK/PD biomarker data highly useful in clinical decision making PK/PD biomarker data highly useful in clinical decision making processprocess

The Clinical Trial Challenge

• We stand at the dawn of the post genomic era when new targets for novel treatments for human cancer are just being discovered and defined

• Basic research is the engine that drives this process

• Clinical researchers have to take these promising agents and test them in the best and most efficient ways possible – Traditional clinical endpoints, and…– Molecular target endpoints in clinical studies

The Challenge!

PreclinicalPharmacology

Traditional animal studies

PK/PDToxicology

Molecular targets

ClinicalPharmacologist

Early Phase IPharmacokinetic

Clinical Trials

Traditional dose and toxicity endpointsTraditional PK/PD

Molecular andbiochemicalendpoints

New Paradigms for Drug Development in the Post Genomic Era

• Expanding role for translational studies in Phase I clinical trials

• Bridge the gap between preclinical pharmacologic studies and early clinical trials

• New molecular and biochemical endpoints are essential for cancer prevention and antimetastatic agents

• This is an exciting time to be developing new anticancer drugs!

New Phase I Paradigms: New Phase I Paradigms: Evolution not Revolution!Evolution not Revolution!New Phase I Paradigms: New Phase I Paradigms:

Evolution not Revolution!Evolution not Revolution!

Documents

Non-Clinical Drug Development Chris H. Takimoto, MD, PhD Institute for Drug Development San Antonio Cancer Institute University of Texas Health Science