Biomarkers in CRC: Are We Any Closer to Our Goal of Personalized Medicine?

Biomarkers in CRC:Are We Any Closer to Our

Goal of Personalized Medicine?

Lee M. Ellis, MD

Depts. of Surgical Oncology and Cancer Biology

U.T. M.D. Anderson Cancer Center

Houston, Texas, USA

Characteristics of a Good Predictive Biomarker

• Consistent across numerous studies, lines of therapy, and numerous drugs within the same class

• Makes biologic sense• Relatively easy assay with rapid turnaround

time• “Yes” or “no” answer

– Objective, not subjective– Not subject to lab-to-lab variation

We Need to Start Thinking in Terms of “Pathways”, Not Individual Molecules

• Systems biology …… focuses on the systematic study of complex interactions in biological systems… (integration instead of reduction) to study them.…..

QuickTime™ and a decompressor

are needed to see this picture.

Wikipedia

HIFLDH/Glut1/VEGF/R

PI3KPTENAkt





Intratumoral mRNA expression of genes involved in angiogenesis and HIF-1 pathway to predict outcome to

VEGFR tyrosine kinase inhibitor (TKI) in patients enrolled in CONFIRM1 and CONFIRM2

P. M. Wilson, D. Yang, M. M. Shi, W. Zhang, C. Jacques, J. C. Barrett, K. Daneneberg, T. Trarbach, G. Folprecht, G. Meinhardt, H. J. Lenz

There Are No Known Predictive Biomarkers for Anti-VEGF Therapy

Proposed Biomarker Outcome

Plasma VEGF Not Predictive

Primary Tissue VEGF

(ISH, IHC)

Not Predictive

Upstream Mediators of VEGF

(ras, raf, P53)

Not Predictive

Other Angiogenic Mediators

(TSP-2)

Not Predictive

Jubb et al. J Clin Oncol 24:217-227, 2006Ince et al. J Natl Cancer Inst. 97(13):981-9, 2005

Holden et al. ASCO. Abstract: 3555, 2005

IFL +/- Bev Trial

Progression-Free Survival

PTK/ZK (n=585) 9.1 moPlacebo (n=583) 7.7 mo

HR 0.89

(95% CI) (0.78, 1.03)

P-value 0.108

100

80

60

40

20

% P

FS

Time since randomization (months)

4 8 12 16 20 24 28 320 36 400

25

50

75

100

0 4 8 12 16 20

Time since randomization, mos

75

FOLFOX4 + PTK/ZKFOLFOX4 + Placebo

PTK/ZK (n=426) 5.6 moPlacebo (n=429) 4.1 mo

HR 0.83

(95% CI) (0.71, 0.98)

P-value 0.026

PF

S %

CONFIRM-1 CONFIRM-2

Analyses in this study done on 42-54 patient tumors/arm.

High LDH and Predictive Value of FOLFOX + PTK/ZK

Overall Survival High LDHProgression Free Survival High LDHCONFIRM1

CONFIRM2

• Premise of Study– LDH regulated by HIF, and serum LDH

may be a predictive marker– Why not investigate genes in the HIF

pathway as predictive markers for PTK/ZK efficacy?

Important Findings in This Study

• Expression levels of genes in a pathway may be associated with potential benefit of the addition of PTK/ZK to FOLFOX– Proof of principle (genomic profiling)

• However, there are divergent results between first line and second line therapy– Did this fail the validation test? or – Is this a reflection of the effect of chemotherapy on

inducible genes?– Small numbers of patients

• Some groups n<50 (sometimes <10)• Less than 10% of overall trial

PFS with PTK/ZK

>HIF-1α 0.021

>LDHA 0.075

>VEGFR2 0.001

CONFIRM 1

Gene p-value Gene p-value

CONFIRM 2

<HIF-1α 0.021

<LDHA 0.031

11.3 v 7.6 mo

9.4 v 3.5 mo 7.6 v 2.7 mo

7.6 v 1.7 mo

8 v 4.1 mo

“The data is the data…you can’t change the data, change the hypothesis”

Josh Fidler

Unexpected results, such as the divergent data in CONFIRM-1 AND CONFIRM-2, provide an

opportunity for the development of new hypotheses regarding potential biomarkers

We must be flexible in our thinking, and be able to alter our hypotheses, and subsequent investigations

i.e. does exposure to chemotherapy effect the HIF pathway?(topotecan can decrease HIF-1: G. Mellilo NCI)

HIF-1 Regulation:

Measuring mRNA May Not Give You the Whole Story

IGFR/EGFR/HER2

PI-3-K

Akt

Transcriptional Regulation(increased mRNA and protein)

HIF-1 mRNAHIF -1 Protein

Hypoxic Regulation(increased protein)

Decreased prolyl hydroxylation and ubiquitination

Decreased proteosomal degradation

HIF -1 Protein

Gene transcription

http://www.srcf.ucam.org/~jrk33/pov/DNA.png

There is Not Much Remaining Interest in PTK/ZK Will findings from this study translate to other drugs in the field

Markers that may be predictive for one TKI, may not be predictive for another

The larger the red circle, the more effective the drug is for the target

Human Kinome Tree

The Non-Specificity of VEGFR Kinase Inhibitors


Ras/EGFR HIF Pathway/

VEGF/R

Consistent across numerous studies and numerous drugs within the same class

Yes, and consistent upon lines of therapy

TBD, not consistent upon lines of therapy

Relatively easy assay with rapid turnaround time

Yes Kind of….

Makes biologic sense Yes ?

“Yes” or “no” answer

Objective, not subjective

Not subject to lab-to-lab variation

Yes No

Is Gene Expression Profiling IN THIS STUDY Any Better

Than Measuring Serum LDH?

Is This Enough to Proceed With a Phase III Trial Selecting for Patients

By Evaluating HIF Pathway?• NO

– Divergent results in CONFIRM-1 and CONFIRM-2 require explanation– There are better drugs with better PK that are already in late phase

clinical trials– However, tissue should be banked on all Phase III trials for study of

potential markers of efficacy of anti-VEGF therapy

• This is “hypothesis generating” data, not validated data• We must continue to search for predictive markers of anti-

VEGF therapy

ASCO 2005 Plenary Session

Here we



• Here we are again….3 years later and the results for CONFIRM-1 have not yet been submitted for publication.• We when enroll patients in clinical trials, they put their trust in us to use their experience to advance our knowledge. We owe it to our patients to publish data from clinical trials in a timely fashion regardless of whether the trial was negative or positive!!!

Disclaimer: This is not a reflection on most of the current authors on this study

Published?

Yes

Yes

No

Yes

Yes

No

Yes

CONFIRM-1





Evaluation of PTEN expression in colorectal cancer (CRC) metastases

(mets) and in primary tumors as predictors of activity of cetuximab plus

irinotecan treatment

F. Loupakis, L. Pollina, I. Stasi, G. Masi, N. Funel,

M. Scartozzi, Petrini I, Santini D, Cascinu S, Falcone A

Activation of Downstream Pathways Activation of Downstream Pathways and EGFR MoAB Resistanceand EGFR MoAB Resistance

18 BMS-ASCO-CRC-Therapy-Ellis

Camp et al, Clin Cancer Res 2005

SrcActPI-3K

AktMut

NucleusNucleus

EGFR MoAB

EGFR

Fak

RasMut

RafMut

MEK

Erk

PTENMut/

Inactivaed

Survival Pathway

40% CRC mutated

Proliferative Pathway

40% CRC mutated

Mut

PTEN Overview

• PTEN is a phosphatase that blocks activation of the Akt/survival pathway– Loss of function of

PTEN is associated with an increase in cell survival signaling

Leslie, Downes Biochem J 2004

Mechanisms of Regulation of PTEN Protein Levels

• Mutation

• Loss of heterozygosity

• Methylation of promoter – decreases transcription

• Certain oncogenes can downregulate transcription

Important Findings in This Study

• PTEN protein expression in primary tumor does not correspond to PTEN expression in mets– PTEN in mets is predictive of response whereas

PTEN in the primary is not

• Kras mutational status in the primary does correlate with mutational status in the metastasis

• The biology of these observations makes sense– Mutational status should not change (Ras)– Methylation status is subject to regulation by the

microenvironment (PTEN)

Logrank Test: p=0.001HR = 0.42 95% CI: 0.17-0.65

PTEN+/KRAS wt

All others

Logrank Test: p=0.005HR = 0,49 95% CI: 0.20-0.75

PTEN+

PTEN-

PTEN/RAS and PFSCan We Better Predict Efficacy with a

Multifactorial Analysis?

Sorry…slides from the investigators..need their permission to use

British Journal of Cancer (2007) 97, 1139-1145.

“Whereas neither EGFR protein/mRNA expression nor gene copy number correlated with cetuximab response, examination of the mutation status of signaling components downstream of EGFR showed that cell lines with activating PIK3CA mutations or loss of PTEN expression (PTEN null) were more resistant to cetuximab than PIK3CA wild type (WT)/PTEN-expressing cell lines”


Ras/EGFR PTEN/EGFR

Consistent across numerous studies and numerous drugs within the same class

Yes Probably

Relatively easy assay with rapid turnaround time

Yes Yes

Makes biologic sense Yes Yes

“Yes” or “no” answer

Objective, not subjective

Not subject to lab-to-lab variation

Yes No (IHC staining)1 2 3

Intensity

1+ 2+ 3+

% +cells

0-2525-50

>50

>4 positive

Predictive Oncology in mCRC: What Have We Learned Today?

VEGF/R Targeting• Gene expression profiling of the HIF pathway may be predictive for efficacy of

PTK/ZK + FOLFOX– “Hypothesis generating” study

• Divergent results in CONFIRM-1 and -2 are confusing

– Investigations of gene expression profiling with other VEGF/R inhibitors are essential in determining the true value of this approach

– We must continue the search for predictors of VEGF/R efficacy

EGFR Targeting• Ras continues to predict for EGFR MoAB efficacy• PTEN levels in metastases predicts for efficacy of cetuximab

– PTEN in primary tumors is not predictive• Proteins that can be regulated by the tumor microenvironment may require biopsy when

determining value as a predictive marker

• Multi-factorial analyses may better predict for efficacy



After ASCO 2008, it is time to update recommendations for CRC!!



Thank You for Your Interest!

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Biomarkers in CRC: Are We Any Closer to Our Goal of Personalized Medicine?