New Strategies and Therapeutic Targets in PDATargets in PDA

Manuel Hidalgo, M.D., Ph.D.g

Multifaceted Biology of PDAgy

M Hidalgo, NEJM 2010

Genomic Diversity of Pancreas Cancer

Jones et al, Science 2008.

Core Gene Pathways in Pancreas Cancer

Jones et al, Science 2008.

PALB2 Mutations

Tischkowitz, CR 2010

Targeting PALB2 Mutations

Villaroel, MCT 2011

Mutational Analysis of Primary Tumor

Human genome reference sequenceHuman genome reference sequence18,655 genes

Capture tumor and normal DNA to enrich for coding exons37,806,033 bases of target sequence s g q

from 165,637 target regions

Sequence tumor and normal DNA usingnext-generation sequencing

15 904 958 898 bases of tumor DNAry A

naly

sis

15,904,958,898 bases of tumor DNA12,445,240,820 bases of normal DNA

Assemble sequence data and compare to target region 34,366,555 bases of target regions with at least 10 readsD

isco

ver

Analyze sequences for potential alterationsAverage of 37 distinct reads at each base in tumor DNAAverage of 28 distinct reads at each base in normal DNA

Compare tumor and normal sequence data to identify tumor-specific mutations

Somatic mutations in 62 genes

alys

is

Bioinformatic analyses of mutated genes

Id tifi ti f Id tifi ti f t t drmat

ic A

na

Identification of cancer genes

Identification of mutatedpathways

Analysis of individual mutationsBio

info

r

Selected Somatic Mutation

Mutation Position

Gene      Symbol

Gene Description Pathway  / Functional GroupTranscript Accession

Mutation   TypeNucleotide      

(genomic)

Nucleotide 

(transcript)

Amino Acid 

(protein)

MEFV Mediterranean fever Inflammatory response CCDS10498.1 chr16:3234255C>A 1759G>T 587V>F Missense

MRM1 mitochondrial rRNA methyltransferase 1 homolog RNA processing CCDS32631.1 chr17:32038218G>C IVS2‐1G>C NA Splice site

MYH10 myosin; heavy chain 10; non‐muscle Cytoskeletal organization CCDS11144.1 chr17:8352644A>C 3074T>G 1025L>W Missense

NES nestin CNS development CCDS1151.1 chr1:154906069T>C 4535A>G 1512K>R Missense

OR13F1 olfactory receptor; family 13; subfamily F; member 1 Sensory perception of smell CCDS35087 1 chr9:106306951A>G 587A>G 196Q>R MissenseOR13F1 olfactory receptor; family 13; subfamily F; member 1 Sensory perception of smell CCDS35087.1 chr9:106306951A>G 587A>G 196Q>R Missense

OR52N1 olfactory receptor; family 52; subfamily N; member 1 Sensory perception of smell CCDS31398.1 chr11:5766168A>C 455T>G 152L>R Missense

PABPC1L poly(A) binding protein; cytoplasmic 1‐like RNA and nucleotide binding CCDS42878.1 chr20:42992784G>T IVS8+3G>T NA Splice site

PAPPA2 pappalysin 2 Cell growth and differentiation CCDS41438.1 chr1:175035873C>A 5184C>A 1728H>Q Missense

PAPSS1 3'‐phosphoadenosine 5'‐phosphosulfate synthase 1 Xenobiotic metabolic process CCDS3676.1 chr4:108795472A>G 929T>C 310L>P Missense

PDE4B phosphodiesterase 4B; cAMP‐specific Cyclic nucletoide concentration  CCDS632.1 chr1:66151609G>T 24G>T 8M>I Missense

PIK3C2A phosphoinositide‐3‐kinase; class 2; alpha polypeptide Phosphatidylinositol signaling CCDS7824.1 chr11:17147055C>G 810G>C 270W>C Missense

PIK3CA phosphoinositide‐3‐kinase; catalytic; alpha polypeptide Phosphatidylinositol signaling CCDS43171.1 chr3:180430545T>G 2726T>G 909F>C Missense

PLD5 phospholipase D family; member 5 Phospholipase activity CCDS1621.1 chr1:240330625T>G 1046A>C 349K>T Missense

PLS3 plastin 3 actin binding CCDS14568.1 chrX:114788511G>A 1678G>A 560D>N Missense

PUS3 pseudouridylate synthase 3 tRNA processing CCDS8466.1 chr11:125268896G>C 1440C>G 480I>M Missense

Bona Fide Candidate: PIK3CA

Validation in PDX Model

1800

2000

2200

CONTROL (n=8)

BEZ235 (n=9-8)

1400

1600

1800

SEM

( )

BKM120 (n=8)

GEMCITABINE (n=8)

1000

1200

e (m

m3 )

±

600

800

or V

olum

e

0

200

400

Tum

o

00 2 4 6 8 10 12 14 16 18 20 22 24 26 28 30

Time (days)

Genetic Evolution of PDA

Yachida, Nat 2010

Molecular Subtypes of PDA

Collison et al, Nat Med 2011.

Drug Response in Molecular Subtypes of PDASubtypes of PDA

Collison et al, Nat Med 2011.

PRODIGE 4/ACCORD 11 Trial

GemcitabineOS (months)

Advanced PDA(n= 342)

OS (months)

Gem FOLFIRINOX

6.8 11.1

FOLFIRINOX

• Primary endpoint: Survival

Conroy et al, NEJM 2011

First Line Treatment

FOLFIRINOX(-/+)

FOLFIRINOX

FOLFIRI(+/+)

5FU ERCC1/Topo1

FOLFOX( )

(-/-)

TS

(-)

(+/-)GEM + Capecitabina

(+)(No Ampl/No WT)

SPARC

(+)GEM + nab

GEM EGFR/K.Ras

GEM + Erlotinib

SPARCGEM

( -)

GEM + Erlotinib(Ampl ó WT)

PSCA Targetg• PSCA is GPI-linked cell surface protein of unclear function.

• Shares 30% homology with stem cell antigen type-2, but expressed mostly in differentiating epithelial cells of GI and p m y ff g p fGU tracts.

• Literature suggests PSCA expression in 87 100% of prostate • Literature suggests PSCA expression in 87-100% of prostate tumors1,2,3 and ~60% of pancreatic tumors.

• Germline SNPs in PSCA associated with risk of gastric and bladder cancer. Strongest association with missense variant in 1st exon, shown to alter PSCA expression levelsin 1st exon, shown to alter PSCA expression levels

AGS-1C4D4 in PDA

Gemcitabine6m Suv Rate

Advanced PDA(n= 196)

2:1 Randomization

6m Suv Rate

Gem Gem + AGS

44 % 61 %2:1 RandomizationGem + AGS1CD4 p = 0.016

• Primary endpoint: 6-m Survival Rate

Wolpin et al, ECCO 2011

Multifaceted Biology of PDAgy

M Hidalgo, NEJM 2010

SPARC (Secreted Protein and Rich in Cystein)

• Sparc is a member of a family of matricellular proteins whose function is to modulate cell-matrix interactions. (Bornstein and Sage, Curr. Opin. Cell Biol.,2002, Brekken and Sage, Matrx Biol., 2001)

• Sparc was shown to be expressed in tumors and the surrounding stroma of Sparc was shown to be expressed in tumors and the surrounding stroma of different types of cancer: breast, melanoma, glioblastoma, ovarian, colorectal and pancreatic.

M i i h i ll i S KO i h • Murine pancreatic tumors grown orthotopically in Sparc KO mice show increased metastatic potential. (Arnold S., et al.,Disease models and mechanism, 2010)

• The expression of SPARC by peritumoral fibroblasts accounts for a poorer prognosis for patients with pancreatic cancer. (Infante J. et al., Journal of Clinical Oncology, 2007)

• Sparc is a target for nanoparticle – albumin bound (Nab) drugs due to its high affinity for binding serum-albumin. (Desai N., et al., Clinical Cancer Research., 2006; Sage et al.,1984)

SPARC Expression in Tumor Stroma pis Associated with Worse Outcome

Infante et al, 2007

Nab Paclitaxel – (Abraxane®) 1 C m ph f ll id l p n i n f n n p ti l 1. Cremophor – free colloidal suspension of nanoparticle

paclitaxel (Abraxane®) stabilized with human serum albumin (130 nm particles )

2 R id di t ib ti t t 2. Rapid distribution to tumor (Sparreboom et al. Clin Cancer Res. 11: 4136, 2005)

3. Intra-tumoral accumulation of nab-paclitaxel - (Desai l Cli C R 12 1317 2006)et al. Clin. Cancer Res. 12: 1317, 2006)

Fluorescent nab-paclitaxel nanoparticles in syringe injected via tail

vein

Imaged Tumor

Phase I/II Trial of Gem + Nab-paclitaxel

• Multicenter trial run through PCRT.F d b l • Fix dose-gemcitabine + escalating dose of nab-paclitaxel.p

• 67 patients with stage IV disease.• Expansion cohort of 44 patients at Expansion cohort of 44 patients at the MTD.R d b PET d • Response assessed by PET and CA199.

• SPARC assessment in tumor biopsies.

Von Hoff et al, JCO 2011

Rapid Response b PET d CT Sby PET and CT Scans

Scan Baseline Post Rx 1 Post Rx 2

Post Rx 3

Post Rx 4

Post Rx 5

Post Rx 6

Post Rx 7

RECIST Total

19.4 15.2 12.6 10.7 10.4 9.3 9.6 7.6

Baseline: 6.03.2007

ET S

cans

Total

% Change

- -21.6 -35.1 -44.8 -46.4 -52.1 -50.5 -60.8

nse

by P

E

Scan

s Baseline: 9.10.2008

d R

espo

n

se b

y C

T S

8.15.2007

Rap

id

Res

pons Cycle 7:

04.13.2009

Von Hoff et al, JCO 2011

Preclinical Platform

Rubio et al, CCR 2006

Average Response Rate in Xenografts (n = 11)

50

60al

siz

e (%

)

40

of it

s in

itia

20

30

ssed

50%

o

10

mor

s re

gres

0GEM ABI GEM+ABI

Tum

Effects of nab-Paclitaxel in Tumor Stroma

Effects of nab-Paclitaxel on GemcitabineDeliveryDelivery

6000

7000

8000

an ±

SEM

)

GEM alone

GEM+ABI22000

24000

an ±

SEM

)

GEM aloneGEM+ABI

3000

4000

5000

6000

g/g

tum

or (M

ea

20000

22000

g/g

tum

or (M

ea0

1000

2000

3000

once

ntra

tion

n

16000

18000

once

ntra

tion

n0

GEM

Co 16000

dFdU

Co

Pancreatic Cancer is Hypovascular

Olive…Tuveson, Science 2009

Anti-Stromal Agent Combination Activity(Panc163)(Panc163)

Roles of Hh Pathway in PDAy

Hidalgo and Maitra, NEJM 2009

Targeting CSC Pathways

Jimeno et al, MCT 2009

High DR-5 Expression in Cancer Stem Cells

Bulk Population

CD24+CD44+ Population

89.4%

30.1%

Cel

l #

l #

DR5

ALDH+ PopulationDR5

Cel

6.22%

94.3%

CD

44

SSC

Cel

l #

4.62%

CD24 ALDEFLUORDR5

Rajeshkumar et al, MCT 2010

Targeting Extrinsic PathwayTargeting Extrinsic Pathway

TGI of Established Human Pancreatic Tumor Xenografts Treated with CS1008, GEM and GEM+CS1008 on day 28

Rajeshkumar et al, MCT 2010

Conclusions• Genetically complex, unstable and heterogenous

diseasedisease.

I i i t t iti f li di i • Incipient opportunities for personalize medicine are emerging.

• Targeting pancreatic cancer stroma is appealing preclinically and clinicallypreclinically and clinically.

• Pancreas CSC directed therapies may be needed • Pancreas CSC directed therapies may be needed for full prevention of cancer failure.

GI Group at CNIOM. Mustenau A. CalleP. Lopez-Casas M. MuñozP M lli R S h

SU2C Team at JHUC. Iacobuzio-DonahueR HrubanP. Morelli R. Sanchez

R. Martinez E. GarciaR. HrubanA. MaitraR. KumarE. OliveiraGI Program at Johns HopkinsV. VelculescuC. Dang

GI rogram at Johns Hopk nsD. LaheruA. De Jesus

TGenTGenD. Von HoffR. Ramanathan

CBID. SidranskyE. Bruckheimer

PCRT

GI Group at CIOCCE. VicenteY. QuijanoA Cubillo

J. Martell

A. StollC. MoriartyPCRT Investigators

A. CubilloJ. RodriguezC. RubioO. Hernando

Pancreas Team at CNIOM. Barbacid/C GuerraP Real

LDT Lab at CIOCCF. Lopez-Rios

P. RealC. HeeschenN. Malats

Imaging Unit at CNIOP. Mulero