Transcript Pten +/ + /p53 L/+

Dual PTEN/P53 suppression
promotes high grade sarcomas
by activating Notch
Eva Hernando, Ph.D.
Challenges in the study of
high grade sarcoma and leiomyosarcoma
• Genomic complexity
• Low number of known recurrent genetic
mutations
• Limited enrolment of patients in clinical trials
 better understanding of the molecular basis
 mouse models that recapitulate human disease
 Preclinical testing (immunotherapy)
Targeted sequencing revealed low incidence
of mutations in classical ‘cancer genes’
• N=40 specimens (NYU School of Medicine and
Northwestern University)
• Platform: OncoMap3 core and extended panels
(1047 mutations in 116 genes)
• 73 candidate mutations on 44 genes
• 6 confirmed mutations on 2 genes:
– TP53 (one V157F and two R273C amino acid
substitutions).
– a low-frequency germ-line single-nucleotide
polymorphism in EGFR (S703F)
Guijarro et al., AJP 2013
Partial PTEN and TP53 deletions are common in
HGUPS and LMS
Barretina et al., Nat Gen 2010
Guijarro et al., AJP 2013
PTEN and TP53 expression is commonly
downregulated in HGUPS and LMS
Guijarro et al., AJP 2013
Smooth-muscle specific conditional inactivation
of Pten and p53
F0
Tagln-cre+/+
F1
F2
Tagln-cre+/- /PtenL/+p53L/+
PtenL/L p53L/L
Tagln-cre+/- /PtenL/+ p53L/+
Tagln-cre/ Ptenwtp53wt
Tagln-cre/ PtenL/+p53L/+
Tagln-cre/ PtenL/+ p53wt
Tagln-cre/ Ptenwtp53L/+
Tagln-cre/ PtenL/Lp53wt
Tagln-cre/ Ptenwtp53L/L
PtenD/+p53D/+ mice have shorter overall survival
than Pten+/+p53D/+
Guijarro et al., AJP 2013
Hernando et al., Nat Med 2007
PtenD/+p53D/+ mice have increased sarcoma
incidence compared to Pten+/+p53D/+
Guijarro et al., AJP 2013
PtenD/+p53D/+ HGS histologically resemble the
equivalent human lesions
Guijarro et al., AJP 2013
Tagln-cre marks smooth muscle cells
Tagln-cre+/+ x ROSA26-loxP-STOP-loxP-LacZ
Guijarro et al., 2013
Tagln-cre is also expressed in
mesenchymal stem cells
Murine HGUPS have complex karyotypes
PtenD/+p53D/+ HGUPS are highly metastatic
Liver
Pancreas
Lung
Pten is haploinsufficient
for sarcoma tumor progression
PtenD/+p53D/+ Pten+/+p53D/+
No mutation
Which advantage does Pten
downregulation
confer to p53 deficient sarcomas?
PtenD/+/p53D/+ bone-marrow mesenchymal stem
cells display higher clonability and proliferation
PtenL/+/p53L/+ and Pten+/+/p53L/+ bone marrow-derived mouse mesenchymal stem cells
+ Adeno-cre infection
Colony formation assay
Proliferation assay
Both, acute and genetically induced concomitant inactivation of Pten and P53 leads to
increased clonogenic and proliferative capacity of mesenchymal stem cells
Pten suppression in Pten+/+/p53D/+ tumor cells
increases clonogenic capacity and invasion
The Notch signaling pathway is up-regulated in
PtenΔ/+Tp53Δ/+ murine HGUPS
The Notch signaling pathway is up-regulated in
PtenΔ/+Tp53Δ/+ murine HGUPS
Gamma-secretase inhibition suppresses the
clonogenic and invasive potential of tumor cells
GSI
Colony formation assay
shPten pro-oncogenic effects are counteracted
by Notch inhibition
Colony formation assay
invasion assay
Model of molecular classification of high grade sarcoma
patients for targeted therapies
Conclusions
• Conditional inactivation of Pten and p53 in mouse
mesenchymal progenitors recapitulates the histological and
cytogenetic features of human HGUPS and LMS
• Pten deficiency confers increased clonogenic and invasive
potential to p53 heterozygous sarcoma cells
• Pten suppression in p53 deficient MSCs and tumor cells
triggers Notch signaling
• Gamma-secretase inhibitors negate the pro-oncogenic
effects of Pten suppression in p53 deficient sarcoma cells
 PTEN/TP53 deficient HGUPS may be susceptible
to Notch inhibition
Hernando Lab
Maria V. Guijarro
Laura Danielson
Miguel Segura
Martha Vega
Avital Gaziel
Silvia Menendez
Luca Paoluzzi
Doug Hanniford
Raffaella Di Micco
Lisa Koetz
Barbara Fontanals
Elena Sokolova
Vivien Low
Olivia Blackburn
Rana Moubarak
Veronica Davalos
Praveen Agrawal
Collaborators
Sonika Dahiya
Jian Jun Wei
Carlos Cordon-Cardo
Pier Paolo Pandolfi
Funding
American Cancer Society
Edna´s Foundation of Hope
Liddy Shriver Sarcoma Intiative
Sarcoma Foundation of America
Melanoma Research Alliance
DOD
NIH/NCI
NIH/NIAMS
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