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Stat3 Transgenic Mice as a Model for Human Basal Subtype of Invasive Bladder Cancer Philip L. Ho, MD Urologic Oncology Fellow The University of Texas at MD Anderson Cancer Center Houston, TX Disclosures None Outline Cancer Stem Cells

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Stat3 Transgenic Mice as a Model for Human Basal Subtype of Invasive Bladder Cancer

Philip L. Ho, MD Urologic Oncology Fellow The University of Texas at MD Anderson Cancer Center Houston, TX

SLIDE 2

Disclosures

None

SLIDE 3

Outline

Cancer Stem Cells
Keratin 14, Basal subtype
Stat3

– Cancer – Transgenic mouse tumors – Mechanism and Targets

SLIDE 4

Introduction

Minimal improvement in bladder urothelial

carcinoma patient outcomes have been made

Standard of care remains surgery and

chemotherapy

– No targeted therapies

Need to further understand biology in order to

develop targeted therapies

SLIDE 5

Cancer Stem Cell Model

CSC CSC CSC

Intratumoral Heterogeneity

1. Tumor-initating

potential

2. Self-renewal

Cancer Stem Cell Progenitor Cells Differentiated Cell

3. Ability to

differentiate

Recapitulate

heterogeneity of

riginal tumor

SLIDE 6

Bladder Cancer Stem Cell Markers

Ho, PL. Kurtova, A., Chan, KS. Nature Reviews Urology 2012

SLIDE 7

Keratins (K)

Structural proteins 

intermediate filaments

Expression

– Cell-type – Differentiation – Function

Cancer

– Diagnosis – Prognosis

Karantza, V. Oncogene 2011

SLIDE 8

Bladder Cancer – Basal Subtype and K14

TCGA McConkey MDACC Kim UNC

The Cancer Genome Atlas Research Network. Nature 2014 Choi et al. Cancer Cell 2014 Damrauer et al. PNAS 2014 Basal

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K14 Gene Expression Correlates with Survival

Overall Survival – Multivariable analysis

HR (low-high) p K14 1.51 (1.11 – 2.03) 0.0077 Stage 1.32 (1.20 – 1.45) <0.001 Grade 1.32 (1.00 – 1.74) 0.047 Gender 1.02 (0.82 – 1.27) 0.86 Age 1.45 (1.23 – 1.71) <0.001

Volkmer et al. PNAS 2012

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HR (low-high) p K14 2.11 (1.18 – 3.80) 0.0038 Stage 23.32 (2.72 – 199.83) 0.0038 Grade 1.29 (0.27 – 6.10) 0.87 Gender 0.87 (0.63 – 1.19) 0.41 Age 1.03 (1.01 – 1.06) 0.042 HR (low-high) p K14 2.42 (1.19 – 4.93) 0.032 Stage 0.89 (0.66 – 1.20) 0.16 Grade 0.89 (0.61 – 1.32) 0.55 Gender 1.20 (0.47 – 3.05) 0.71 Age 1.03 (1.00 – 1.06) 0.069

Multivariable analysis Multivariable analysis

K14 Protein Expression Correlates with Survival

Volkmer et al. PNAS 2012

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K14 Reporter Construct

DAPI Tm K14

Transduced with hK14.tdTomato (Tm) virus

FACS Tm+/K14+ cells Tm-/K14- cells

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K14 Reporter Construct

sphere number

A B C Tm- Tm+

Fold Change sphere number

Cells Tm+ 5000 7/7 500 7/7 50 6/7 10 2/7 Cells Tm- 5000 7/7 500 5/7 50 1/7 10 0/7

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Bladder Cancer – Basal Subtype and K14

TCGA subgroup McConkey group

…Stat3

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Stat3 – basal subtype

Choi, W, Porten, S, Kim, S, Willis, D,…McConkey, DJ. Cancer Cell 2014

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STAT Proteins

Signal Transducer and activator of transcription (STAT)

Proteins

Family of proteins that mediate
Growth factor signaling
Apoptosis
Angiogenesis
Immune responses
Tumor development
Bromberg. Stat proteins and oncogenesis. Journal of Clinical Investigation. May 2002

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STAT3 activation

Target genes include

– Antiapoptotic proteins – Proliferation-associated proteins – Proangiogenic factors

Regulator of EMT transcriptions

factors

– Twist, Snail

Bromberg. Journal of Clinical Investigation. May 2002

Chan et al. Oncogene 2008. Pedranzini et al. Journal of Clinical Investigation. 2004. Wendt et al. JAK STAT. 2014

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STAT3 and Cancer

Constitutively active in many human

malignancies

Naturally occurring mutations in Stat3 have

yet to be identified

Dysregulation may play role in constitutive

activation of Stat3

Bromberg. Journal of Clinical Investigation. May 2002

Myers Jr. et al. Science 323 (2009): 723-24.

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STAT3 and Bladder Cancer

Ho, PL …Chan, KS. Cancer Research 2012

Nuclear, active Stat3 was present in 100% of high-grade, invasive human UC tumors but

nly in 37.5% of low‐stage tumors. Stat3 was also found to co‐localize with CK14

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2 Pathways of Bladder Cancer

Model?

Dinney, CP, McConkey, DJ et al. Cancer Cell 2004 Ho, PL., Kurtova, A., Chan, KS. Nature Reviews Urology 2012

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K5.Stat3C Transgenic Mice

 K5.Stat3C transgenic mice with constitutively active Stat3 driven by the basal keratin 5 promoter were exposed to nitrosamine (BBN) through their drinking water.

Bovine keratin 5 promoter

Stat3C

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K5.Stat3C Transgenic Mice

Stat3-transgenic Wild-type

Ho, PL & Chan, KS. Cancer Research 2012

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K5.Stat3C Transgenic Mice

Ho, PL & Chan, KS. Cancer Research 2012

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K5.Stat3C Tumors – Sphere-forming Assay

Ho, PL & Chan, KS. Cancer Research 2012

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Stat3  ?

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Stat3  ?

Immunofluorescence staining of SOX2 in Stat3-driven lesions

SOX2

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Activation of Stat3

Yu et al. Nature Reviews Immunology. January 2007

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siRNA inhibition of Src and Jak

JAK2 inhibitor - Ruxolitinib Src inhibitor - Dasatinib

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Conclusion

K14 – marker for cancer stem cells
Stat3 – drives basal subtype of invasive

bladder cancer

Stat3 transgenic mouse

– Invasive bladder cancer model – Preclinical studies to investigate inhibitory drugs

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Acknowledgements

Baylor College of Medicine

Keith S. Chan PhD
Seth P. Lerner MD

Chan Lab

Antonnia Kurtova
Jing Xiao PhD
Erica J. Lay
Yanting Qi PhD
Ross E. Krasnow MD

MD Anderson Cancer Center

Ashish M. Kamat MD
David J. McConkey PhD
Colin P. N. Dinney MD