1. Characterizing the DDR pathway
in acutely treated testicular germ
cell tumors
Alicia Braxton
2. Testicular Germ Cell Tumors
• Extremely sensitive to chemotherapy
• Curable with chemo + surgery
– Over 99% of patients
• Curable even after metastasis occurs
– 85% success rate of treatment once metastasis
develop (5 years post treatment)
Huddart, R. A. et al . (2003). Management of Testicular Germ Cell Tumors. American
Journal Of Cancer, 2(5), 325-334.
3. Somatic tumors are chemoresistant
• Chemotherapy resistance poses a challenge for
treatment
• Accumulate mutations in DDR pathways
– Ex: p53
• Infer: Compromised repair pathways
chemotherapy resistance
Germ Cells avoid mutations in their DDR
pathways.
Perhaps this makes the TGCT sensitive to
chemotherapy treatment.
Bartkova, J. et al. (2007). DNA Damage Response in human testes and testicular germ cell tumors:
biology and implications for therapy. International Journal of Andrology, 30, 282-291
4. Aim: What mechanisms make TGCT sensitive
to chemotherapy?
If we are able to identify the mechanism
conferring chemotherapy sensitivity to TGCT,
perhaps we could apply that knowledge to
the treatment of more resistant somatic cell
tumors.
5. Types of TGCTs
Gonadal Germ Cell Cancers
Ovarian
Testicular
Seminoma Non-seminoma
Yolk Sac
Tumor
Choriocarcinoma
Teratoma
Embryonal
Carcinoma
Teratocarcinoma
Rare
Most Common
Cancer of Young Men
Tim Pierpont
6. Skeletal Muscle
(Mesoderm)
Embryonal Carcinoma
(Pluripotent Cell Type)
Ciliated Respiratory
(Endoderm)
Neural Like
(Ectoderm)
Embryonal Carcinoma
(Pluripotent Cell Type)
Teratoma and Embyronal Carcinoma Components
Teratocarcinomas
Tim Pierpont
7. Cancer Stem Cell
• 2 properties
– Self renew
– Differentiate
• Current cancer treatment strategy
• Target cancer stem cells: Inhibit regrowth of
tumor
Tim Pierpont
8. A Novel Mouse Model for TGCTs
• Novel TGCT on mice 129 background
• Undergone 2 oncogenic events
• Original thinking: Post-natal development of a
more malignant tumor in the testes
LSL-KrasG12D STOP 1
G12D
Ptenflox Exon 5
Inactive
Active PtenΔ5 INACTIVE
Oncogenic!
Tim Pierpont
Lox-KrasG12D 1
G12D
ACTIVE
Oncogenic!
Stra8-Cre
9. Human treatment of TGCT
• Chemotherapy + Inguinal orchidectomy
• Chemo: BEP protocol
– Bleomyocin
– Etoposide
– Cisplatin
• 3-4 cycles of treatment depending on stage of
cancer
• Cisplatin and etoposide introduced in late
70s/early 80s
Huddart, R. A. et al . (2003). Management of Testicular Germ Cell Tumors. American
Journal Of Cancer, 2(5), 325-334.
10. Cisplatin
• Platinum containing chemotherapuetic agent
• Covalent cross-linking of DNA double
stranded breaks apoptosis
• Used in people to treat TGCT, as well as
ovarian, bladder and cervical cancers
• Malignant embryonic carcinoma cells-
sensitive to Cisplatin
http://chemocare.com/chemotherapy/drug-info/cisplatin.aspx#.U80vNfldUsA
Cisplatin : Pharmacology, clinical uses and adverse effects. 2012.
Hauppauge, NY, USA: Nova Science Publishers, Inc.
14. Increased survival with treated allograft tumors
10k
Treated
1mil
Treated
100k
Treated
25k
Cumulativesurvival
Days
0%
20%
40%
60%
80%
100%
0 605040302010
Tim Pierpont
15. We know the mice are sensitive to Cisplatin
treatment: Now we are working to determine
the mechanism behind their sensitivity
16. What makes TGCT chemosensitive?
• Mice with tumors injected IP with Cisplatin
• Tumors collected at various time points post
treatment
– 6, 12, 24, 36, 48, and 96 hours
• Cassette, embed, section and fix samples to
slide
• Stain using IHC techniques
17. Initial Markers of Interest
• Oct4- TF that promotes pluripotency
– Expressed by malignant embryonic carcinoma
cells
• TUNEL- tags apoptotic cells
• yH2AX- identifies double stranded breaks in
DNA
– shows the DNA damage
• Ki67- marks proliferating cells
Help us gauge at which time points to look
closer with more specific markers: p53, pCHK1,
and other markers of DDR pathway
25. Future Research
• Repeat TUNEL on all samples
• Look into other markers of the DDR pathway
– p53, pCHK1
– Differentiation-Sox17
• Collect another 6 hour post treatment sample
– Oct4 negative existing samples
• More acute time point
– 2 hours
26. Thanks To
Tim Pierpont
Amy Lyndaker
Claire Anderson
Pei Xin Lim
Erin Daugherity
Kelly Hume
Joanna Mleczko
Elizabeth Moore
Yashira Negrón
David Karambizi
Cindy Luan
Ellen Hong
Zanah Francis
Charlton Tsai
Thank you!
Cornell Pathologists
Donald H. Schlafer,
Teresa L. Southard
Committee
Robert Weiss
Doina Tumbar
John Schimenti
Duke University
Matthew S. Cook
Blanche Capel
Collaborators
