BRCA1 Associated epigenetic

regulation of p73 mediates

effector pathway for
chemosensitivity in ovarian
carcinoma
 INTRODUCTION
 Ovarian carcinoma is the fifth commonest carcinoma in the world.
 Commonest gynecological cancer in united kingdom.
 It arises due to germ line mutation of the BRCA1 tumor suppressor
gene.

 Mutation in BRCA1 result in loss of p53 activity.

 despite the loss of p53 BRAC1 cell exhibit increased sensitivity in
platinum based chemotherapy.

 As well as improved overall survival.

 Effector pathway behind this concept is poorly understood.

According to the multiple studies on BRCA1 deficient human
and murine carcinoma , Ibrahim etall shows that selective up
regulation of trans activating isoform of p53 related gene P73
(TAP73) is involve in this phenomenon.

TAP73 serves as an important contributor to the platinum

induced DNA damage response and to chemosensitivity in
ovarian carcinoma.


TAP73 does not play any role in BRCA1 expressing cells but
knock down of TAP73 can abolish the effect of cisplatin.
 MATERIAL AND METHODS
Tissue culture and reagents for study of murine model
Murine ovarian carcinoma cell lines (T1,T2,T3,TBR2,TBR5 and
TBR6).
Western blot analysis to confirm the absence of whole length
BRCA1 protein.
These lines were maintained in 10% fetal bovine serum ,100
IU/ML Penicillin and 100 microgram /ml of streptomycin.
For study of human cells
UWB 1.289 human cell lines.
1:1 RPMI 1640/MEGM supplemented with 3% fetal bovine serum.
RNA analysis ,lentiviral/retroviral production
And chromatin immunoprecipitation
 QRT-PCR analysis
 Cotransfection is used to generate lentiviral and retroviral stock.
Bisulphite treatment and methylation analysis
 200 to 500ng of genomic DNA were used.
 After confirmation of desired product methylation was quantitated in
sequence product by measuring the heights of the C and T peaks.
Patient sample acquisition and processing
 Tumor specimens were obtained from from the MGH gynecological
tissue repository.
 RNA/DNA was isolated from dry frozen tumor samples.
 RESULTS
Cisplatin sensitivity of BRCA1 deficient ovarian
carcinoma cells
 BRCA1 associated tumor lines consistently exhibited significantly
increased sensitivity to cisplatin comparison to wild type in murine
model.
 In human assay showed the same result confirming the increase
platinum sensitivity of UWB 1.289 cells relative to their isogenic
BRCA1 counter part.
 Ibrahim et all found that cisplatin induces apoptotic death in both
murine and human BRCA1 deficient cells ,as evidence by cleavage of
poly (ADP ribose) polymerase on western blot analysis.
Expression of P73 and its target genes correlate with
cisplatin sensitivity in BRCA1 deficient cells
 After cisplatin induction Ibrahim etall observed robust induction of
the P53 regulated proapoptotic genes Noxa ,Puma and P53AIP1 in
both human and murine BRCA1 deficient cells but not in wild type.
 These finding suggest a distinct cell death pathway in the absence of
BRCA1 which involve P53 family member P73.
TAP73 is necessary for cisplatin sensitivity
 TAP73 knock down essentially abolished cisplatin induced Noxa
expression in the three murine BRCA1 deficient ovarian carcinoma
cells.
 Knock down of TAP73 induced significant resistant in human BRCA1
deficient UWB 1.289 cells although it did not affect proliferation and
cell viability in the absence of cisplatin.
The ZEB1 transcriptional repressor of TAP73 in BRCA1
expressing cells
 Ibrahim et all observed that binding of ZEB1 with TAP73 potently
repressed TAP73 mRNA expression.
 This is the main reason that why the phenomenon of TAP73 exist only
in BRCA1 deficient ovarian carcinoma.

Methylation of ZEB1 locus controls the binding of ZEB1

with TAP73
 To confirm this model Ibrahim et all treated ZEB1 with 5 azacytidine
(demethylating agent) which result in 100 fold increase in ZEB1
binding in BRCA1 deficient cells.
 It shows that methylation of ZEB1 locus will decrease the binding of
ZEB1 and TAP73 and increase the expression of TAP73 which
ultimately increase the chemosensitivity for cisplatin.
 DISCUSSION
 TAP73 is an important contributor to chemosensitivity in
BRCA1 deficient carcinoma tumor.

 Ablation of TAP73 promotes chemoresistance.

 The description of pathway responsible for cell death was
good but not proper.

 actual concept behind this phenomenon is not present in this

research paper.

 Research is still going on to find the actual concept