Featured Poster Abstract 84:

CD8 T cell-mediated immune responses are critical to the increased efficacy of Doxil in

BRCA1 deficient tumors

Gina Mantia-Smaldone, Victoria Gamerman, Phyllis Gimotty, Regina Loomis, Lukas Ronner, Christopher Morse, Sandra Orsulic,

Stephen Rubin, George Coukos, Sarah Adams

BRCA-deficient tumors are vulnerable to the immunomodulatory effects of Doxil

• 56% of women with recurrent BRCA-associated ovarian cancer demonstrated a response to Doxil1

• In addition to causing DNA damage, Doxil increases the vulnerability of ovarian cancer cells to T cell attack

• Immunophenotypic changes in response to Doxil are more pronounced in BRCA-/- cells

• BRCA-/- tumors recruit higher numbers of T cells after Doxil exposure

• We hypothesized that the immunogenicity of BRCA-/- tumors contributes to the improved clinical response to Doxil among women with hereditary ovarian cancer

• The role of T cells in tumor clearance following exposure to Doxil was tested using a murine BRCA1-/- ovarian cancer model 1Adams SF et al, Gynecol Oncol 2011

Higher numbers of CD3+ Tumor Infiltrating Lymphocytes are present in murine BRCA1 -

tumors following Doxil therapy

0

20

40

60

80

100

low CD3 high CD3

Perc

ent

Control

Doxil

Endpoint: Weight ≥30 grams

Antibody

-7 -6 -5 0 3 10 17Experimental Day

Doxil

Tumor Innoculation

Antibody q3-4d

Doxil Doxil Endpoint: Weight ≥30 grams

Antibody

-7 -6 -5 0 3 10 17Experimental Day

Doxil

Tumor Innoculation

Antibody q3-4d

Doxil DoxilEvaluated in a cohort of 23 women with BRCA mutations and 41 women with sporadic cancer treated at three institutions between 2000-2010 for recurrent cancer. The response to Doxil was associated with a significant improval in overall survival

Doxil increased intratumoral T cell recruitment and overall survival in mice with BRCA1- tumors

Doxil therapy significantly prolonged the survival of

animals with BRCA1- tumors

Doxil exposure increased cytotoxic CD8 T cells and

reduced suppressive FoxP3 T cells

Both CD4 and CD8 T cell populations were present in BRCA1- tumors and ascites

Survival measured from the time of tumor inoculation until animals reached 30g due to ascites accumulation

Survival (Days)

P=0.0407

Control

Doxil

Flow cytometry was performed on tumor, ascites, spleen and peripheral blood mononuclear cells (PBMCs). Illustrated are the portion of CD45+/CD3+ T cells present in representative tumor or ascites samples for placebo or Doxil treated mice.

The survival advantage associated with Doxil treatment persisted in the absence of CD4 T cells

An immunodepleting antibody successfully eliminated CD4 T cells from BRCA1- tumors and

ascites

In the absence of CD4 T cells, the proportion of CD8 T cells was

higher in Doxil-treated animals

After treatment with anti-CD4 antibody, a population of CD4-/CD8- cells persisted

Doxil significantly prolonged the survival of CD4-depleted animals

with BRCA1- tumors

Survival measured from the time of tumor inoculation until animals reached 30g due to ascites accumulation

Survival (Days)

** p< 0.05

P=0.0079

Doxil

Control

There was no statistically significant difference in survival after Doxil treatment in the absence of CD8 T cells

An immunodepleting antibody successfully eliminated all CD8 T cells from BRCA1- tumors and

ascites

In mice lacking CD8 T cells, the proportion of mature CD4 T cells,

but not suppressive FoxP3 T cells, was higher after Doxil

treatment

The survival among animals depleted of CD8 cells was not

improved with Doxil treatment

After treatment with anti-CD8 antibody, a population of CD4-/CD8- cells persisted

Survival measured from the time of tumor inoculation until animals reached 30g due to ascites accumulation

* p< 0.05P=0.0952

Doxil

Control

Survival (Days)

*

• Doxil improved survival in a BRCA1- model of ovarian cancer• This effect persisted in the absence of CD4 T cells but is diminished in the absence

of CD8 T cells• Cytotoxic CD8 T cells are necessary for the enhanced response to Doxil observed in

BRCA1- tumor bearing mice• BRCA1- deficient ovarian cancers may be more susceptible to immunotherapeutic

strategies

Limitations: • These results represent a small pilot study and

need to be confirmed in larger cohorts• Unfortunately the national Doxil shortage has

limited our ability to conduct planned experiments

This work was supported by a grant from the

Conclusions