Abstract 4743: Ex vivo expanded cytokine-induced killer cells maintain cytotoxic activity against ovarian cancer in an in vivo model.
Authors: Ingersoll Susan Blaydes , Srivastava Milan , Ahmad Sarfraz , Stavitzski Nicole M. , Oyer Jeremiah L. , Ali Ghazanfar , Banks Robert K. , Finkler Neil J. , Edwards John R. , Holloway Robert W. 
Published: August 2014

Introduction: Ovarian cancer (OC) is the leading cause of death among gynecological malignancies and thus there is a compelling need to develop novel treatment protocols. The study aim was to asses the anti-tumor efficacy of ex vivo expanded cytokine-induced killer cells (CIK) against OC using in vitro and in vivo models.

Methods: CIK were expanded from peripheral blood mononuclear cells (PBMC) isolated from OC patients (n=21) and healthy donors (HD; n=11), in a cocktail of IL-2, IL-12, and anti-CD3 antibody ± IL-7 (10 ng/ml). Cell populations present were determined by flow cytometry. CIK cytotoxicity was assessed against an OC cell line (SKOV3-AF2) in presence of IL-2 and IFNα-2b. To determine in vivo activity, OC cells (1x106) were injected intraperitoneally (IP) into female athymic nude mice (n=100). Day-7 post-OC cell injection, mice were injected IP with treatment cells [CIK (5x106) from OC patient or PBMC (5x106) from HD]. Also on day-7, IL-2 treatment was initiated (4,000 U/IP-injection) and continued thrice weekly. Control mice with and without treatment cells or IL-2 were also included. Mice were sacrificed when they became moribund due to tumor burden at which time solid tumor and ascitic fluid were measured and collected. Cell and cytokine treatment efficacy was measured by overall survival of the mice.

Results: CIK expanded exponentially from both HD (149-fold) and OC patients (139-fold) over 3-weeks; expansion cultures consisted primarily of T-cells (&gt;98%) by days 7-8. However, CIK expansion was doubled in the presence of IL-7 (159 vs. 81-fold; p&lt;0.05). CIK from OC patients and HD elicited a significant cytotoxic response against OC cells (26.8% and 24.8%, respectively) compared to negative controls lacking CIK (0.4%; p&lt;0.01). In the in vivo experiments, mice tolerated all CIK, PBMC and cytokine dosages tested as evidenced by no significant weight loss (&lt;10%) or treatment-associated morbidity (100% survival, post-treatment cell injection). Mice that received CIK + IL-2 (expanded in the absence of IL-7) or PBMC + IL-2 demonstrated a survival advantage at 9-weeks (50% and 55%, respectively) compared to untreated mice (9%), treated with IL-2 alone (13%) or PBMC alone (14%). There was no significant difference in average tumor weight or ascitic fluid between treatment groups, probably due to mice being sacrificed when they were moribund due to tumor burden (not a specific time post-tumor cell injection).

Conclusions: Ex vivo expanded CIK demonstrated an anti-OC effect both in vitro and in a xenograft model. CIK expanded from HD and OC patients elicited a significant cytotoxicity against OC cells in vitro. In addition, OC bearing mice treated with CIK + IL-2 or PBMC + IL-2 showed improved survival. Additional mouse studies are underway to test the efficacy of CIK, PBMC, and their combination. Data generated will provide the basis for development of an immunotherapy-based Phase I trial for OC.

Citation Format: Susan Blaydes Ingersoll, Milan Srivastava, Sarfraz Ahmad, Nicole M. Stavitzski, Jeremiah L. Oyer, Ghazanfar Ali, Robert K. Banks, Neil J. Finkler, John R. Edwards, Robert W. Holloway. Ex vivo expanded cytokine-induced killer cells maintain cytotoxic activity against ovarian cancer in an in vivo model. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 4743. doi:10.1158/1538-7445.AM2013-4743