Figure 2: FluoroSpot analysis of anti-tumor immunity in Hepa1-6-luc tumor bearing mice. C57BL/6 mice with established Hepa1-6-luc tumors were dosed with anti-PD-1 or isotype control antibodies. White blood cells from 200µL of peripheral blood were cultured with irradiated Hepa1-6-luc cells. A) Quantitation of the frequency of IFNγ and IL-4 secreting cells using a CTL ImmunoSpot® S6 Universal analyzer. B) Analysis of the IFNγ-producing cell frequency vs tumor volume in each mouse. C). Th1/Th2 balance in each mouse assessed by measuring the ratio of IFNγ/IL-4 producing cells.
Similar to the mechanism of action for a vaccine, a growing tumor elicits an adaptive immune response that consists of antibody and cytokine producing tumor-specific cells1. The quality of the response can be characterized by examination of IFNγ and IL-4. IFNγ has been well documented to inhibit tumor growth by enhancing T cell-mediated by cytotoxicity. In contrast, IL-4 promotes tumor growth through a variety of immunosuppressive effects. This dichotomy is often referred to as a Th1/Th2 balance2. As shown in Figure 2, anti-PD-1 increased the frequency of tumor-responsive cells producing IFNγ following ex vivo stimulation. In contrast, the frequency of IL-4 producing cells was not significantly increased, suggesting checkpoint inhibition triggered an enhancement of anti-tumor protective immunity. In support of this, mice with the highest frequencies of IFNγ producing cells harbored tumors with the smallest volumes. Furthermore, anti-PD-1 increased the ratio of IFNγ/IL-4 producing cells, which again was most pronounced in mice with the smallest tumors.
Taken together, this dataset demonstrates how ELISpot/FluoroSpot can further reveal drug effects on the tumor-specific immune response. Moreover, because the assay can be performed with small volumes of blood, ELISpot/FluoroSpot can be applied to efficacy arms longitudinally. Thus, opportunity to gain mechanistic insight on drug activity without adding a second sampling arm to the study is provided. For more information on how ELISpot/FluoroSpot can be applied to your preclinical oncology and immuno-oncology research, contact the scientists at Labcorp Preclinical Oncology.