Abstract B051: Antitumor activity of dual PD-1 and LAG-3 blockade reversed by adoptive transfer of tumor-draining lymph node cells
Authors: Liang Wei , Valdovinos Lilibet , Yapjoco Kaitlyn , Hinata Kaede , Sharma Geeta , Broudy Thomas B 
Published: March 2020

Immune checkpoint inhibitors (ICIs) emerged as promising therapies for various cancer types. Yet, therapeutics that target single immune checkpoint protein (ICP) have met with obstacles including low response rates and acquired tumor resistance. To overcome these obstacles, combination therapies that simultaneously target multiple ICPs are being actively pursued in the field. In this study, we utilized the B16F10 tumors, a syngeneic mouse melanoma model that showed limited responses to numerous ICIs against single ICP, and assessed the efficacy of dual blockade of PD-1 and LAG-3 (an ICP expressed on the surface of T and NK cells) through i.p. injection of neutralizing antibodies [anti-PD-1 (clone: RMP1-14) and anti-LAG-3 (clone: C9B7W), 10 mg/kg each, twice weekly]. Moreover, to interrogate the effects of altered immune landscape in the tumor microenvironment (TME) on tumor responses to ICIs, we performed a single-time adoptive transfer of tumor-draining lymph node (TDLN) cells harvested from B16F10 tumor-bearing donor mice (5 M TDLN cells per recipient mouse, i.v.), 2 days before the first antibody treatments, and assessed its impact on tumor responses in recipient mice. TDLN cells were either used immediately after isolation or cultured in vitro with anti-CD3/IL-2 for 5 days that resulted in rapid proliferation and overpopulation (&gt;98%) of T cells. Our results showed that combination treatment with anti-PD-1 and anti-LAG-3, compared to isotype controls, significantly inhibited B16F10 tumor growth (&gt;60%) in mice that did not received donor TDLN cells. FACS analyses of blood and tumor samples indicated that anti-PD-1/anti-LAG-3 significantly enhanced the percentages of CD8+ T cells and CD49b+ NK cells as well as the ratio between CD8+ T cells and CD4+CD25+CD127-/low regulatory T cells, in B16F10 tumors but not in the peripheral blood. Interestingly, adoptive transfer of freshly-isolated TDLN cells completely abrogated the antitumor activity, whereas infusion with anti-CD3/IL-2-treated TDLN cells only partially alleviated the antitumor activity of anti-PD-1/anti-LAG-3. FACS analyses indicated that, in mice receiving donor TDLN cells, anti-PD-1/anti-LAG-3 was not able to stimulate intratumoral infiltration of CD8+ T cells or NK cells or increase the ratio between CD8+ T cells and regulatory T cells. In conclusion, combined treatment of anti-PD-1 and anti-LAG-3 significantly inhibits B16F10 tumor growth associated with activation of immune responses in the TME, which is at least partially reversed by adoptive transfer of TDLN cells. Our results support the usage of the B16F10 mouse melanoma model for testing the efficacy of combination therapies or bi-specific antibodies against both PD-1 and LAG-3. The reason behind tumor resistance induced by adoptive transfer of TDLN cells remains unclear. It is important to identify this mechanism of resistance, as the knowledge can benefit patient selection or provide a rationale for future combination therapies that concomitantly target PD-1/LAG-3 and this resistance machinery.

Citation Format: Wei Liang, Lilibet Valdovinos, Kaitlyn Yapjoco, Kaede Hinata, Geeta Sharma, Thomas B Broudy. Antitumor activity of dual PD-1 and LAG-3 blockade reversed by adoptive transfer of tumor-draining lymph node cells [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference on Molecular Targets and Cancer Therapeutics; 2019 Oct 26-30; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2019;18(12 Suppl):Abstract nr B051. doi:10.1158/1535-7163.TARG-19-B051