Tumor-Specific Antibody, Cetuximab, Enhances the In Situ Vaccine Effect of Radiation in Immunologically Cold Head and Neck Squamous Cell Carcinoma

Abstract

In head and neck squamous cell carcinoma (HNSCC) tumors that over-expresses huEGFR, the anti-EGFR antibody, cetuximab, antagonizes tumor cell viability and sensitizes to radiation therapy. However, the immunologic interactions between cetuximab and radiation therapy are not well understood. We transduced two syngeneic murine HNSCC tumor cell lines to express human EGFR (MOC1- and MOC2-huEGFR) in order to facilitate evaluation of the immunologic interactions between radiation and cetuximab. Cetuximab was capable of inducing antibody-dependent cellular cytotoxicity (ADCC) in MOC1- and MOC2-huEGFR cells but showed no effect on the viability or radiosensitivity of these tumor cells, which also express muEGFR that is not targeted by cetuximab. Radiation enhanced the susceptibility of MOC1- and MOC2-huEGFR to ADCC, eliciting a type I interferon response and increasing expression of NKG2D ligands on these tumor cells. Co-culture of splenocytes with cetuximab and MOC2-huEGFR cells resulted in increased expression of IFNγ in not only NK cells but also in CD8+ T cells, and this was dependent upon splenocyte expression of FcγR. In MOC2-huEGFR tumors, combining radiation and cetuximab induced tumor growth delay that required NK cells, EGFR expression, and FcγR on host immune cells. Combination of radiation and cetuximab increased tumor infiltration with NK and CD8+ T cells but not regulatory T cells. Expression of PD-L1 was increased in MOC2-huEGFR tumors following treatment with radiation and cetuximab. Delivering anti–PD-L1 antibody with radiation and cetuximab improved survival and resulted in durable tumor regression in some mice. Notably, these cured mice showed evidence of an adaptive memory response that was not specifically directed against huEGFR. These findings suggest an opportunity to improve the treatment of HNSCC by combining radiation and cetuximab to engage an innate anti-tumor immune response that may prime an effective adaptive immune response when combined with immune checkpoint blockade. It is possible that this approach could be extended to any immunologically cold tumor that does not respond to immune checkpoint blockade alone and for which a tumor-specific antibody exists or could be developed.

Multifunctional nanoparticle potentiates the in situ vaccination effect of radiation therapy and enhances response to immune checkpoint blockade

“…From the B78 melanoma re-challenged mice, we collected splenocytes and co-cultured them with B16 melanoma cells that are parental to B78 and share common TAAs (Fig. 5g ) 7 , 47 . Using flow cytometry on these B16-co-cultured splenocytes, we observed increased levels of early activation marker, CD69 + , on both CD4 + and CD8 + T cells from mice rendered disease-free by PIC + RT + anti-CTLA-4, as compared to B16-co-cultured splenocytes from naïve control mice (Fig.…”

Harnessing CD16-Mediated NK Cell Functions to Enhance Therapeutic Efficacy of Tumor-Targeting mAbs

“…Several lines of evidence support a crucial role for NK cells in the CD16-dependent therapeutic efficacy of tumor-targeting mAbs. The composition and quality of tumorinfiltrating immune compartment are modified upon mAb treatment; in particular, several studies reported that intratumoral and circulating NK cell numbers increase upon tumor-targeting mAb-based combination regimens and that this correlated with favorable outcome [95][96][97][98][99]. Moreover, the observation that anti-lymphoma activity of rituximab was improved in the presence of anti-KIR mAb (IPH2102-lirilumab), in a syngeneic murine lymphoma model [100], and the evidence that NKG2A blockade by specific mAb (IPH2201monalizumab) potentiated cetuximab-dependent NK antitumor functions [101], together with the observation that Treg-mediated suppression of ADCC correlates with a lower clinical efficacy in cetuximab-treated HNSCC patients [102], further stress the concept that CD16-dependent NK cell effector functions heavily contribute to explain the therapeutic efficacy of tumor-targeting mAbs.…”

Local TLR4 stimulation augments in situ vaccination induced via local radiation and anti-CTLA-4 checkpoint blockade through induction of CD8 T-cell independent Th1 polarization

“…Flow cytometry was performed as previously described, 32 using fluorescent beads (UltraComp Beads eBeads, 176 Invitrogen, #01-2222-42) to determine compensation and fluorescence minus one methodology to determine gating ( online supplemental figure S2 ). For in vivo analysis, tumors and tumor draining lymph nodes were harvested and gently dissociated.…”