Immune checkpoint inhibitors (ICI) are highly effective for a minority of patients with cancer. Pre-clinical research into mechanisms of ICI resistance is reliant upon anti-PD-1 or anti-PD-L1 antibody treatment in mouse models of cancer. Common approaches utilize commercially-available rat isotype anti-PD-L1 antibodies given by intraperitoneal (IP) injection at relatively high doses (5mg/kg every 2-3 days), often for extended periods. However, there has been limited investigation into the pharmacokinetics of these antibodies with single and repeat dosing, which could inform rational dose scheduling. Non-species-matched antibodies are known to induce accelerated anti-drug immune responses, compared to species-matched antibodies. As a result, we hypothesized that matched-isotype anti-PD-L1 (based on the grafting of atezolizumab variable regions onto a mouse IgG2a scaffold) and unmatched-isotype rat IgG2b anti-PD-L1 would be associated with distinct pharmacokinetics on single and repeated dosing. We first confirmed that both matched and unmatched antibodies have nanomolar-range affinity for mouse PD-L1 by surface plasmon resonance (0.85nM and 0.37nM, respectively). Using a single IP injection in 8-week-old male C57BL/6J mice followed by repeated tail-vein plasma sampling and an indirect anti-PD-L1 ELISA, we detected a trend towards prolonged median half-life for the matched (n=17 mice) versus unmatched (n=19) antibody (91.5 versus 39.0 hours). We detected a particularly wide range of half-life measurements for matched antibody experiments (19.8 to 337.7 hours), suggesting that antibody clearance can be highly heterogeneous. Using repeated IP injection (2 doses, 14 days apart), we identified markedly accelerated antibody clearance for both antibodies resulting in comparable antibody half-lives after dose 2 (11.9 versus 12.5 hours, n=13 and n=8 mice, respectively), suggestive of a time-dependent anti-drug adaptive immune response. These findings have significant implications for in vivo antibody treatment experiments, demonstrating that repeat dosing is associated with rapid antibody clearance, irrespective of the antibody species. These data should be taken into consideration when planning longer-term antagonism of PD-1/PD-L1 signaling with antibodies in mice and strengthen the basis for short term window studies that examine the intratumoral immune biology. Furthermore, the high variability in antibody half-life suggests that antibody pharmacokinetics could confound ICI treatment response studies in vivo. Citation Format: Breanna Demestichas, Sam Kleeman, Matthew Chvasta, Tobias Janowitz. Comparative mouse pharmacokinetics of matched- and unmatched-isotype anti-PD-L1 antibodies. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 5672.
Immune checkpoint inhibitors (ICI) targeting PD-1/PD-L1 and CTLA4 are effective treatments in multiple cancer types, but durable responses are seen only in a minority of patients. Resistance to ICI is likely to reflect a combination of tumor-intrinsic, such as tumor mutational burden, and tumor-extrinsic factors, such as systemic immune suppression and host genetics. With the advent of large-scale germline genotyping studies, there is emerging evidence that host-specific genetic factors can substantially modify both disease risk and responsiveness to therapy. There has been limited characterization to date of the interaction between host genetics and response to ICI. Here, we perform harmonized mutation calling in available normal exome sequencing from 668 metastatic cancer patients treated with ICI (derived from n=7 studies, spanning n=3 cancer types and PD-1/PD-L1/CTLA4-directed therapy) and performed exome-wide association studies in European ancestry patients. For each cancer type (melanoma, renal cell carcinoma, urothelial carcinoma), we conducted exome-wide Cox regression (with PC1-4 and sex as covariates) to identify coding single-nucleotide polymorphisms (SNPs) associated with overall survival (OS). On fixed-effect meta-analysis, we identified a single coding variant (rs140221307, C allele, MAF 0.5% in European populations) that met genome-wide significance (p=1.11 × 10−15, HR=4.86, 95% CI 3.30-7.15). This SNP causes a substitution of Trp to Arg at amino acid 320 at the interface between the transmembrane and extracellular domains of interleukin 17 receptor A (IL17RA). While the secretion mechanism is uncharacterized, soluble IL17RA can be readily detected in plasma and the survival-reducing variant is associated with substantially elevated plasma IL17RA (p=1.90 × 10−9, beta=1.01, Sun et al. 2018) as well as reduced monocyte counts in UK Biobank (p=2.20 × 10−308, beta=-0.60). We have established a prime editing strategy to induce this variant in cultured human monocytes to investigate its biochemical and functional consequences, and are performing validation studies in independent patient cohorts. Soluble IL17RA would be expected to function as a decoy receptor to antagonize IL17/IL17RA signaling, and so these data suggest that activation of IL-17 receptor signaling might improve responsiveness to ICI therapy. Citation Format: Sam O. Kleeman, Michael Chan, Matthew Chvasta, Tobias Janowitz. Exome-wide association study identifies coding variant in IL17RA associated with survival in cancer patients treated with immunotherapy. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 5226.
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