Daniel J. Powell

Systematic analysis of CD39, CD103, CD137, and PD‐1 as biomarkers for naturally occurring tumor antigen‐specific TILs

Abstract The detection of tumor‐specific T cells in solid tumors is integral to interrogate endogenous antitumor responses and to advance downstream therapeutic applications. Multiple biomarkers are reported to identify endogenous tumor‐specific tumor‐infiltrating lymphocytes (TILs), namely CD137, PD‐1, CD103, and CD39; however, a direct comparison of these molecules has yet to be performed. We evaluated these biomarkers in primary human ovarian tumor samples using single‐cell mass cytometry to compare their relative phenotypic profiles, and examined their response to autologous tumor cells ex vivo. PD‐1 + , CD103 + , and CD39 + TILs all contain a CD137 + cell subset, while CD137 + TILs highly co‐express the aforementioned markers. CD137 + TILs exhibit the highest expression of cytotoxic effector molecules compared to PD‐1 + , CD103 + , or CD39 + TILs. Removal of CD137 + cells from PD‐1 + , CD103 + , or CD39 + TILs diminish their IFN‐γ secretion in response to autologous tumor cell stimulation, while CD137 + TILs maintain high HLA‐dependent IFN‐γ secretion. CD137 + TILs exhibited an exhausted phenotype but with CD28 co‐expression, suggesting possible receptiveness to reinvigoration via immune checkpoint blockade. Together, our findings demonstrate that the antitumor abilities of PD‐1 + , CD103 + , and CD39 + TILs are mainly derived from a subset of CD137‐expressing TILs, implicating CD137 as a more selective biomarker for naturally occurring tumor‐specific TILs.

Folate Receptor Beta as a Direct and Indirect Target for Antibody-Based Cancer Immunotherapy

Folate receptor beta (FRβ) is a folate binding receptor expressed on myeloid lineage hematopoietic cells. FRβ is commonly expressed at high levels on malignant blasts in patients with acute myeloid leukemia (AML), as well as on M2 polarized tumor-associated macrophages (TAMs) in the tumor microenvironment of many solid tumors. Therefore, FRβ is a potential target for both direct and indirect cancer therapy. We demonstrate that FRβ is expressed in both AML cell lines and patient-derived AML samples and that a high-affinity monoclonal antibody against FRβ (m909) has the ability to cause dose- and expression-dependent ADCC against these cells in vitro. Importantly, we find that administration of m909 has a significant impact on tumor growth in a humanized mouse model of AML. Surprisingly, m909 functions in vivo with and without the infusion of human NK cells as mediators of ADCC, suggesting potential involvement of mouse macrophages as effector cells. We also found that TAMs from primary ovarian ascites samples expressed appreciable levels of FRβ and that m909 has the ability to cause ADCC in these samples. These results indicate that the targeting of FRβ using m909 has the potential to limit the outgrowth of AML in vitro and in vivo. Additionally, m909 causes cytotoxicity to TAMs in the tumor microenvironment of ovarian cancer warranting further investigation of m909 and its derivatives as therapeutic agents in patients with FRβ-expressing cancers.

PTEN Loss and BRCA1 Promoter Hypermethylation Negatively Predict for Immunogenicity in BRCA-Deficient Ovarian Cancer

PURPOSE Ovarian cancers can exhibit a prominent immune infiltrate, but clinical trials have not demonstrated substantive response rates to immune checkpoint blockade monotherapy. We aimed to understand genomic features associated with immunogenicity in BRCA1/2 mutation–associated cancers. MATERIALS AND METHODS Using the Cancer Genome Atlas whole-exome sequencing, methylation, and expression data, we analyzed 66 ovarian cancers with either germline or somatic loss of BRCA1/2 and whole-exome sequencing, immunohistochemistry, and CyTOF in 20 ovarian cancers with germline BRCA1/2 pathogenic variants from Penn. RESULTS We found two groups of BRCA1/2 ovarian cancers differing in their immunogenicity: (1) 37 tumors significantly enriched for PTEN loss (11, 30%) and BRCA1 promoter–hypermethylated (10, 27%; P = .0016) and (2) PTEN wild-type (28 of 29 tumors) cancers, with the latter group having longer overall survival (OS; P = .0186, median OS not reached v median OS = 66.1 months). BRCA1/2-mutant PTEN loss and BRCA1 promoter–hypermethylated cancers were characterized by the decreased composition of lymphocytes estimated by gene expression ( P = .0030), cytolytic index ( P = .034), and cytokine expression but higher homologous recombination deficiency scores ( P = .00013). Large-scale state transitions were the primary discriminating feature ( P = .001); neither mutational burden nor neoantigen burden could explain differences in immunogenicity. In Penn tumors, PTEN loss and high homologous recombination deficiency cancers exhibited fewer CD3+ ( P = .05), CD8+ ( P = .012), and FOXP3+ ( P = .0087) T cells; decreased PRF1 expression ( P = .041); and lower immune costimulatory and inhibitory molecule expression. CONCLUSION Our study suggests that within ovarian cancers with genetic loss of BRCA1/2 are two subsets exhibiting differential immunogenicity, with lower levels associated with PTEN loss and BRCA hypermethylation. These genomic features of BRCA1/2-associated ovarian cancers may inform considerations around how to optimally deploy immune checkpoint inhibitors in the clinic.

A B7-H4–Targeting Antibody–Drug Conjugate Shows Antitumor Activity in PARPi and Platinum-Resistant Cancers with B7-H4 Expression

Abstract Purpose: Platinum and PARP inhibitors (PARPi) demonstrate activity in breast and ovarian cancers, but drug resistance ultimately emerges. Here, we examine B7-H4 expression in primary and recurrent high-grade serous ovarian carcinoma (HGSOC) and the activity of a B7-H4-directed antibody–drug conjugate (B7-H4-ADC), using a pyrrolobenzodiazepine-dimer payload, in PARPi- and platinum-resistant HGSOC patient-derived xenograft (PDX) models. Experimental Design: B7-H4 expression was quantified by flow cytometry and IHC. B7-H4-ADC efficacy was tested against multiple cell lines in vitro and PDX in vivo. The effect of B7-H4-ADC on cell cycle, DNA damage, and apoptosis was measured using flow cytometry. Results: B7-H4 is overexpressed in 92% of HGSOC tumors at diagnosis (n = 12), persisted in recurrent matched samples after platinum treatment, and was expressed at similar levels across metastatic sites after acquired multi-drug resistance (n = 4). Treatment with B7-H4-ADC resulted in target-specific growth inhibition of multiple ovarian and breast cancer cell lines. In platinum- or PARPi-resistant ovarian cancer cells, B7-H4-ADC significantly decreased viability and colony formation while increasing cell-cycle arrest and DNA damage, ultimately leading to apoptosis. Single-dose B7-H4-ADC led to tumor regression in 65.5% of breast and ovarian PDX models (n = 29), with reduced activity in B7-H4 low or negative models. In PARPi and platinum-resistant HGSOC PDX models, scheduled B7-H4-ADC dosing led to sustained tumor regression and increased survival. Conclusions: These data support B7-H4 as an attractive ADC target for treatment of drug-resistant HGSOC and provide evidence for activity of an ADC with a DNA-damaging payload in this population. See related commentary by Veneziani et al., p. 1434

A phase 1 trial of adoptive transfer of vaccine-primed autologous circulating T cells in ovarian cancer

We have previously shown that vaccination with tumor-pulsed dendritic cells amplifies neoantigen recognition in ovarian cancer. Here, in a phase 1 clinical study ( NCT01312376 /UPCC26810) including 19 patients, we show that such responses are further reinvigorated by subsequent adoptive transfer of vaccine-primed, ex vivo-expanded autologous peripheral blood T cells. The treatment is safe, and epitope spreading with novel neopeptide reactivities was observed after cell infusion in patients who experienced clinical benefit, suggesting reinvigoration of tumor-sculpting immunity.