Targeting Protein Tyrosine Phosphatases to Improve Cancer Immunotherapies

Advances in immunotherapy have brought significant therapeutic benefits to many cancer patients. Nonetheless, many cancer types are refractory to current immunotherapeutic approaches, meaning that further targets are required to increase the number of patients who benefit from these technologies. Protein tyrosine phosphatases (PTPs) have long been recognised to play a vital role in the regulation of cancer cell biology and the immune response. In this review, we summarize the evidence for both the pro-tumorigenic and tumour-suppressor function of non-receptor PTPs in cancer cells and discuss recent data showing that several of these enzymes act as intracellular immune checkpoints that suppress effective tumour immunity. We highlight new data showing that the deletion of inhibitory PTPs is a rational approach to improve the outcomes of adoptive T cell-based cancer immunotherapies and describe recent progress in the development of PTP inhibitors as anti-cancer drugs

Deletion of the protein tyrosine phosphatase PTPN22 for adoptive T cell therapy facilitates CTL effector function but promotes T cell exhaustion

Background: Adoptive cell therapy (ACT) is a promising strategy for treating cancer, yet it faces several challenges such as lack of long-term protection due to T cell exhaustion induced by chronic TCR stimulation in the tumor microenvironment. One benefit of ACT, however, is that it allows for cellular manipulations, such as deletion of the phosphotyrosine phosphatase non-receptor type 22 (PTPN22), which improves CD8+ T cell antitumor efficacy in ACT. We tested whether Ptpn22KO cytolytic T cells (CTLs) were also more effective than Ptpn22WT CTL in controlling tumors in scenarios that favor T cell exhaustion. Methods: Tumor control by Ptpn22WT and Ptpn22KO CTL was assessed following adoptive transfer of low numbers of CTL to mice with subcutaneously implanted MC38 tumors. Tumor infiltrating lymphocytes were isolated for analysis of effector functions. An in vitro assay was established to compare CTL function in response to acute and chronic restimulation with antigen-pulsed tumor cells. The expression of effector and exhaustion-associated proteins by Ptpn22WT and Ptpn22KO T cells was followed over time in vitro and in vivo using the ID8 tumor model. Finally, the effect of PD-1 and TIM-3 blockade on Ptpn22KO CTL tumor control was assessed using monoclonal antibodies and CRISPR/Cas9-mediated knockout. Results: Despite having improved effector function at the time of transfer, Ptpn22KO CTL became more exhausted than Ptpn22WT CTL, characterized by more rapid loss of effector functions, and earlier and higher expression of inhibitory receptors (IRs), particularly the terminal exhaustion marker TIM-3. TIM-3 expression, under the control of the transcription factor NFIL3, was induced by IL-2 signaling which was enhanced in Ptpn22KO cells. Antitumor responses of Ptpn22KO CTL were improved following PD-1 blockade in vivo, yet knockout or antibody-mediated blockade of TIM-3 did not improve but further impaired tumor control, indicating TIM-3 signaling itself did not drive the diminished function seen in Ptpn22KO CTL. Conclusions: This study questions whether TIM-3 plays a role as an IR and highlights that genetic manipulation of T cells for ACT needs to balance short-term augmented effector function against the risk of T cell exhaustion in order to achieve longer-term protection

OT-I TCR Transgenic Mice to Study the Role of PTPN22 in Anti-cancer Immunity

Phosphotyrosine phosphatase non-receptor type 22 (PTPN22) is a key regulator of immune cell activation and responses. Genetic polymorphisms of PTPN22 have been strongly linked with an increased risk of developing autoimmune diseases, while analysis of PTPN22-deficient mouse strains has determined that PTPN22 serves as a negative regulator of T cell antigen receptor signaling. As well as these key roles in maintaining immune tolerance, PTPN22 acts as an intracellular checkpoint for T cell responses to cancer, suggesting that PTPN22 might be a useful target to improve T cell immunotherapies. To assess the potential for targeting PTPN22, we have crossed Ptpn22-deficient mice to an OT-I TCR transgenic background and used adoptive T cell transfer approaches in mouse cancer models. We provide basic methods for the in vitro expansion of effector OT-I cytotoxic T lymphocytes, in vitro phenotypic analysis, and in vivo adoptive T cell transfer models to assess the role of PTPN22 in anti-cancer immunity

Systemic inflammation, the peripheral blood transcriptome and primary melanoma

Peripheral blood transcriptomes from 383 newly-diagnosed melanoma patients were subjected to differential gene expression analysis. The hypotheses were that impaired systemic immunity is associated with poorer prognosis (thicker tumors and fewer tumor infiltrating lymphocytes (TILs)) and evidence of systemic inflammation (high-sensitivity C-reactive protein (hsCRP) and fibrinogen levels). Higher fibrinogen levels were associated with thicker primaries. In single gene analysis hsCRP levels were significantly associated with higher blood CD274 expression, (coding for PD-L1), but each was independently prognostic, with hsCRP associated with increased mortality, and higher CD274 protective, independent of age. Pathway analysis identified downregulation of immune cell signalling pathways in the blood of people with thicker tumors and notable upregulation of STAT1 in people with brisk TILs. Transcriptomic data provided evidence for increased NFB signalling with higher inflammatory markers but with reduction in expression of HLA class II molecules and higher CD274 suggesting that aberrant systemic inflammation is a significant mediator of reduced immune function in melanoma. In summary, transcriptomic data revealed evidence of reduced immune function in patients with thicker tumors and fewer TILs, at diagnosis. Inflammatory markers were associated with thicker primaries and independently with death from melanoma suggesting that systemic inflammation contributes to that reduced immune function

Costimulatory molecule DNAM-1 is essential for optimal differentiation of memory natural killer cells during mouse cytomegalovirus infection.

Recent studies demonstrate that natural killer (NK) cells have adaptive immune features. Here, we investigated the role of the costimulatory molecule DNAM-1 in the differentiation of NK cells in a mouse model of cytomegalovirus (MCMV) infection. Antibody blockade of DNAM-1 suppressed the expansion of MCMV-specific Ly49H(+) cells during viral infection and inhibited the generation of memory NK cells. Similarly, DNAM-1-deficient (Cd226(-/-)) Ly49H(+) NK cells exhibited intrinsic defects in expansion and differentiation into memory cells. Src-family tyrosine kinase Fyn and serine-threonine protein kinase C isoform eta (PKCη) signaling through DNAM-1 played distinct roles in the generation of MCMV-specific effector and memory NK cells. Thus, cooperative signaling through DNAM-1 and Ly49H are required for NK cell-mediated host defense against MCMV infection