PD-L1 and PD-L2 are ligands for the PD-1 immune inhibiting checkpoint that can be induced in tumors by interferon exposure, leading to immune evasion. This process is important for immunotherapy ...based on PD-1 blockade. We examined the specific molecules involved in interferon-induced signaling that regulates PD-L1 and PD-L2 expression in melanoma cells. These studies revealed that the interferon-gamma-JAK1/JAK2-STAT1/STAT2/STAT3-IRF1 axis primarily regulates PD-L1 expression, with IRF1 binding to its promoter. PD-L2 responded equally to interferon beta and gamma and is regulated through both IRF1 and STAT3, which bind to the PD-L2 promoter. Analysis of biopsy specimens from patients with melanoma confirmed interferon signature enrichment and upregulation of gene targets for STAT1/STAT2/STAT3 and IRF1 in anti-PD-1-responding tumors. Therefore, these studies map the signaling pathway of interferon-gamma-inducible PD-1 ligand expression.
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•PD-L1 is primarily regulated by interferon gamma signaling in melanoma cells•PD-L2 is regulated by both interferon beta and gamma signaling•Regulation of PD-1 ligands works mainly through the JAK1/2-STAT1/3-IRF1 axis
Garcia-Diaz et al. performed a small hairpin RNA screen and genetic and functional studies to map the signaling pathways that result in reactive PD-L1 and PD-L2 on melanoma cells upon interferon gamma exposure. The authors highlight the importance of the JAK1/JAK2-STAT1/STAT2/STAT3-IRF1 axis for clinical responses to PD-1 blockade therapy.
Interleukin-2 (IL-2) is a component of most protocols of adoptive cell transfer (ACT) therapy for cancer, but is limited by short exposure and high toxicities. NKTR-214 is a kinetically-engineered ...IL-2 receptor βγ (IL-2Rβγ)-biased agonist consisting of IL-2 conjugated to multiple releasable polyethylene glycol chains resulting in sustained signaling through IL-2Rβγ. We report that ACT supported by NKTR-214 increases the proliferation, homing and persistence of anti-tumor T cells compared to ACT with IL-2, resulting in superior antitumor activity in a B16-F10 murine melanoma model. The use of NKTR-214 increases the number of polyfunctional T cells in murine spleens and tumors compared to IL-2, and enhances the polyfunctionality of T and NK cells in the peripheral blood of patients receiving NKTR-214 in a phase 1 trial. In conclusion, NKTR-214 may have the potential to improve the antitumor activity of ACT in humans through increased in vivo expansion and polyfunctionality of the adoptively transferred T cells.
Interleukin-2 (IL-2) is a cytokine required for effector T cell expansion, survival, and function, especially for engineered T cells in adoptive cell immunotherapy, but its pleiotropy leads to ...simultaneous stimulation and suppression of immune responses as well as systemic toxicity, limiting its therapeutic use. We engineered IL-2 cytokine-receptor orthogonal (
) pairs that interact with one another, transmitting native IL-2 signals, but do not interact with their natural cytokine and receptor counterparts. Introduction of
IL-2Rβ into T cells enabled the selective cellular targeting of
IL-2 to engineered CD4
and CD8
T cells in vitro and in vivo, with limited off-target effects and negligible toxicity.
IL-2 pairs were efficacious in a preclinical mouse cancer model of adoptive cell therapy and may therefore represent a synthetic approach to achieving selective potentiation of engineered cells.
Cancer testis antigens (CTA) are a large family of tumor-associated antigens expressed in human tumors of different histological origin, but not in normal tissues except for testis and placenta. This ...tumor-restricted pattern of expression, together with their strong
in vivo immunogenicity, identified CTA as ideal targets for tumor-specific immunotherapeutic approaches, and prompted the development of several clinical trials of CTA-based vaccine therapy. Driven by this practical clinical interest, a more detailed characterization of CTA biology has been recently undertaken. So far, at least 70 families of CTA, globally accounting for about 140 members, have been identified. Most of these CTA are expressed during spermatogenesis, but their function is still largely unknown. Epigenetic events, particularly DNA methylation, appear to be the primary mechanism regulating CTA expression in both normal and transformed cells, as well as in cancer stem cells. In view of the growing interest in CTA biology, the aim of this review is to provide the most recent information on their expression, regulation and function, together with a brief summary of the major clinical trials involving CTA as therapeutic agents. The pharmacologic modulation of CTA expression profiles on neoplastic cells by DNA hypomethylating drugs will also be discussed as a feasible approach to design new combination therapies potentially able to improve the clinical efficacy of currently adopted CTA-based immunotherapeutic regimens in cancer patients.
Purpose
Pharmacologic DNA hypomethylation holds strong promises in cancer immunotherapy due to its immunomodulatory activity on neoplastic cells. Searching for more efficient DNA hypomethylating ...agents to be utilized to design novel immunotherapeutic strategies in cancer, we investigated the immunomodulatory properties of the new DNA hypomethylating agent SGI-110, that is resistant to in vivo inactivation by cytidine deaminase.
Experimental design
Cutaneous melanoma, mesothelioma, renal cell carcinoma, and sarcoma cells were treated in vitro with SGI-110. RT-PCR, quantitative RT-PCR, quantitative methylation-specific PCR, and flow cytometric analyses were performed to investigate changes induced by SGI-110 in the constitutive immune profile of cancer cells. The recognition by gp100-specific CTL of gp100-positive melanoma cells, treated or not with SGI-110, was tested by LDH release assays.
Results
SGI-110 induced/up-regulated the expression of investigated cancer/testis antigens (CTA) (i.e., MAGE-A1, MAGE-A2, MAGE-A3, MAGE-A4, MAGE-A10, GAGE 1-2, GAGE 1-6, NY-ESO-1, and SSX 1-5) in all cancer cell lines studied, both at mRNA and at protein levels. Quantitative methylation-specific PCR analyses identified a hypomethylation of MAGE-A1 and NY-ESO-1 promoters in SGI-110-treated neoplastic cells, demonstrating a direct role of pharmacologic DNA demethylation in CTA induction. SGI-110 also up-regulated the expression of HLA class I antigens and of ICAM-1, resulting in an improved recognition of cancer cells by gp100-specific CTL.
Conclusions
Our findings show that SGI-110 is a highly attractive therapeutic agent to comprehensively increase immunogenicity and immune recognition of neoplastic cells, and provide the scientific rationale for its clinical development to design novel chemo-immunotherapeutic approaches in cancer patients.
In a global warming context, the advancement and compression of maturity in early ripening grape varieties suited to sparkling wine making can easily expedite harvest within the first two weeks of ...August (in northern Italy). Such earliness, albeit safeguarding acid retention, involves potential suboptimal grape composition and triggers logistical issues related to grape harvest and delivery. Hence, this three-year study beginning in 2020 was conducted on cane-pruned Chardonnay vines, grown in the Franciacorta district of Lombardy, to assess if a single-step delayed winter pruning was able to postpone vine phenology and ripening, without harming yield potential. Control (C) vines pruned midwinter were compared with those subjected to late winter pruning (LWP), performed when the apical shoots growing on the unpruned canes had reached the 2-3 unfolded leaf stage (T1) as well as 7-8 days later (T2). In 2022, a fourth treatment was added, consisting of a two-step procedure with finishing performed at T1 (LWP-two canes). Vegetative growth, yield components, ripening dynamics, and maturity at harvest were followed in each year of this study. Budburst delay induced by LWP treatments across seasons varied between 4 and 9 days, whereas harvest was postponed by 5–14 days. The extent of delay was especially pronounced in 2020, when the removed leaf area (LA) was also found to be the highest. The higher the removed LA, the higher the yield constrain was in the concomitant year. In 2020 and 2021, despite the harvest delay, LWP vines were able to assure full matching with desirable features in must composition, set at total soluble solids (TSS) of about 18°Brix and a titratable acidity (TA) of at least 8 g/L. In the very hot and dry phase of 2022, none of the treatments facilitated the required ripening status, while data showed that an even earlier ripening would not have allowed the harvest to reach the minimum TSS level. Notably, even in 2021 and 2022, when the removed LA was quite low, a delaying effect was prompted in the seasonal trends of all main ripening parameters. This suggested that besides the amount of LA removal, other unknown factors drive the postponement of ripening. In 2022, the behavior of added-treatment LWP-two canes was found to be quite similar to that of the C vines. Depending upon the desired yield level and local climate feature, the protocol entailing a single-step late winter pruning, performed at any time between the T1-T2 time window used in this study, is deemed as effective in achieving a significant harvest delay while maintaining or even improving the compositional patterns recorded for midwinter pruned vines.
BackgroundOvercoming the suppressive tumor microenvironment (TME) remains an important unmet challenge for chimeric antigen receptor (CAR) T cell therapies. A key suppressive factor, transforming ...growth factor β (TGF-β), is a primary driver of T cell suppression reducing T cell receptor (TCR)-mediated cytotoxicity and driving the differentiation of immunosuppressive regulatory T cells (Tregs) in the TME.MethodsTo overcome TGF-β-specific immunosuppression, we employed a TGF-β-targeting CAR co-expressed in T cells with a human papillomavirus type 16 (HPV16) E711-19-specific, HLA-A*02:01-restricted TCR (E7 TCR).ResultsIn comparison to T cells expressing E7 TCR alone or E7 TCR with an irrelevant CD19-targeting CAR, T cells co-transduced with E7 TCR and TGF-β CAR showed enhanced proliferation and cytokine production, while maintained cytotoxicity throughout repeat antigen challenge assays with HPV16+ Ca Ski tumor cells. The inclusion of TGF-β CAR also reduced PD1+ expression and Treg differentiation after repeat antigen challenges. Transcriptional analysis further confirmed reduced FOXP3 expression as well as enhanced proinflammatory genes such as TNF and IFNG.ConclusionsIn combination, these data clearly show that a TGF-β CAR can enhance TCR function and limit Treg differentiation and is therefore likely to improve the function and persistence of TCR therapies in the TME.
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