Enhancement Of Immunotherapy Research Articles

Multi-omics analysis reveals the unique landscape of DLD in the breast cancer tumor microenvironment and its implications for immune-related prognosis

BackgroundCuproptosis, i.e., copper-induced programmed cell death, has potential implications in cancer therapy. However, the impact of the cuproptosis-related gene (CRG) dihydrolipoyl dehydrogenase (DLD) on breast cancer (BC) prognosis remains underexplored. MethodsWe employed real-time quantitative PCR and multiplexed immunostaining techniques to quantify DLD expression in both BC and the adjacent non-cancerous tissues. Immunofluorescence analysis was employed to assess the influence of DLD on immune cells and immunological checkpoints in the BC microenvironment. DLD knockdown experiments were conducted in BC cell lines MDA-MB-468 and SK-BR-3, with knockdown efficiency validated via western blot. Subsequently, we performed the cell counting kit-8 (CCK-8) assay, clone formation assay, Transwell migration assay, and invasion assay. To construct a prognostic model, we employed a Lasso-Cox regression analysis of immune-related genes associated with DLD. Additionally, we established a competing endogenous RNA network based on CRGs to evaluate potential regulatory pathways. ResultsCompared to the adjacent tissues, BC tissues exhibited markedly elevated DLD expression levels. In vitro experiments demonstrated that DLD knockdown effectively inhibited BC cell migration, invasion, and proliferation. DLD exhibited positive correlations with CD68+ macrophages and PD-L1 in the tumor, as well as with macrophages and CD4+ T cells in the stroma. Tumor regions with high DLD expression were enriched in PD-L1 and macrophages, while stromal regions with high DLD expression contained CD4+ T cells and macrophages. The AUC values for 1-, 3-, and 5-year overall survival in TCGA-BRCA training set were 0.67, 0.66, and 0.66, respectively. A nomogram with a C-index of 0.715 indicated that risk score, tumor stage, and age could serve as independent prognostic factors for BC. ConclusionOur findings underscore the significant predictive significance of DLD in BC and its influence on the tumor microenvironment. DLD represents a promising diagnostic and prognostic marker for BC, offering novel avenues for the identification of therapeutic targets and the enhancement of immunotherapy in BC.

Triply Enhanced Immunotherapy via Dual Glycan Reforming Integrated with Perforation.

The enhancement of immunotherapy is an emerging direction to develop highly effective and practical cancer therapeutic methods. Here a triply enhanced immunotherapy drug (TEID) is designed for ingeniously integrating in situ dual glycan reforming with perforation on cell membrane. The TEID is composed of galactose and neuraminidase conjugated streptolysin O (SLO-Gal and SLO-NEU), which are encapsulated in a hyaluronic acid (HA) shell for targeted recognition to tumor tissue via cell surface CD44. After targeted delivery and HAase-mediated degradation in the tumor region, the TEID releases SLO-Gal and SLO-NEU, which can easily anchor Gal and NEU on the tumor cell membrane via the perforation of SLO to perform dual glycan reforming for the introduction of Gal and the cleavage of sialic acid. The former can activate immune cells to secret cytokines for immune-killing, and the latter can weaken the immune inhibition to improve the immunotherapeutic efficacy. Meanwhile, the perforation of SLO can promote the delivery of cytokines into the tumor cells to further enhance the efficacy. The designed triply enhanced immunotherapy strategy opens a significant and promising route to promote clinical immunotherapy of cancer.

Abstract 2161: Fasting fueled ketogenesis inhibits ovarian cancer and promotes anti-tumor T cell response

Abstract Epithelial Ovarian Cancer (EOC) is the most lethal gynecologic cancer with no predominant genetic alteration identified that can be therapeutically targeted. Low-cost non-toxic approaches such as dietary modulations alter the responses of host and the cancer cells. Limited scientific investigations have examined the link between nutrition and EOC. Here, we tested the outcome of intermittent fasting (IF) on EOC progression with focus on fasting driven anti-tumor immune responses. Female C57/B6 mice were injected intraperitoneally with 5x106 mouse epithelial ovarian cancer ID8 (p53-/-, ID8 p53-/-, PTEN-/- and ID8 p53-/-, BRCA-/-) cells and fed either ad libitum (RD) or subjected to IF, where they underwent 16 hour fasting and 8 hour feeding window for 5 days/week. Tumor progression was monitored by ascites formation and abdominal circumference. Immune profiling was performed by flow cytometry using BD FACS Calibur. Untargeted metabolomics was performed in plasma from RD and IF mice using LC-MS/MS. Enhancement of immunotherapy was tested by combining IF with anti-PD 1 (100 µg/mice, every 5th day, 3 times) in ID8 p53-/- tumor bearing mice and similar parameters were examined. IF reduced the growth of ID8 (p53-/-, ID8 p53-/-, PTEN-/- and ID8 p53-/-, BRCA-/-) tumors, increased overall survival when compared to RD mice. IF promoted anti-tumor T cell response as seen by increase in CD4+ and CD8+ cells with enhanced expression of IFNγ, granzyme B and perforin compared to RD mice bearing ID8 cells in the ascites and blood. A striking observation was that the CD4+ T cells from IF mice exhibited a strong Th1 phenotype (CD4+IFNγ+), while Th2 phenotype (CD4+IL4+) was significantly decreased. IF when combined with anti-PD-1 treatment significantly improved survival and potentiated the T cell mediated anti-tumor response. Metabolic analysis revealed an enrichment in ketone body biosynthetic pathway as reflected by 5fold increase in serum ketone bodies such as beta hydroxy butyrate (BHB) and acetoacetate, when compared to RD. Receptors that have been reported to response to BHB as a ligand, specifically GPR109A and GPR41, were highly expressed on T cells derived from IF mice. BHB treatment enhanced the CD8+ T cell effector function as reflected by increased levels of IFNγ, granzyme B and perforin compared to untreated CD8+ T cells. Our study shows that IF significantly restricts EOC growth, improve survival, potentiate chemotherapy and immunotherapy anti-tumor response by augmenting T cell mediated anti-tumor immune responses. We also provide evidence that IF generated bioactive metabolite BHB which can be a potential substitute for availing the ant-tumor benefits of IF. Thus, our data provides strong evidence for IF and its metabolic mediator BHB for ameliorating EOC progression and as a viable approach in maintaining and sustaining an effective anti-tumor T cell response alone and in combination with immunotherapy. Citation Format: Mary Priyanka Udumula, Laila Poisson, Lin Chun-Hui, Nivedita Tiwari, Harshit Singh, Giri Shailendra, Ramandeep Rattan. Fasting fueled ketogenesis inhibits ovarian cancer and promotes anti-tumor T cell response [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 2161.

Abstract 3960: Low doses of Docetaxel in combination with anti-prostate cancer immunotherapy lead to complete tumor-free outcome through Galectin-3 silencing

Abstract Based on animal model results and the analysis of clinical data from metastatic and castration resistant prostate cancer (mCRPC) patients, we were able to demonstrate the advantages and the mechanisms by which Docetaxel(DTX)-based chemotherapy induces the correct pre-conditioning of the tumour microenvironment to allow high cytotoxic T cell activity to avoid prostate tumour growth and recurrence. First, analysis of the gene expression of human PCa cells showed that treatment with DTX decreases the expression of Galectin-3 (Gal-3) in either chemo-sensitive or -resistant prostate cancer (PCa) cell lines. In addition, using a murine PCa model (derived from TRAMP mice), treatment of TRAMP-C1 (TC1) cells or tumors with a low dose of DTX lowered the expression of Gal-3 but not Gal-1 (among others) in this cell line and tumors as well (reversible silencing of about 98% (in vitro) and 55% (in vivo)). More importantly, Gal-3 downregulation was confirmed in clinical samples of mCRPC patients treated by taxane-based chemotherapy. To evaluate whether this DTX-dependent Gal-3 downregulation in PCa tumors could be one of the principal molecular factors responsible for chemotherapy enhancement of immunotherapy, we decide to inoculate mice with an autologous BM-DC vaccine loaded with lysates from TC1 cells pretreated by DTX in vitro or expressing an anti-Gal-3 shRNA. This protocol completely protects the animals to the growth of Gal-3 deficient-PCa tumors. We then decided to go further in the understanding of the mechanisms that promote this effective anti-PCa immune response, and analysed the proliferation ability of lymphocytes in the presence or absence of TC1 tumor cells, expressing or not Gal-3. The results were encouraging as we demonstrated that Gal-3 down-regulation in tumors (through the use of TC1-shGal-3 cells or TC1-treated with low doses of DTX) was sufficient to restore the capability of CD8+ T cells to be activated and to proliferate as well as in the absence of tumor cells. Interestingly, we showed by ex vivo cytotoxicity assays that these activated and proliferating CD8+ T lymphocytes after vaccination of mice with TC1-lysate loaded BM-DC are capable of killing Gal-3 downregulated TC1 cells and protect prostate cancer recurrence and metastasis development in mice after primary tumor-resection. In conclusion, our results strongly suggest that Gal-3 is one of the principal immunosuppressor in prostate cancer that limits the success of immunotherapy. We then show that DTX-based chemotherapy would promote a decrease of Gal-3 expression by the tumors to create a permissive tumor microenvironment for successful immunotherapy. Finally, we propose a rational protocol combining no toxic/low doses of Docetaxel with simplified immunotherapy to completely control tumor recurrence for all PCa patients and chemoresistant as well. Citation Format: Daniel Compagno, Carolina Tiraboschi, Lucas Gentilini, Enrique Corapi, Felipe M. Jaworski, Anne Chauchereau, Diego J. Laderach. Low doses of Docetaxel in combination with anti-prostate cancer immunotherapy lead to complete tumor-free outcome through Galectin-3 silencing [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 3960.

Analysis of the Expression of Surface Receptors on NK Cells and NKG2D on Immunocytes in Peripheral Blood of Patients with Nasopharyngeal Carcinoma

Background:The aberrant expression of surface receptors on immunocytes may represent potential markers of tumor escape for nasopharyngeal carcinoma (NPC). The aim of this study was to investigate the expression of representative receptors on natural killer (NK) cells and NK group 2, member D (NKG2D) on immunocytes in the peripheral blood of patients with NPC.Methods:Patients (n = 64) with NPC prior to initiation of treatment were defined as the study group. Healthy volunteers (n = 31) served as the control group. The expression of NK cells and NKT cells; the triggering receptors NKp30, NKp44, and NKp46 on NK cells; the activating receptor NKG2D on NK cells, CD4+ T cells, and CD8+ T cells; and the inhibitory receptors CD158b and CD159a on NK cells were analyzed by flow cytometry in the two groups.Results:Here, our study showed that no differences were observed in terms of the numbers of NK cells or NKT cells, or the expression of CD158b and CD159a on the surface of NK cells between the two groups. Nevertheless, the expression levels of NKp30 and NKp46 on NK cells in the NPC patients were significantly lower than in the healthy individuals (P < 0.05). No differences existed in the expression of NKG2D on NK cells, but NKG2D on CD8+ T cells showed a markedly lower expression in the study group (P < 0.001).Conclusions:Our findings may reflect a possible mechanism of immune evasion for NPC. The enhancement of immunotherapy concerning NKp30, NKp46, and NKG2D may be an innovative treatment strategy for patients with NPC.

Phase I Study of Recombinant Human Interleukin-7 Administration in Subjects with Refractory Malignancy

Interleukin-7 (IL-7) has critical and nonredundant roles in T-cell development, hematopoiesis, and postdevelopmental immune functions as a prototypic homeostatic cytokine. Based on a large body of preclinical evidence, it may have multiple therapeutic applications in immunodeficiency states, either physiologic (immunosenescence), pathologic (HIV), or iatrogenic (postchemotherapy and posthematopoietic stem cell transplant), and may have roles in immune reconstitution or enhancement of immunotherapy. We report here on the toxicity and biological activity of recombinant human IL-7 (rhIL-7) in humans. Subjects with incurable malignancy received rhIL-7 subcutaneously every other day for 2 weeks in a phase I interpatient dose escalation study (3, 10, 30, and 60 microg/kg/dose). The objectives were safety and dose-limiting toxicity determination, identification of a range of biologically active doses, and characterization of biological and, possibly, antitumor effects. Mild to moderate constitutional symptoms, reversible spleen and lymph node enlargement, and marked increase in peripheral CD3(+), CD4(+), and CD8(+) lymphocytes were seen in a dose-dependent and age-independent manner in all subjects receiving >or=10 microg/kg/dose, resulting in a rejuvenated circulating T-cell profile, resembling that seen earlier in life. In some subjects, rhIL-7 induced in the bone marrow a marked, transient polyclonal proliferation of pre-B cells showing a spectrum of maturation as well as an increase in circulating transitional B cells. This study shows the potent biological activity of rhIL-7 in humans over a well-tolerated dose range and allows further exploration of its possible therapeutic applications.

Perspective on Potential Clinical Applications of Recombinant Human Interleukin‐7

Interleukin-7 has critical and nonredundant roles in T cell development, hematopoiesis, and postdevelopmental immune functions as a prototypic homeostatic cytokine. Based on a large body of preclinical evidence, it may have multiple therapeutic applications in immunodeficiency states, either physiologic (immuno-senescence), pathologic (HIV) or iatrogenic (postchemotherapy and posthematopoietic stem cell transplant) and may have roles in immune reconstitution or enhancement of immunotherapy. Early clinical development trials in humans show that, within a short time, rhIL-7 administration results in a marked preferential expansion of both naive and memory CD4 and CD8 T cell pools with a tendency toward enhanced CD8 expansion. As a result, lymphopenic or normal older hosts develop an expanded circulating T cell pool with a profile that resembles that seen earlier in life with increased T cell repertoire diversity. These results, along with a favorable toxicity profile, open a wide perspective of potential future clinical applications.