Search

Type I interferon (IFN-I) and IFN-γ foster antitumor immunity by facilitating T cell responses. Paradoxically, IFNs may promote T cell exhaustion by activating immune checkpoints. The downstream regulators of these disparate responses are incompletely understood. Here, we describe how interferon regulatory factor 1 (IRF1) orchestrates these opposing effects of IFNs. IRF1 expression in tumor cells blocks Toll-like receptor- and IFN-I-dependent host antitumor immunity by preventing interferon-stimulated gene (ISG) and effector programs in immune cells. In contrast, expression of IRF1 in the host is required for antitumor immunity. Mechanistically, IRF1 binds distinctly or together with STAT1 at promoters of immunosuppressive but not immunostimulatory ISGs in tumor cells. Overexpression of programmed cell death ligand 1 (PD-L1) in Irf1-/- tumors only partially restores tumor growth, suggesting multifactorial effects of IRF1 on antitumor immunity. Thus, we identify that IRF1 expression in tumor cells opposes host IFN-I- and IRF1-dependent antitumor immunity to facilitate immune escape and tumor growth.

Background: S-588410, a cancer peptide vaccine (CPV), comprises five HLA-A*24:02-restricted peptides from five cancer-testis antigens. In a phase 2 study, S-588410 was well-tolerated and exhibited antitumor efficacy in patients with urothelial cancer. Therefore, we aimed to evaluate the efficacy, immune response, and safety of S-588410 in patients with completely resected esophageal squamous cell carcinoma (ESCC). Methods: This phase 3 study involved patients with HLA-A*24:02-positive and lymph node metastasis-positive ESCC who received neoadjuvant therapy followed by curative resection. After randomization, patients were administered S-588410 and placebo (both emulsified with Montanide™ ISA 51VG) subcutaneously. The primary endpoint was relapse-free survival (RFS). The secondary endpoints were overall survival (OS), cytotoxic T-lymphocyte (CTL) induction, and safety. Statistical significance was tested using the one-sided weighted log-rank test with the Fleming–Harrington class of weights. Results: A total of 276 patients were randomized (N = 138/group). The median RFS was 84.3 and 84.1 weeks in the S-588410 and placebo groups, respectively (P = 0.8156), whereas the median OS was 236.3 weeks and not reached, respectively (P = 0.6533). CTL induction was observed in 132/134 (98.5%) patients who received S-588410 within 12 weeks. Injection site reactions (137/140 patients [97.9%]) were the most frequent treatment-emergent adverse events in the S-588410 group. Prolonged survival was observed in S-588410-treated patients with upper thoracic ESCC, grade 3 injection-site reactions, or high CTL intensity. Conclusions: S-588410 induced immune response and had acceptable safety but failed to reach the primary endpoint. A high CTL induction rate and intensity may be critical for prolonging survival during future CPV development. [ABSTRACT FROM AUTHOR]

Conventional treatments have shown a limited efficacy for pancreatic cancer, and immunotherapy is an emerging option for treatment of this highly fatal malignancy. Neoantigen is critical to improving the efficacy of tumor-specific immunotherapy. The cancer and peripheral blood specimens from an HLA-A0201–positive pancreatic cancer patient were subjected to next-generation sequencing, and bioinformatics analyses were performed to screen high-affinity and highly stable neoepitopes. The activation of cytotoxic T lymphocytes (CTLs) by dendritic cells (DCs) loaded with mutBCL2A111–20 neoepitope targeting a BCL2A1 mutant epitope was investigated, and the cytotoxicity of mutBCL2A111–20 neoepitope–specific CTLs to pancreatic cancer cells was evaluated. The mutBCL2A111–20 neoepitope was found to present a high immunogenicity and induce CTLs activation and proliferation, and these CTLs were cytotoxic to mutBCL2A111–20 neoepitope–loaded T2 cells and pancreatic cancer PANC-1-Neo and A2-BxPC-3-Neo cells that overexpressed mutBCL2A111–20 neoepitopes, appearing to be a targeting neoepitope specificity. In addition, high BCL2A1 expression correlated with a low 5-yr progression-free interval among pancreatic cancer patients. Our findings provide experimental supports to individualized T cell therapy targeting mutBCL2A111–20 neoepitopes, and provide an option of immunotherapy for pancreatic cancer. [ABSTRACT FROM AUTHOR]

Methods: Control group consisted of 50 patients treated with the FOLFIRI combined with Bevacizumab regimen. The experimental group consisted of 60 patients treated with the Sintilimab combined with Bevacizumab regimen. By comparing the expression levels of CD8+ T lymphocytes, TAMs, and CAFs before and after treatment, short-term efficacy after treatment, and adverse drug reactions between the two groups, we comprehensively evaluated the impact of Sintilimab combined with Bevacizumab on patients with MSS/pMMR advanced colorectal cancer who failed first-line treatment. Results: There was a statistically significant difference in the percentage of CD8+ T lymphocytes, TAMs, and CAFs before and after treatment between the two groups (P<0.05);Immunohistochemical scoring of CD8+ T lymphocytes, TAMs, and CAFs showed significant differences between the groups post-treatment (P<0.05). The experimental group demonstrated statistically significant differences in immunohistochemical scoring of CD8+ T lymphocytes, TAMs, and CAFs before and after treatment (P<0.05). There was a statistically significant difference in the therapeutic effect between the two groups of tumors (P<0.05). The experimental group had greater PFS, mPFS, ORR, and DCR than did the control group. There was no statistically significant difference in the occurrence rate of drug-related adverse reactions after treatment between the two groups (P>0.05). The results of the Cox proportional hazards model analysis indicate that age, gender, and group are independent risk factors affecting MSS/pMMR advanced colorectal cancer patients treated with second-line therapy in this study. Patients aged ≤60 years, male patients, and those in the experimental group showed better treatment responses in this study. Conclusion: By administering immune checkpoint inhibitors in combination with bevacizumab to patients with advanced colorectal cancer with MSS/pMMR disease for whom first-line treatment failed, not only did the patients' prognosis improve, but the adverse drug reactions were also safe and controllable. [ABSTRACT FROM AUTHOR]

Somatic cell therapy based on the infusion of donor-derived cytotoxic T lymphocytes (CTL) able to recognize patients' leukemia blasts (LB) is a promising approach to control leukemia relapse after allogeneic HSCT. The success of this approach strongly depends on the ex vivo generation of high-quality donor-derived anti-leukemia CTL in compliance with Good Manufacturing Practices (GMP). We previously described a procedure for generating large numbers of donor-derived anti-leukemia CTL through stimulation of CD8-enriched lymphocytes with dendritic cells (DCs) pulsed with apoptotic LB in the presence of interleukin (IL)-12, IL-7 and IL-15. Here we report that the use of IFN-DC and the addition of IFNα2b during the priming phase significantly improve the generation of an efficient anti-leukemia T cells response in vitro. Using this approach, 20 high-risk pediatric patients given haploidentical HSCT for high-risk acute leukemia were enrolled and 51 batches of advanced therapy medical products (ATMP), anti-leukemia CTL, were produced. Quality controls demonstrated that all batches were sterile, free of mycoplasma and conformed to acceptable endotoxin levels. Genotype analysis confirmed the molecular identity of the ATMP based on the starting biological material used for their production. The majority of ATMP were CD3+/CD8+ cells, with a memory/terminal activated phenotype, including T-central memory populations. ATMP were viable after thawing, and most ATMP batches displayed efficient capacity to lyse patients' LB and to secrete interferon-γ and tumor necrosis factor-α. These results demonstrated that our protocol is highly reproducible and allows the generation of large numbers of immunologically safe and functional anti-leukemia CTL with a high level of standardization. [ABSTRACT FROM AUTHOR]

Abstract Background Lytic Epstein–Barr virus (EBV) infection plays a major role in the pathogenesis of nasopharyngeal carcinoma (NPC). For patients with recurrent or metastatic NPC and resistant to conventional therapies, adoptive cell therapy using EBV-specific cytotoxic T cells (EBV-CTLs) is a promising option. However, the long production period (around 3 to 4 weeks) and low EBV-CTL purity (approximately 40% of total CD8 T cells) in the cell product limits the application of EBV-CTLs in clinics. Thus, this study aimed to establish a protocol for the rapid production of EBV-CTLs. Methods By culturing peripheral blood mononuclear cells (PBMCs) from EBV-seropositive donors with EBV-specific peptides and interleukin (IL)-2, IL-15, and interferon α (IFN-α) for 9 days, we identified that IL-15 can enhance IL-2-mediated CTL activation and significantly increase the yield of CTLs. Results When IFN-α was used in IL-2/IL-15-mediated CTL production from days 0 to 6, the productivity of EBV-CTLs and EBV-specific cytotoxicity significantly were reinforced relative to EBV-CTLs from IL-2/IL-15 treatment. Additionally, IFN-α-induced production improvement of virus-specific CTLs was not only the case for EBV-CTLs but also for cytomegalovirus-specific CTLs. Conclusion We established a novel protocol to rapidly expand highly pure EBV-CTLs from PBMCs, which can produce EBV-CTLs in 9 days and does not require feeder cells during cultivation.

Abstract Objectives The aim of this study was to evaluate the characteristics of immunocyte associated with bloodstream infection (BSI) caused by Klebsiella pneumoniae (Kpn). Methods Patients with BSI-Kpn were included from 2015 to 2022 in our hospital. Immunocyte subpopulations of enrolled BSI-Kpn patients were tested on the same day of blood culture using multicolor flow cytometry analysis. Antibiotic susceptibility test was determined by agar dilution or broth dilution method. All included isolates were subjected to whole genome sequencing and comparative genomics analysis. Clinical and genetic data were integrated to investigate the risk factors associated with clinical outcome. Results There were 173 patients with non-duplicate BSI-Kpn, including 81 carbapenem-resistant Kpn (CRKP), 30 extended-spectrum β-lactamases producing Kpn (ESBL-Kpn), 62 none CRKP or ESBL-Kpn (S-Kpn). Among 68 ST11-CRKP isolates, ST11-O2v1:KL64 was the most common serotypes cluster (77.9%, 53/68), followed by ST11-OL101: KL47 (13.2%, 9/68). Compared with CSKP group, subpopulations of immunocyte in patients with CRKP were significantly lower (P

Immune thrombocytopenia (ITP) is an autoimmune disease characterized by a low platelet (PLT) count and a high risk of bleeding, the clinical treatment for which still needs to be upgraded. Based on the critical role of human leukocyte antigen class II heterodimer β5 (HLA-DRB5) in immune system, we herein investigated its effect on ITP. ITP murine models were established by the injection of guinea pig anti-mouse platelet serum (GP-APS), and the PLT of mouse peripheral blood was counted during the modeling. Quantitative real-time reverse transcription polymerase chain reaction, western blot and immunofluorescence assay was performed to quantify expressions of HLA-DRB5, major histocompatibility complex II (MHC-II) and co-stimulatory molecules (CD80, CD86). Flow cytometry was conducted to analyze the percentage of CD8+ T cells. As a result, the PLT count was decreased in mouse peripheral blood. Expressions of HLA-DRB5, MHC-II and co-stimulatory molecules, as well as the percentage of CD8+ T cells were elevated in peripheral blood of ITP mice. HLA-DRB5 knockdown mitigated ITP by increasing peripheral PLT level, downregulating expressions of MHC-II and co-stimulatory molecules and inactivating CD8+ T cells. Collectively, the downregulation of HLA-DRB5 restores the peripheral PLT count in ITP mice by reducing MHC-II-mediated antigen presentation of macrophages to inhibit the activation of CD8+ T cells.

Background/Aim: Clear cell carcinoma is a prevalent histological type of ovarian cancer in East Asia, particularly in Japan, known for its resistance to chemotherapeutic agents and poor prognosis. ARID1A gene mutations, commonly found in ovarian clear cell carcinoma (OCCC), contribute to its pathogenesis. Recent data revealed that the ARID1A mutation is related to better outcomes of cancer immunotherapy. Thus, this study aimed to investigate the immunotherapy treatment susceptibility of OCCC bearing ARID1A mutations. Materials and Methods: Expression of ARID1A was analyzed using western blotting in ovarian cancer cell lines. OCCC cell lines JHOC-9 and RMG-V were engineered to overexpress NY-ESO-1, HLAA* 02:01, and ARID1A. Sensitivity to chemotherapy and T cell receptor-transduced T (TCR-T) cells specific for NYESO- 1 was assessed in ARID1A-restored cells compared to ARID1A-deficient wild-type cells. Results: JHOC-9 cells and RMG-V cells showed no expression of ARID1A protein. Overexpression of ARID1A in JHOC-9 and RMG-V cells did not impact sensitivity to gemcitabine. While ARID1A overexpression decreased sensitivity to cisplatin in RMG-V cells, it had no such effect in JHOC-9 cells. ARID1A overexpression reduced the reactivity of NY-ESO-1-specific TCR-T cells, as observed by the IFNγ ESLIPOT assay. Conclusion: Cancer immunotherapy is an effective approach to target ARID1A-deficient clear cell carcinoma of the ovary. [ABSTRACT FROM AUTHOR]

Objectives: The aim of this study was to evaluate the characteristics of immunocyte associated with bloodstream infection (BSI) caused by Klebsiella pneumoniae (Kpn). Methods: Patients with BSI-Kpn were included from 2015 to 2022 in our hospital. Immunocyte subpopulations of enrolled BSI-Kpn patients were tested on the same day of blood culture using multicolor flow cytometry analysis. Antibiotic susceptibility test was determined by agar dilution or broth dilution method. All included isolates were subjected to whole genome sequencing and comparative genomics analysis. Clinical and genetic data were integrated to investigate the risk factors associated with clinical outcome. Results: There were 173 patients with non-duplicate BSI-Kpn, including 81 carbapenem-resistant Kpn (CRKP), 30 extended-spectrum β-lactamases producing Kpn (ESBL-Kpn), 62 none CRKP or ESBL-Kpn (S-Kpn). Among 68 ST11-CRKP isolates, ST11-O2v1:KL64 was the most common serotypes cluster (77.9%, 53/68), followed by ST11-OL101: KL47 (13.2%, 9/68). Compared with CSKP group, subpopulations of immunocyte in patients with CRKP were significantly lower (P < 0.01). In patients with ST11-O2v1:KL64 BSI-Kpn, the level of cytotoxic T lymphocytes (CD3 + CD8 +) is the highest, while the B lymphocytes (CD3-CD19 +) was the least. In addition, the level of immunocyte in patients with Kpn co-harbored clpV-ybtQ-qacE were lower than that in patients with Kpn harbored one of clpV, ybtQ or qacE and without these three genes. Furthermore, co-existence of clpV-ybtQ-qacE was independently associated with a higher risk for 30-day mortality. Conclusions: The results demonstrate that patients with BSI-CRKP, especially for ST11-O2v1:KL64, exhibit lower leukomonocyte counts. In addition, BSI-Kpn co-harbored clpV-ybtQ-qacE is correlated to higher 30-day mortality. [ABSTRACT FROM AUTHOR]

Chronic infection with the hepatitis B virus (HBV) induces progressive hepatic impairment. Achieving complete eradication of the virus remains a formidable challenge. Cytotoxic T lymphocytes, specific to viral antigens, either exhibit a numerical deficiency or succumb to an exhausted state in individuals chronically afflicted with HBV. The comprehension of the genesis and dissemination of stem cell memory T cells (TSCMs) targeting HBV remains inadequately elucidated. We identified TSCMs in subjects with chronic HBV infection and scrutinized their efficacy in a murine model with human hepatocyte transplants, specifically the TK‐NOG mice. TSCMs were discerned in all subjects under examination. Introduction of TSCMs into the HBV mouse model precipitated a severe necro‐inflammatory response, resulting in the elimination of human hepatocytes. TSCMs may constitute a valuable tool in the pursuit of a remedial therapy for HBV infection. [ABSTRACT FROM AUTHOR]

Cytotoxic T lymphocyte (CTL) motility is an important feature of effective CTL responses and is impaired when CTLs become exhausted, e.g. during chronic retroviral infections. A prominent T cell exhaustion marker is programmed cell death protein 1 (PD-1) and antibodies against the interaction of PD-1 and PDligand 1 (PD-L1) are known to improve CTL functions. However, antibody blockade affects all PD-1/PD-L1-expressing cell types, thus, the observed effects cannot be attributed selectively to CTLs. To overcome this problem, we performed CRISPR/ Cas9 based knockout of the PD-1 coding gene PDCD1 in naïve Friend Retrovirus (FV)-specific CTLs. We transferred 1,000 of these cells into mice where they proliferated upon FV-infection. Using intravital two-photon microscopy we visualized CTL motility in the bone marrow and evaluated cytotoxic molecule expression by flow cytometry. Knockout of PDCD1 improved the CTL motility at 14 days post infection and enhanced the expression of cytotoxicity markers. Our data show the potential of genetic tuning of naive antiviral CTLs and might be relevant for future designs of improved T cell-mediated therapies. [ABSTRACT FROM AUTHOR]

Diospyros peregrina is a dioecious plant which is native to India. It belongs to the family of Ebenaceae and is extensively used to treat various ailments, such as leucorrhoea and other uterine-related problems. Though few studies have been on D. peregrina for their anti-tumour response, little is known. Therefore, this intrigued us to understand its immunomodulator capabilities on various types of cancer extensively. Our primary focus is on NSCLC (Non-Small Cell Lung Cancer), which is ranked as the second largest form of cancer in the world, and the treatments demand non-invasive agents to target NSCLC effectively. In an objective to generate an efficient Lung Cancer Associated Antigen (LCA) specific anti-tumour immune response, LCA was presented using dendritic cells (DCs) in the presence of D. peregrina fruit preparation (DFP). Moreover, we also investigated DFP's role in the differentiation of T-helper (TH) cells. Therefore, this study aimed at better LCA presentation mediated by DFP by activating the LCA pulsed DCs and T helper cell differentiation for better immune response. DCs were pulsed with LCA for tumour antigen presentation in vitro, with and without DFP. Differentially pulsed DCs were irradiated to co-culture with autologous and allogeneic lymphocytes. Extracellular supernatants were collected for the estimation of cytokine levels by ELISA. LDH release assay was performed to test Cytotoxic T lymphocytes (CTLs) mediated lung tumour cell cytotoxicity. Thus, DFP may be a potential vaccine to generate anti-LCA immune responses to restrict NSCLC. A graphical representation of the cascade of events pertaining to the augmentation of the process of cancer antigen presentation, TH cell differentiation and subsequent tumour cell cytotoxicity in presence or absence of DFP [ABSTRACT FROM AUTHOR]

Abstract In situ cancer vaccination is an attractive strategy that stimulates protective antitumor immunity. Cytotoxic T lymphocytes (CTLs) are major mediators of the adaptive immune defenses, with critical roles in antitumor immune response and establishing immune memory, and are consequently extremely important for in situ vaccines to generate systemic and lasting antitumor efficacy. However, the dense extracellular matrix and hypoxia in solid tumors severely impede the infiltration and function of CTLs, ultimately compromising the efficacy of in situ cancer vaccines. To address this issue, a robust in situ cancer vaccine, Au@MnO2 nanoparticles (AMOPs), based on a gold nanoparticle core coated with a manganese dioxide shell is developed. The AMOPs modulated the unfavorable tumor microenvironment (TME) to restore CTLs infiltration and function and efficiently induced immunogenic cell death. The Mn2+‐mediated stimulator of the interferon genes pathway can be activated to further augment the therapeutic efficacy of the AMOPs. Thus, the AMOPs vaccine successfully elicited long‐lasting antitumor immunity to considerably inhibit primary, recurrent, and metastatic tumors. This study not only highlights the importance of revitalizing CTLs efficacy against solid tumors but also makes progress toward overcoming TME barriers for sustained antitumor immunity.

Abstract Cytotoxic T lymphocytes (CTLs) play critical antitumor roles, encompassing diverse subsets including CD4+, NK, and γδ T cells beyond conventional CD8+ CTLs. However, definitive CTLs biomarkers remain elusive, as cytotoxicity-molecule expression does not necessarily confer cytotoxic capacity. CTLs differentiation involves transcriptional regulation by factors such as T-bet and Blimp-1, although epigenetic regulation of CTLs is less clear. CTLs promote tumor killing through cytotoxic granules and death receptor pathways, but may also stimulate tumorigenesis in some contexts. Given that CTLs cytotoxicity varies across tumors, enhancing this function is critical. This review summarizes current knowledge on CTLs subsets, biomarkers, differentiation mechanisms, cancer-related functions, and strategies for improving cytotoxicity. Key outstanding questions include refining the CTLs definition, characterizing subtype diversity, elucidating differentiation and senescence pathways, delineating CTL-microbe relationships, and enabling multi-omics profiling. A more comprehensive understanding of CTLs biology will facilitate optimization of their immunotherapy applications. Overall, this review synthesizes the heterogeneity, regulation, functional roles, and enhancement strategies of CTLs in antitumor immunity, highlighting gaps in our knowledge of subtype diversity, definitive biomarkers, epigenetic control, microbial interactions, and multi-omics characterization. Addressing these questions will refine our understanding of CTLs immunology to better leverage cytotoxic functions against cancer.

Abstract Background Esophageal cancer (EC) is a global canker notorious for causing high mortality due to its relentless incidence rate, convoluted with unyielding recurrence and metastasis. However, these intricacies of EC are associated with an immoderate expression of NY-ESO-1 antigen, presenting a lifeline for adoptive T cell therapy. We hypothesized that naturally isolated higher-affinity T cell receptors (TCRs) that bind to NY-ESO-1 would allow T lymphocytes to target EC with a pronounced antitumor response efficacy. Also, targeting TRPV2, which is associated with tumorigenesis in EC, creates an avenue for dual-targeted therapy. We exploited the dual-targeting antitumor efficacy against EC. Methods We isolated antigen-specific TCRs (asTCRs) from a naive library constructed with TCRs obtained from enriched cytotoxic T lymphocytes. The robustness of our asTCRs and their TCR-T cell derivatives, Tranilast (TRPV2 inhibitor), and their bivalent treatment were evaluated with prospective cross-reactive human-peptide variants and tumor cells. Results Our study demonstrated that our naive unenhanced asTCRs and their TCR-Ts perpetuated their cognate HLA-A*02:01/NY-ESO-1(157–165) specificity, killing varying EC cells with higher cytotoxicity compared to the known affinity-enhanced TCR (TCRe) and its wild-type (TCR0) which targets the same NY-ESO-1 antigen. Furthermore, the TCR-Ts and Tranilast bivalent treatment showed superior EC killing compared to any of their monovalent treatments of either TCR-T or Tranilast. Conclusion Our findings suggest that dual-targeted immunotherapy may have a superior antitumor effect. Our study presents a technique to evolve novel, robust, timely therapeutic strategies and interventions for EC and other malignancies.

Cytotoxic T lymphocytes (CTLs) play critical antitumor roles, encompassing diverse subsets including CD4+, NK, and γδ T cells beyond conventional CD8+ CTLs. However, definitive CTLs biomarkers remain elusive, as cytotoxicity-molecule expression does not necessarily confer cytotoxic capacity. CTLs differentiation involves transcriptional regulation by factors such as T-bet and Blimp-1, although epigenetic regulation of CTLs is less clear. CTLs promote tumor killing through cytotoxic granules and death receptor pathways, but may also stimulate tumorigenesis in some contexts. Given that CTLs cytotoxicity varies across tumors, enhancing this function is critical. This review summarizes current knowledge on CTLs subsets, biomarkers, differentiation mechanisms, cancer-related functions, and strategies for improving cytotoxicity. Key outstanding questions include refining the CTLs definition, characterizing subtype diversity, elucidating differentiation and senescence pathways, delineating CTL-microbe relationships, and enabling multi-omics profiling. A more comprehensive understanding of CTLs biology will facilitate optimization of their immunotherapy applications. Overall, this review synthesizes the heterogeneity, regulation, functional roles, and enhancement strategies of CTLs in antitumor immunity, highlighting gaps in our knowledge of subtype diversity, definitive biomarkers, epigenetic control, microbial interactions, and multi-omics characterization. Addressing these questions will refine our understanding of CTLs immunology to better leverage cytotoxic functions against cancer. [ABSTRACT FROM AUTHOR]

Background: Esophageal cancer (EC) is a global canker notorious for causing high mortality due to its relentless incidence rate, convoluted with unyielding recurrence and metastasis. However, these intricacies of EC are associated with an immoderate expression of NY-ESO-1 antigen, presenting a lifeline for adoptive T cell therapy. We hypothesized that naturally isolated higher-affinity T cell receptors (TCRs) that bind to NY-ESO-1 would allow T lymphocytes to target EC with a pronounced antitumor response efficacy. Also, targeting TRPV2, which is associated with tumorigenesis in EC, creates an avenue for dual-targeted therapy. We exploited the dual-targeting antitumor efficacy against EC. Methods: We isolated antigen-specific TCRs (asTCRs) from a naive library constructed with TCRs obtained from enriched cytotoxic T lymphocytes. The robustness of our asTCRs and their TCR-T cell derivatives, Tranilast (TRPV2 inhibitor), and their bivalent treatment were evaluated with prospective cross-reactive human-peptide variants and tumor cells. Results: Our study demonstrated that our naive unenhanced asTCRs and their TCR-Ts perpetuated their cognate HLA-A*02:01/NY-ESO-1(157–165) specificity, killing varying EC cells with higher cytotoxicity compared to the known affinity-enhanced TCR (TCRe) and its wild-type (TCR0) which targets the same NY-ESO-1 antigen. Furthermore, the TCR-Ts and Tranilast bivalent treatment showed superior EC killing compared to any of their monovalent treatments of either TCR-T or Tranilast. Conclusion: Our findings suggest that dual-targeted immunotherapy may have a superior antitumor effect. Our study presents a technique to evolve novel, robust, timely therapeutic strategies and interventions for EC and other malignancies. [ABSTRACT FROM AUTHOR]

Background: Vitiligo is an acquired disease characterized by progressive skin depigmentation resulting from an autoimmune response targeting epidermal melanocytes. Few studies have shown specific cytotoxic T-lymphocyte response against Melan -- A which is the most specific marker of melanocytes. There are also other studies which have confirmed the presence of residual melanocytes even in the vitiliginous skin. Objectives: To evaluate the melanocyte content in lesional, perilesional [within 2 cm] and distant normal skin of non- segmental vitiligo patients. Materials and Methods: A total of 40 cases of non-segmental were enrolled in the study based on the prevelance of vitiligo in India. After taking informed consent, skin biopsy from the lesional, perilesional and normal skin was sent to pathology for immunohistochemical analysis of melanocyte content in the above zones using Melan -- A marker. Results: All 40 lesional sites were negative for Melan-A staining. The Melan-A positive melanocytes were reduced in immediate perilesional skin compared to distant normal skin. Paired t-test was applied between perilesional skin and normal skin and the result was statistically significant at p<0.001. Conclusion: Melan-A positive residual melanocytes were not present in the lesional skin in our study as opposed to very few studies which confirmed their presence in the vitiliginous skin even in the long standing cases. But there was significant difference between perilesional and normal skin. [ABSTRACT FROM AUTHOR]

Introduction: Tuberculosis (TB) remains the first cause of death from infection caused by a bacterial pathogen. Chemotherapy does not eradicate Mycobacterium tuberculosis (Mtb) fromhuman lungs, andthe pathogen causes a latent tuberculosis infectionthatcannotbepreventedbythecurrently availableBacilleCalmetteGuerin (BCG) vaccine, which is ineffective inthe prevention of pulmonary TBinadults. HLAE-restricted CD8+ T lymphocytes are essential players in protective immune responses against Mtb. Hence, expanding this population in vivo or ex vivo may be crucial for vaccination or immunotherapy against TB. Methods: The enzymatically inactive Bordetella pertussis adenylate cyclase (CyaA) toxoid is an effective tool for delivering peptide epitopes into the cytosol of antigen-presenting cells (APC) for presentation and stimulation of specific CD8+ T-cell responses. In this study, we have investigated the capacity of the CyaA toxoid to deliver Mtb epitopes known to bind HLA-E for the expansion of human CD8+ T cells in vitro. Results: Our results show that the CyaA-toxoid containing five HLA-E-restricted Mtb epitopes causes significant expansion of HLA-E-restricted antigen-specific CD8+ T cells, which produce IFN-g and exert significant cytotoxic activity towards peptide-pulsed macrophages. Discussion: HLA-E represents a promising platform for the development of new vaccines; our study indicates that the CyaA construct represents a suitable delivery system of the HLA-E-binding Mtb epitopes for ex vivo and in vitro expansion of HLA-E-restricted CD8+ T cells inducing a predominant Tc1 cytokine profile with a significant increase of IFN-γ production, for prophylactic and immunotherapeutic applications against Mtb. [ABSTRACT FROM AUTHOR]

Eribulin inhibits microtubule polymerization and improves the overall survival of patients with recurrent metastatic breast cancer. A subgroup analysis revealed a low neutrophil to lymphocyte ratio (NLR) (<3) to be a prognostic factor of eribulin treatment. We thus hypothesized that eribulin might be related to the immune response for breast cancer cells and we analyzed the effects of eribulin on the immune system. Immunohistochemical staining revealed that human leukocyte antigen (HLA) class I expression was increased in clinical samples after eribulin treatment. In vitro assays revealed that eribulin treatment increased HLA class I expression in breast cancer line cells. RNA‐sequencing demonstrated that eribulin treatment increased the expression of the NOD‐like family CARD domain‐containing 5 (NLRC5), a master regulator of HLA class I expression. Eribulin treatment increased the NY‐ESO‐1‐specific T‐cell receptor (TCR) transduced T (TCR‐T) cell response for New York oesophageal squamous cell carcinoma 1 (NY‐ESO‐1) overexpressed breast cancer cells. The eribulin and TCR‐T combined therapy model revealed that eribulin and immunotherapy using TCR‐T cells has a synergistic effect. In summary, eribulin increases the expression of HLA class 1 via HLA class 1 transactivatior NLRC5 and eribulin combination with immunotherapy can be effective for the treatment of breast cancer. [ABSTRACT FROM AUTHOR]

Hepatocellular carcinoma (HCC) is a highly fatal malignancy with limited therapeutic options and high recurrence rates. Recently, immunotherapeutic agents such as immune checkpoint inhibitors (ICIs) have emerged as a new paradigm shift in oncology. ICIs, such as programmed cell death protein 1 (PD-1) inhibitors, have provided a new source of hope for patients with advanced HCC. Yet, the eligibility criteria of HCC patients for ICIs are still a missing piece in the puzzle. Circular RNAs (circRNAs) have recently emerged as a new class of non-coding RNAs that play a fundamental role in cancer pathogenesis. Structurally, circRNAs are resistant to exonucleolytic degradation and have a longer half-life than their linear counterparts. Functionally, circRNAs possess the capability to influence various facets of the tumor microenvironment, especially at the HCC tumor–immune synapse. Notably, circRNAs have been observed to control the expression of immune checkpoint molecules within tumor cells, potentially impeding the therapeutic effectiveness of ICIs. Therefore, this renders them potential cancer-immune biomarkers for diagnosis, prognosis, and therapeutic regimen determinants. In this review, the authors shed light on the structure and functional roles of circRNAs and, most importantly, highlight the promising roles of circRNAs in HCC immunomodulation and their potential as promising biomarkers and immunotherapeutic regimen determinants. [ABSTRACT FROM AUTHOR]

Objective: The aim of this study was to explore the safety and efficacy of multiple peptide-pulsed autologous dendritic cells (DCs) combined with cytotoxic T lymphocytes (CTLs) in patients with cancer. Methods: Five patients diagnosed with cancer between November 2020 and June 2021 were enrolled and received DC-CTLs therapy. Peripheral blood was collected and antigenic peptides were analyzed. The phenotype and function of DC-CTLs and the immune status of patients were detected using flow cytometry or IFN-g ELISPOT analysis. Results: DCs acquired a mature phenotype and expressed high levels of CD80, CD86, CD83, and HLA-DR after co-culture with peptides, and the DC-CTLs also exhibited high levels of IFN-g. Peripheral blood mononuclear cells from posttreatment patients showed a stronger immune response to peptides than those prior to treatment. Importantly, four of five patients maintained a favorable immune status, of which one patient's disease-free survival lasted up to 28.2 months. No severe treatment-related adverse events were observed. Conclusion: Our results show that multiple peptide-pulsed DCs combined with CTLs therapy has manageable safety and promising efficacy for cancer patients, which might provide a precise immunotherapeutic strategy for cancer. [ABSTRACT FROM AUTHOR]

Mycobacterium bovis bacilli Calmette-Guerin (BCG) is a licensed vaccine against tuberculosis. It requires attenuated live bacteria to be effective, possibly because actively secreted proteins play a critical role in inducing anti-tuberculosis immunity. BCG also functions as an effective adjuvant. Moreover, the effects of BCG components as adjuvants are not important as those of attenuated live BCG, which is used in cancer immunotherapy. However, the BCG secreted proteins have not been paid attention in anticancer immunity. To understand mycobacterial secreted proteins' function, we investigate immune responses to BCG culture filtrate proteins (CFP). Here, CFP strongly induce both antigenspecific CD4+ T cells and specific CD8+ T cells, which may be functional cytotoxic T lymphocytes (CTLs). In this study, we clearly demonstrate that CFP acts as an adjuvant for CTL induction against specific co-administered proteins and propose CFP as a new protein adjuvant. The CTL response shows potent anticancer effects in mice. These findings could provide insight into the contribution of mycobacterial secreted proteins in both anticancer and antimycobacterial immunity. [ABSTRACT FROM AUTHOR]

Cytotoxic T lymphocyte (CTL) motility is an important feature of effective CTL responses and is impaired when CTLs become exhausted, e.g. during chronic retroviral infections. A prominent T cell exhaustion marker is programmed cell death protein 1 (PD-1) and antibodies against the interaction of PD-1 and PD-ligand 1 (PD-L1) are known to improve CTL functions. However, antibody blockade affects all PD-1/PD-L1-expressing cell types, thus, the observed effects cannot be attributed selectively to CTLs. To overcome this problem, we performed CRISPR/Cas9 based knockout of the PD-1 coding gene PDCD1 in naïve Friend Retrovirus (FV)-specific CTLs. We transferred 1,000 of these cells into mice where they proliferated upon FV-infection. Using intravital two-photon microscopy we visualized CTL motility in the bone marrow and evaluated cytotoxic molecule expression by flow cytometry. Knockout of PDCD1 improved the CTL motility at 14 days post infection and enhanced the expression of cytotoxicity markers. Our data show the potential of genetic tuning of naive antiviral CTLs and might be relevant for future designs of improved T cell-mediated therapies.

Neoadjuvant chemotherapy (NACT) was developed with the aims of shrinking tumors or stopping cancer cells from spreading before surgery. Unfortunately, not all breast cancer patients will benefit from NACT, and thus, patients must weigh the risks and benefits of treatment prior to the initiation of therapy. Currently, the data for predicting the efficacy of NACT is limited. Molecular testing, such as Oncotype DX, MammaPrint, and Curebest 95GC, have been developed to assist which breast cancer patients will benefit from the treatment. Patients with an increased level of Human Leukocyte Antigen-DR isotype, tumor-infiltrating lymphocytes, Fizzy-related protein homolog, and a decreased level of tumor-associated macrophages appear to benefit most from NACT.

The 2019 coronavirus disease (COVID-19) pandemic has had an unprecedented impact on health and economies around the world. Direct myocardial injury and cytokine storm, leading to destabilization of preexisting plaques and accelerated formation of new plaques, are two mechanisms that trigger the acute coronary syndrome in COVID-19. There is insufficient data on the immune status of patients with acute coronary syndrome who have undergone COVID-19. The aim of the study was to study T and B cell, humoral immunity depending on the number of cytotoxic T lymphocytes (CD8+) in patients with acute coronary syndrome who underwent COVID-19. Materials and methods of research: 65 men with unstable angina pectoris and acute myocardial infarction (acute coronary syndrome) from 40 to 65 years old, who had previously had COVID-19, were examined. A study of peripheral blood was carried out: complete blood count (Medonic device, Sweden), general and specific IgM, IgG, IgA, compliment fragments (Vector Best, Russia). Subpopulations of T and B lymphocytes were determined by flow cytometry. In persons with acute coronary syndrome who underwent COVID-19 with predominantly normal and elevated levels of cytotoxic T cells, a more severe course of the disease was observed: patients with acute myocardial infarction prevailed, they had longer mortality, longer treatment duration, and stent thrombosis was more common. In patients with elevated cytotoxic T cells, there was a maximum increase in erythrocytes, hemoglobin, hematocrit, lymphocytes of both the total number and subpopulations – T helpers, T-NK lymphocytes, NK lymphocytes, T lymphocytes of early and late activation, B1 and B2 lymphocytes, index of NBT-induced test. In patients with normal levels of NK cells, compared with other groups, there was an increase in spontaneous NBT activity and index, a significant decrease in C3a and C5a complement fragments. Prevalence of stent thrombosis and mortality in the group of patients with normal levels of cytotoxic T cells may indicate torpidity of the immune system in these patients with poor outcomes.

IntroductionTuberculosis (TB) remains the first cause of death from infection caused by a bacterial pathogen. Chemotherapy does not eradicate Mycobacterium tuberculosis (Mtb) from human lungs, and the pathogen causes a latent tuberculosis infection that cannot be prevented by the currently available Bacille Calmette Guerin (BCG) vaccine, which is ineffective in the prevention of pulmonary TB in adults. HLA-E-restricted CD8+ T lymphocytes are essential players in protective immune responses against Mtb. Hence, expanding this population in vivo or ex vivo may be crucial for vaccination or immunotherapy against TB.MethodsThe enzymatically inactive Bordetella pertussis adenylate cyclase (CyaA) toxoid is an effective tool for delivering peptide epitopes into the cytosol of antigen-presenting cells (APC) for presentation and stimulation of specific CD8+ T-cell responses. In this study, we have investigated the capacity of the CyaA toxoid to deliver Mtb epitopes known to bind HLA-E for the expansion of human CD8+ T cells in vitro.ResultsOur results show that the CyaA-toxoid containing five HLA-E-restricted Mtb epitopes causes significant expansion of HLA-E-restricted antigen-specific CD8+ T cells, which produce IFN-γ and exert significant cytotoxic activity towards peptide-pulsed macrophages.DiscussionHLA-E represents a promising platform for the development of new vaccines; our study indicates that the CyaA construct represents a suitable delivery system of the HLA-E-binding Mtb epitopes for ex vivo and in vitro expansion of HLA-E-restricted CD8+ T cells inducing a predominant Tc1 cytokine profile with a significant increase of IFN-γ production, for prophylactic and immunotherapeutic applications against Mtb.

ABSTRACTCancer-testis antigen CT23 is a class of tumor-associated antigens (TAA) characterized by restricted expression in male germ cells and a variety of tumor tissues. Numerous studies have shown that CT23 is closely related to tumor cell viability, proliferation, metastasis and invasion. CT23 is immunogenic and can cause specific immune response in tumor patients. Therefore, it is considered to be one of the best target antigens for designing therapeutic tumor vaccines and T-cell-mediated tumor immunotherapy. In this study, we initially obtained seven HLA-A*0201-restricted CT23 epitope candidate peptides through the T cell epitope prediction program. Subsequently, a T2 cell binding assay revealed the potential binding of all candidate peptides with HLA-A2 molecules. Notably, peptide P7 (ALLVLCYSI) exhibited the highest affinity, as evidenced by a fluorescence index (FI) of 2.19. Dendritic cells (DCs) loaded with CT23 candidate peptide can stimulate CD8+T cell activation and proliferation, and compared with other candidate peptides, candidate peptide P7 is superior. The cytotoxic T lymphocytes (CTLs) stimulated by the peptide P7 had killing effect on tumor cells (HLA-A*0201+, CT23+), but no killing effect on tumor cells (HLA-A*0201−, CT23+). The CTLs induced by the peptide P7 also had a specific killing effect on T2 cells bearing the peptide P7. In summary, our findings suggest that the CT23 peptide P7 (ALLVLCYSI) can induce immune responses and holds potential for tumor-specific CTL therapy.

Neoantigen-reactive cytotoxic T lymphocytes play a vital role in precise cancer cell elimination. In this study, we demonstrate the effectiveness of personalized neoantigen-based T cell therapy in inducing tumor regression in two patients suffering from heavily-burdened metastatic ovarian cancer. Our approach involved the development of a robust pipeline for ex vivo expansion of neoantigen-reactive T lymphocytes. Neoantigen peptides were designed and synthesized based on the somatic mutations of the tumors and their predicted HLA binding affinities. These peptides were then presented to T lymphocytes through co-culture with neoantigen-loaded dendritic cells for ex vivo expansion. Subsequent to cell therapy, both patients exhibited significant reductions in tumor marker levels and experienced substantial tumor regression. One patient achieved repeated cancer regression through infusions of T cell products generated from newly identified neoantigens. Transcriptomic analyses revealed a remarkable increase in neoantigen-reactive cytotoxic lymphocytes in the peripheral blood of the patients following cell therapy. These cytotoxic T lymphocytes expressed polyclonal T cell receptors (TCR) against neoantigens, along with abundant cytotoxic proteins and pro-inflammatory cytokines. The efficacy of neoantigen targeting was significantly associated with the immunogenicity and TCR polyclonality. Notably, the neoantigen-specific TCR clonotypes persisted in the peripheral blood after cell therapy. Our findings indicate that personalized neoantigen-based T cell therapy triggers cytotoxic lymphocytes expressing polyclonal TCR against ovarian cancer, suggesting its promising potential in cancer immunotherapy.

Prophylactic tumor vaccines hold great promise against tumor occurrence. However, their clinical efficacy remains low due to inadequate activation of strong-sustainable immunity. Herein, a biomembrane hydrogel was designed as a powerful single-shot prophylactic tumor vaccine. Mannose-decorated hybrid biomembrane (MHCM) modified with oxidized sodium alginate (OSA) was designed as a gelator (O-MHCM), where the hybrid biomembrane (HCM) is a hybridization of bacterial outer membrane vesicles (OMV) and tumor cell membranes (TCM). The O-MHCM enables quick gelation subcutaneously where the cysteine protease inhibitor E64 is encapsulated in hydrogel micropores. After a single vaccination of E64@O-MHCM hydrogel, MHCM and E64 are released sustainably due to OSA moiety degradation. The MHCM enables active targeting to dendritic cells (DC) and effective DC maturation. Meanwhile, the E64 enables sufficient antigen availability for subsequent cross presentation. Ultimately, strong and sustainable T lymphocyte-mediated immunity was elicited, demonstrating a strong prophylactic effect against breast tumors. This study provides a long-lasting platform to prevent tumor occurrence, opening an innovative avenue for the design of a single-shot prophylactic tumor vaccine. Developing a single-shot prophylactic tumor vaccine to elicit strong-sustainable immunity is of great interest clinically. Here, a prophylactic tumor vaccine was designed using an injectable biomembrane hydrogel for achieving strong-sustainable immunity. The mannose-tailored hybrid biomembrane was modified with oxidized sodium alginate to result in a gelator, which enabled the formation of the hydrogel after subcutaneous injection. Cysteine protease inhibitor E64 was incorporated into the micropores of the hydrogel. The hydrogel induced strong-sustainable immunity through the continuous release of active components. This was facilitated by the mannose moiety, which enabled active targeting, as well as the antigen and adjuvant function of biomembrane, and the E64-enabled suppression of antigen degradation. The biomembrane hydrogel demonstrated powerful prevention of 4T1 breast tumors. This study offers an attractive strategy for designing a single-shot prophylactic tumor vaccine. [Display omitted] [ABSTRACT FROM AUTHOR]

Oral squamous cell carcinomas (OSCCs) are genetically heterogeneous and exhibit diverse stromal and immune microenvironments. Acquired resistance to standard chemo‐, radio‐, and targeted therapies remains a major hurdle in planning effective treatment modalities for OSCC patients. Since Caspase 8 (CASP8) is frequently mutated in OSCCs, we were interested to explore a potential interaction between tumour‐infiltrating lymphocytes (TILs) and CASP8 activation using high‐content image analysis of human tumour (n = 32) sections. Despite the lymphocyte‐rich tumour microenvironment, we observed lower activation of CASP8 (0–10% of tumour area) and its downstream effector CASP3 (0–6%) in tumours than in normal oral epithelium. Conversely, we found apoptosis was high for all the lymphocyte subtypes examined (38–52% of lymphocytes within tumour islands). Tumours with higher Fas ligand (FasL) expression had a significantly higher proportion of cleaved CASP3/8 positive cytotoxic T cells within the tumour islands (p = 0.05), and this was associated with the presence of lymph node metastatic disease [odds ratio: 1.046, 95% confidence interval (1.002–1.091), p = 0.039]. Our finding of extensive activation of the extrinsic pathway of apoptosis in TILs, together with evidence of higher FasL in CASP8 mutated tumours, may be useful in predicting the course of disease in individual patients. © 2023 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of The Pathological Society of Great Britain and Ireland. [ABSTRACT FROM AUTHOR]

Rheumatoid factor (RF) and anti-citrullinated protein antibodies (ACPAs) are the most frequently used rheumatoid arthritis (RA) diagnostic markers, but they are unable to anticipate the patient's evolution or response to treatment. The aim of this study was to identify possible severity biomarkers to predict an upcoming flare-up or remission period. To address this objective, sera and anticoagulated blood samples were collected from healthy controls (HCs; n = 39) and from early RA (n = 10), flare-up (n = 5), and remission (n = 16) patients. We analyzed leukocyte phenotype markers, regulatory T cells, cell proliferation, and cytokine profiles. Flare-up patients showed increased percentages of cluster of differentiation (CD)3+CD4− lymphocytes (p < 0.01) and granulocytes (p < 0.05) but a decreased natural killer (NK)/T lymphocyte ratio (p < 0.05). Analysis of leukocyte markers by principal component analysis (PCA) and receiver operating characteristic (ROC) curves showed that CD45RO+ (p < 0.0001) and CD45RA+ (p < 0.0001) B lymphocyte expression can discriminate between HCs and early RA patients, while CD3+CD4− lymphocyte percentage (p < 0.0424) and CD45RA+ (p < 0.0424), CD62L+ (p < 0.0284), and CD11a+ (p < 0.0185) B lymphocyte expression can differentiate between flare-up and RA remission subjects. Thus, the combined study of these leukocyte surface markers could have potential as disease severity biomarkers for RA, whose fluctuations could be related to the development of the characteristic pro-inflammatory environment. [ABSTRACT FROM AUTHOR]

Anticancer drug‐mediated induction of immunogenic cell death (ICD) blocks metastasis or recurrence in cancer cells by promoting specific immune activity against cancer antigens. However, this strategy has failed to afford adequate treatment efficiency. Overcoming the failure of ICD‐mediated cancer therapy, lipid nanoparticles (LNPs) containing cancer cell surface proteins are synthesized using sonication and extrusion without microfluidics. In addition, these LNPs are decorated with high‐mobility group box 1 protein and calreticulin, indicators of ICD, and named artificial ICD LNPs (AiLNPs). Administration of AiLNPs effectively targets dendritic cells (DCs) and induces DC activation in mice. Moreover, treating CT‐26 tumor‐bearing mice with AiLNPs inhibits tumor growth by inducing CT‐26 antigen‐specific T‐cell immunity. Furthermore, AiLNPs containing Lewis lung carcinoma (LLC1) membrane proteins can prevent metastatic LLC1 tumor growth in the lung via LLC1 antigen‐specific T‐cell activation. Finally, AiLNPs synthesized with human breast cancer membrane proteins activate DC‐mediated antigen‐specific T‐cell immunity, effectively killing tumor cells. Therefore, AiLNPs are expected to be developed as a patient‐specific cancer treatment to prevent cancer recurrence and metastasis. [ABSTRACT FROM AUTHOR]

Lejia Zhang,&ast; Kuangwu Pan,&ast; Siyuan Huang, Xiliu Zhang, Xinyu Zhu, Yi He, Xun Chen, Yuquan Tang, Lingyu Yuan, Dongsheng Yu Hospital of Stomatology, Sun Yat-Sen University, Guangdong Provincial Key Laboratory of Stomatology, Guanghua School of Stomatology, Guangzhou, People’s Republic of China&ast;These authors contributed equally to this workCorrespondence: Dongsheng Yu, Hospital of Stomatology, Sun Yat-Sen University, Guangdong Provincial Key Laboratory of Stomatology, Guanghua School of Stomatology, Guangzhou, People’s Republic of China, Email yudsh@mail.sysu.edu.cnPurpose: Nanomaterial-based photodynamic therapy (PDT) has been commonly used for the treatment of cancerous tumors. Despite significant achievements made in this field, the intrinsic impact of nanomaterials-based PDT on the mechanical properties of oral squamous cell carcinoma (OSCC) cells is not entirely understood. Here, we used atomic force microscopy (AFM) to measure the stiffness of OSCC cells subjected to PDT in co-culture systems to evaluate the T cell-mediated cancer cell-killing effects.Methods: In this study, AFM was used to assess the stiffness of PDT-subjected cells. The phototoxicity of graphdiyne oxide (GDYO) was assessed using confocal laser scanning microscopy (CLSM), measurements of membrane cholesterol levels, and assessments of the F-actin cytoskeleton. A co-culture system was used to evaluate the effects of CD8+ T cells (cytotoxic T lymphocytes), demonstrating how PDT modulates the mechanical properties of cancer cells and activates T cell responses. The antitumor immunotherapeutic effect of GDYO was further evaluated in a murine xenograft model.Results: GDYO increased the mechanical stiffness of tumor cells and augmented T-cell cytotoxicity and inflammatory cytokine secretion (IFN-γ and TNF-α) under laser in vitro. Furthermore, GDYO-based PDT exerted inhibitory effects on OSCC models and elicited antitumor immune responses via specific cytotoxic T cells.Conclusion: These results highlight that GDYO is a promising candidate for OSCC therapy, shifting the mechanical forces of OSCC cells and breaking through the barriers of the immunosuppressive tumor microenvironment. Our study provides a novel perspective on nanomaterial-based antitumor therapies.Graphical Abstract: Keywords: graphdiyne oxide, photodynamic therapy, stiffness, immunotherapy, cytotoxic T lymphocytes, oral squamous cell carcinoma cells

Summary: Background: In individuals with malignancy or HIV-1 infection, antigen-specific cytotoxic T lymphocytes (CTLs) often display an exhausted phenotype with impaired capacity to eliminate the disease. Existing cell-based immunotherapy strategies are often limited by the requirement for adoptive transfer of CTLs. We have developed an immunotherapy technology in which potent CTL responses are generated in vivo by vaccination and redirected to eliminate target cells using a bispecific Redirector of Vaccine-induced Effector Responses (RoVER). Methods: Following Yellow fever (YF) 17D vaccination of 51 healthy volunteers (NCT04083430), single-epitope YF-specific CTL responses were quantified by tetramer staining and multi-parameter flow cytometry. RoVER-mediated redirection of YF-specific CTLs to kill antigen-expressing Raji-Env cells, autologous CD19+ B cells or CD4+ T cells infected in vitro with a full-length HIV-1-eGFP was assessed in cell killing assays. Moreover, secreted IFN-γ, granzyme B, and TNF-α were analyzed by mesoscale multiplex assays. Findings: YF-17D vaccination induced strong epitope-specific CTL responses in the study participants. In cell killing assays, RoVER-mediated redirection of YF-specific CTLs to autologous CD19+ B cells or HIV-1-infected CD4+ cells resulted in 58% and 53% killing at effector to target ratio 1:1, respectively. Interpretation: We have developed an immunotherapy technology in which epitope-specific CTLs induced by vaccination can be redirected to kill antigen-expressing target cells by RoVER linking. The RoVER technology is highly specific and can be adapted to recognize various cell surface antigens. Importantly, this technology obviates the need for adoptive transfer of CTLs. Funding: This work was funded by the Novo Nordisk Foundation (Hallas Møller NNF10OC0054577).

Cytotoxic T lymphocytes (CTLs) are important in controlling some viral infections, and therapies involving the transfer of large numbers of cancer-specific CTLs have been successfully used to treat several types of cancers in humans. While the molecular mechanisms of how CTLs kill their targets are relatively well understood, we still lack a solid quantitative understanding of the kinetics and efficiency by which CTLs kill their targets in vivo. Collagen–fibrin-gel-based assays provide a tissue-like environment for the migration of CTLs, making them an attractive system to study T cell cytotoxicity in in vivo-like conditions. Budhu.et al. systematically varied the number of peptide (SIINFEKL)-pulsed B16 melanoma cells and SIINFEKL-specific CTLs (OT-1) and measured the remaining targets at different times after target and CTL co-inoculation into collagen–fibrin gels. The authors proposed that their data were consistent with a simple model in which tumors grow exponentially and are killed by CTLs at a per capita rate proportional to the CTL density in the gel. By fitting several alternative mathematical models to these data, we found that this simple "exponential-growth-mass-action-killing" model did not precisely describe the data. However, determining the best-fit model proved difficult because the best-performing model was dependent on the specific dataset chosen for the analysis. When considering all data that include biologically realistic CTL concentrations ( E ≤ 10 7 cell / mL ), the model in which tumors grow exponentially and CTLs suppress tumor's growth non-lytically and kill tumors according to the mass–action law (SiGMA model) fit the data with the best quality. A novel power analysis suggested that longer experiments (∼3–4 days) with four measurements of B16 tumor cell concentrations for a range of CTL concentrations would best allow discriminating between alternative models. Taken together, our results suggested that the interactions between tumors and CTLs in collagen–fibrin gels are more complex than a simple exponential-growth-mass–action killing model and provide support for the hypothesis that CTLs' impact on tumors may go beyond direct cytotoxicity. [ABSTRACT FROM AUTHOR]

Evasion from immunity is a major obstacle to the achievement of successful cancer immunotherapy. Hybrids derived from cell–cell fusion are theoretically associated with tumor heterogeneity and progression by conferring novel properties on tumor cells, including drug resistance and metastatic capacity; however, their impact on immune evasion remains unknown. Here, we investigated the potency of tumor–macrophage hybrids in immune evasion. Hybrids were established by co‐culture of a melanoma cell line (A375 cells) and type 2 macrophages. The hybrids showed greater migration ability and greater tumorigenicity than the parental melanoma cells. The hybrids showed heterogeneous sensitivity to New York esophageal squamous cell carcinoma‐1 (NY‐ESO‐1)‐specific T‐cell receptor‐transduced T (TCR‐T) cells and two out of four hybrid clones showed less sensitivity to TCR‐T compared with the parental cells. An in vitro tumor heterogeneity model revealed that the TCR‐T cells preferentially killed the parental cells compared with the hybrids and the survival rate of the hybrids was higher than that of the parental cells, indicating that the hybrids evade killing by TCR‐T cells efficiently. Analysis of a single‐cell RNA sequencing dataset of patients with melanoma revealed that a few macrophages expressed RNA encoding melanoma differentiation antigens including melan A, tyrosinase, and premelanosome protein, which indicated the presence of hybrids in primary melanoma. In addition, the number of potential hybrids was correlated with a poorer response to immune checkpoint blockade. These results provide evidence that melanoma–macrophage fusion has a role in tumor heterogeneity and immune evasion. © 2023 The Pathological Society of Great Britain and Ireland. [ABSTRACT FROM AUTHOR]

Lichen planus (LP) is a cutaneomucosal chronic inflammatory disease characterized by a CD8+ cytotoxic T‐lymphocytes (CTL) infiltrate. In erosive oral LP, we found HPV16‐specific activated CTL in lesions, supporting a pathogenic contribution of HPV16. Here, we investigated whether a similar scenario occurs in other clinical forms of LP and in lichen sclerosus et atrophicus (LSA), another chronic disease also affecting the mucosa and/or the skin. Blood CTL from LP and LSA patients expressed significant higher levels of granzyme B, perforin and CD107a proteins than healthy donors. Expansions of TCRVß3+ CTL, with presence of TCR clonotypes identical to those previously detected in erosive oral LP, were found both in blood and mucosal/skin lesions of LP, and not of LSA patients. These expansions were enriched with HPV16‐specific CD8+ T‐cells as shown by their recognition of the E711‐20 immunodominant epitope. In LSA patients, the peripheral repertoire of CTL was oligoclonal for TCRVß6+ CTL. Finally, although patients with LP and LSA have developed antibodies against HPV16 capsid L1, antibodies against HPV16 E6 were only observed in patients with LP. Overall, our data collectively suggest an involvement of HPV16‐specific CTL in different clinical forms of LP, not only in erosive oral LP, while a different scenario operates in LSA. [ABSTRACT FROM AUTHOR]

Nef is an HIV-encoded accessory protein that enhances pathogenicity by down-regulating major histocompatibility class I (MHC-I) expression to evade killing by cytotoxic T lymphocytes (CTLs). A potent Nef inhibitor that restores MHC-I is needed to promote immune-mediated clearance of HIV-infected cells. We discovered that the plecomacrolide family of natural products restored MHC-I to the surface of Nef-expressing primary cells with variable potency. Concanamycin A (CMA) counteracted Nef at subnanomolar concentrations that did not interfere with lysosomal acidification or degradation and were nontoxic in primary cell cultures. CMA specifically reversed Nef-mediated down-regulation of MHC-I, but not CD4, and cells treated with CMA showed reduced formation of the Nef:MHC-I:AP-1 complex required for MHC-I down-regulation. CMA restored expression of diverse allotypes of MHC-I in Nef-expressing cells and inhibited Nef alleles from divergent clades of HIV and simian immunodeficiency virus, including from primary patient isolates. Lastly, we found that restoration of MHC-I in HIV-infected cells was accompanied by enhanced CTL-mediated clearance of infected cells comparable to genetic deletion of Nef. Thus, we propose CMA as a lead compound for therapeutic inhibition of Nef to enhance immune-mediated clearance of HIV-infected cells.

Background: Chronic lymphocytic leukemia (CLL) is the most common type of leukemia in dogs. It is characterized by the proliferation of neoplastic lymphocytes in the bone marrow, which are morphologically normal (mature), but non-functional. CLL in canines commonly originates in cytotoxic T lymphocytes (TCD8+), and although there is controversy regarding the prognostic value of the immunophenotype, this cell lineage may be associated with a good prognosis. Case description: A 10-year-old, entire female, mixed-breed dog was brought to the University Hospital of the Veterinary Faculty (UdelaR) for consultation because a routine pre-surgical check-up revealed lymphocytic leukocytosis, normocytic anemia, and hyperglobulinemia due to an oligoclonal gammopathy. The ultrasound revealed splenomegaly. PCR performed on blood was negative for Ehrlichia canis. Blood and bone marrow flow cytometry was performed to complement the diagnosis and carry out the immunophenotype, which showed CLL of CD8+ T-cell lineage. The clinical suspicion of CLL was confirmed by myelogram. Chemotherapy treatment based on alkylating agents and glucocorticoids was established. So far, the patient has an overall survival of 13 months with a good response to treatment. Conclusion: The combination of immunophenotyping test, the myelogram, and the hematological and biochemical profile confirmed the presence of T-CLL in our patient. Flow cytometry, increasingly used in veterinary medicine, allowed us to confirm the diagnosis of CLL originating in cytotoxic T lymphocytes in our patient, through the presence of positive staining of primary antibodies specific for the canine species CD45, CD3, CD5 and CD8 and the absence of staining for CD4, CD21 and CD34. [Open Vet J 2022; 12(6.000): 868-876]

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants of concern (VOCs) possess mutations that confer resistance to neutralizing antibodies within the Spike protein and are associated with breakthrough infection and reinfection. By contrast, less is known about the escape from CD8+ T cell-mediated immunity by VOC. Here, we demonstrated that all SARS-CoV-2 VOCs possess the ability to suppress major histocompatibility complex class I (MHC-I) expression. We identified several viral genes that contribute to the suppression of MHC I expression. Notably, MHC-I upregulation was strongly inhibited after SARS-CoV-2 but not influenza virus infection in vivo. While earlier VOCs possess similar capacity as the ancestral strain to suppress MHC-I, the Omicron subvariants exhibited a greater ability to suppress surface MHC-I expression. We identified a common mutation in the E protein of Omicron that further suppressed MHC-I expression. Collectively, our data suggest that in addition to escaping from neutralizing antibodies, the success of Omicron subvariants to cause breakthrough infection and reinfection may in part be due to its optimized evasion from T cell recognition. [ABSTRACT FROM AUTHOR]

Background: With the approval of the HIV-1 capsid inhibitor, lenacapavir, capsid sequencing will be required for managing lenacapavir-experienced individuals with detectable viremia. Successful sequence interpretation will require examining new capsid sequences in the context of previously published sequence data. Methods: We analyzed published HIV-1 group M capsid sequences from 21,012 capsid-inhibitor naïve individuals to characterize amino acid variability at each position and influence of subtype and cytotoxic T lymphocyte (CTL) selection pressure. We determined the distributions of usual mutations, defined as amino acid differences from the group M consensus, with a prevalence ≥ 0.1%. Co-evolving mutations were identified using a phylogenetically-informed Bayesian graphical model method. Results: 162 (70.1%) positions had no usual mutations (45.9%) or only conservative usual mutations with a positive BLOSUM62 score (24.2%). Variability correlated independently with subtype-specific amino acid occurrence (Spearman rho = 0.83; p < 1 × 10−9) and the number of times positions were reported to contain an HLA-associated polymorphism, an indicator of CTL pressure (rho = 0.43; p = 0.0002). Conclusions: Knowing the distribution of usual capsid mutations is essential for sequence quality control. Comparing capsid sequences from lenacapavir-treated and lenacapavir-naïve individuals will enable the identification of additional mutations potentially associated with lenacapavir therapy. [ABSTRACT FROM AUTHOR]

Immune suppressive factors of the tumor microenvironment (TME) undermine viability and exhaust the activities of the intratumoral cytotoxic CD8 + T lymphocytes (CTL) thereby evading anti-tumor immunity and decreasing the benefits of immune therapies. To counteract this suppression and improve the efficacy of therapeutic regimens, it is important to identify and understand the critical regulators within CD8 + T cells that respond to TME stress and tumor-derived factors. Here we investigated the regulation and importance of activating transcription factor-4 (ATF4) in CTL using a novel Atf4ΔCD8 mouse model lacking ATF4 specifically in CD8 + cells. Induction of ATF4 in CD8 + T cells occurred in response to antigenic stimulation and was further increased by exposure to tumor-derived factors and TME conditions. Under these conditions, ATF4 played a critical role in the maintenance of survival and activities of CD8 + T cells. Conversely, selective ablation of ATF4 in CD8 + T cells in mice rendered these Atf4ΔCD8 hosts prone to accelerated growth of implanted tumors. Intratumoral ATF4-deficient CD8 + T cells were under-represented compared to wild-type counterparts and exhibited impaired activation and increased apoptosis. These findings identify ATF4 as an important regulator of viability and activity of CD8 + T cells in the TME and argue for caution in using agents that could undermine these functions of ATF4 for anti-cancer therapies. [ABSTRACT FROM AUTHOR]

Equine Infectious Anemia Virus (EIAV) is an important infection in equids, and its similarity to HIV creates hope for a potential vaccine. We analyze a within-host model of EIAV infection with antibody and cytotoxic T lymphocyte (CTL) responses. In this model, the stability of the biologically relevant endemic equilibrium, characterized by the coexistence of long-term antibody and CTL levels, relies upon a balance between CTL and antibody growth rates, which is needed to ensure persistent CTL levels. We determine the model parameter ranges at which CTL and antibody proliferation rates are simultaneously most influential in leading the system towards coexistence and can be used to derive a mathematical relationship between CTL and antibody production rates to explore the bifurcation curve that leads to coexistence. We employ Latin hypercube sampling and least squares to find the parameter ranges that equally divide the endemic and boundary equilibria. We then examine this relationship numerically via a local sensitivity analysis of the parameters. Our analysis is consistent with previous results showing that an intervention (such as a vaccine) intended to control a persistent viral infection with both immune responses should moderate the antibody response to allow for stimulation of the CTL response. Finally, we show that the CTL production rate can entirely determine the long-term outcome, regardless of the effect of other parameters, and we provide the conditions for this result in terms of the identified ranges for all model parameters. [ABSTRACT FROM AUTHOR]