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Background Exposomes are critical drivers of carcinogenesis. However, how they modulate tumor behavior remains unclear. Extensive clinical data show cigarette smoke to be a key exposome that promotes aggressive tumors, higher rates of metastasis, reduced response to chemoradiotherapy, and suppressed anti-tumor immunity. We sought to determine whether smoke itself can modulate aggressive tumor behavior in head and neck squamous cell carcinoma (HNSCC) through reprogramming of the cellular reductive state. Methods Using established human and murine HNSCC cell lines and syngeneic mouse models, we utilized conventional western blotting, steady state and flux metabolomics, RNA sequencing, quantitative proteomics and flow cytometry to analyze the impact of smoke exposure on HNSCC tumor biology and anti-tumor immunity. Results Cigarette smoke persistently activated Nrf2 target genes essential for maintenance of the cellular reductive state and survival under conditions of increased oxidative stress in HNSCC regardless of human papillomavirus (HPV) association. In contrast to e-cigarette vapor, conventional cigarette smoke mobilizes cellular metabolism toward oxidative stress adaptation, resulting in development of cross-resistance to cisplatin. In parallel, smoke exposure modulates expression of PDL1 and the secretory phenotype of HNSCC cells resulting in an altered tumor immune microenvironment (TIME) in syngeneic mouse models and downregulated expression of antigen presentation and costimulatory genes in myeloid cells. Conclusion The cigarette smoke exposome is a potent activator of the Nrf2 pathway and appears to be the primary trigger for a tripartite phenotype of aggressive HNSCC consisting of: (1) reduced chemotherapy sensitivity, (2) enhanced metastatic potential and (3) suppressed anti-tumor immunity.

Treatment of HIV infection with either antiretroviral (ARV) therapy or neutralizing monoclonal antibodies (NAbs) leads to a reduction in HIV plasma virus. Both ARVs and NAbs prevent new rounds of viral infection, but NAbs may have the additional capacity to accelerate the loss of virus-infected cells through Fc gamma receptor (FcγR)-mediated effector functions, which should affect the kinetics of plasma-virus decline. Here, we formally test the role of effector function in vivo by comparing the rate and timing of plasma-virus clearance in response to a single-dose treatment with either unmodified NAb or those with either reduced or augmented Fc function. When infused into viremic simian HIV (SHIV)-infected rhesus macaques, there was a 21% difference in slope of plasma-virus decline between NAb and NAb with reduced Fc function. NAb engineered to increase FcγRIII binding and improve antibody-dependent cellular cytotoxicity (ADCC) in vitro resulted in arming of effector cells in vivo, yet led to viral-decay kinetics similar to NAbs with reduced Fc function. These studies show that the predominantmechanism of antiviral activity of HIV NAbs is through inhibition of viral entry, but that Fc function can contribute to the overall antiviral activity, making them distinct from standard ARVs.

Over the course of evolution, many proteins have undergone adaptive structural changes to meet the increasing homeostatic regulatory demands of multicellularity. Aminoacyl tRNA synthetases (aaRS), enzymes that catalyze the attachment of each amino acid to its cognate tRNA, are such proteins that have acquired new domains and motifs that enable non-canonical functions. Through these new domains and motifs, aaRS can assemble into large, multi-subunit complexes that enhance the efficiency of many biological functions. Moreover, because the complexity of multi-aminoacyl tRNA synthetase (mARS) complexes increases with the corresponding complexity of higher eukaryotes, a contribution to regulation of homeostatic functions in multicellular organisms is hypothesized. While mARS complexes in lower eukaryotes may enhance efficiency of aminoacylation, little evidence exists to support a similar role in chordates or other higher eukaryotes. Rather, mARS complexes are reported to regulate multiple and variegated cellular processes that include angiogenesis, apoptosis, inflammation, anaphylaxis, and metabolism. Because all such processes are critical components of immune homeostasis, it is important to understand the role of mARS complexes in immune regulation. Here we provide a conceptual analysis of the current understanding of mARS complex dynamics and emerging mARS complex roles in immune regulation, the increased understanding of which should reveal therapeutic targets in immunity and immune-mediated disease.

CIS43 is a potent neutralizing human mAb that targets a highly conserved “junctional” epitope in the Plasmodium falciparum (Pf) circumsporozoite protein (PfCSP). Enhancing the durability of CIS43 in vivo will be important for clinical translation. Here, 2 approaches were used to improve the durability of CIS43 in vivo while maintaining potent neutralization. First, the Fc domain was modified with the LS mutations (CIS43LS) to increase CIS43 binding affinity for the neonatal Fc receptor (FcRn). CIS43LS and CIS43 showed comparable in vivo protective efficacy. CIS43LS had 9- to 13-fold increased binding affinity for human (6.2 nM versus 54.2 nM) and rhesus (25.1 nM versus 325.8 nM) FcRn at endosomal pH 6.0 compared with CIS43. Importantly, the half-life of CIS43LS in rhesus macaques increased from 22 days to 39 days compared with CIS43. The second approach for sustaining antibody levels of CIS43 in vivo is through adeno-associated virus (AAV) expression. Mice administered once with AAV-expressing CIS43 had sustained antibody levels of approximately 300 μg/mL and mediated protection against sequential malaria challenges up to 36 weeks. Based on these data, CIS43LS has advanced to phase I clinical trials, and AAV delivery provides a potential next-generation approach for malaria prevention.

Anti-HIV-1 broadly neutralizing antibodies (bNAbs) have the dual potential of mediating virus neutralization and antiviral effector functions through their Fab and Fc domains, respectively. So far, bNAbs with enhanced Fc effector functions in vitro have only been tested in NHPs during chronic simian-HIV (SHIV) infection. Here, we investigate the effects of administering in acute SHIV AD8-EO infection either wild-type (WT) bNAbs or bNAbs carrying the S239D/I332E/A330L (DEL) mutation, which increases binding to FcγRs. Emergence of virus in plasma and lymph nodes (LNs) was delayed by bNAb treatment and occurred earlier in monkeys given DEL bNAbs than in those given WT bNAbs, consistent with faster clearance of DEL bNAbs from plasma. DEL bNAb-treated monkeys had higher levels of circulating virus-specific IFNγ single-producing CD8 + CD69 + T cells than the other groups. In LNs, WT bNAbs were evenly distributed between follicular and extrafollicular areas, but DEL bNAbs predominated in the latter. At week 8 post-challenge, LN monocytes and NK cells from DEL bNAb-treated monkeys upregulated proinflammatory signaling pathways and LN T cells downregulated TNF signaling via NF-κB. Overall, bNAbs with increased affinity to FcγRs shape innate and adaptive cellular immunity, which may be important to consider in future strategies of passive bNAb therapy. In this work, the authors study the immunological and virological effects of administering either wild-type anti-HIV-1 broadly neutralizing antibodies (bNAbs) or bNAbs with a mutation that increases binding to Fc-gamma receptors (FcγRs) to rhesus macaques in the acute phase of SHIV AD8-EO infection.

BackgroundPancreatic ductal adenocarcinoma (PDAC) is a devastating disease for which 5-year overall survival of early-stage disease is < 50% among patients fit enough to undergo aggressive surgical resection and combination chemotherapy. Immunotherapy for PDAC has shown limited efficacy in part due to heterogeneity of tumor antigens and an immunosuppressive tumor microenvironment. DOC1021 is a cell-based immunotherapy derived from the full complement of autologous tumor antigens. It leverages p38MAPK and mTORC1 signaling cascades to initiate cDC1-like skewing of monocyte-derived DC, generating downstream development of CD8+ tissue-homing, cytolytic memory effectors. Here we report results of Group A of a phase I study in which patients with potentially resectable PDAC received DOC1021 after surgical resection and standard neoadjuvant and/or adjuvant therapy.MethodsDOC1021 was prepared from mobilized peripheral blood mononuclear cells loaded with autologous tumor lysate and amplified tumor mRNA extracted from tumor tissue harvested at the time of resection. After completion of all standard therapy, vaccine was injected by CT guidance near lymph nodes in the post-operative surgical bed every other week for three doses administered concurrently with 6 weekly subcutaneous doses of peg-IFN. Two dose levels of 3.5 x 106 and 1.5 x 107 total vaccine cells were tested.ResultsSeven patients (median age: 58.0 years, range: 49–71) underwent DOC1021 vaccination after R0 or R1 resection and standard neoadjuvant and/or adjuvant therapy. Five patients received all 3 planned doses whereas 2 patients received only 1 or 2 doses due to peg-IFN attributable cytopenias. A protocol amendment incorporating dose modifications of peg-IFN dosing was subsequently instituted. Vaccines generated for 11 additional subjects were not administered due to progression during adjuvant chemotherapy. The most common DOC1021-related adverse events were mild flu-like symptoms, and no dose-limiting toxicities were observed. At the time of this analysis, 5 patients are alive (3 of whom remain relapse-free), with post-operative survival times of 45, 44, 25, 9.7 and 9.6 months. Gene expression profiling of individual DOC1021 vaccines and peripheral blood T-cell responses are ongoing.ConclusionsDOC1021 immunotherapy after surgical resection and standard neoadjuvant and/or adjuvant therapy is safe and feasible in patients with potentially resectable PDAC. An ongoing arm (Group B) is evaluating DOC1021 vaccination after surgery but prior to administration of adjuvant chemotherapy.AcknowledgementsThis study was supported in part by a grant from Cancer Cures 4 Kids (to WKD) with additional support from Diakonos Oncology Corporation. This study was further supported in part by the Baylor College of Medicine flow cytometry core facility under the academic leadership of Dr. Christine Beeton and the expert assistance of Mr. Joel M. Sederstrom.Ethics ApprovalThis study is being completed under oversight from the WCG institutional review board (reference #20192574), the Baylor College of Medicine institutional review board, an independent data safety monitoring board, and the Food and Drug Administration. All patients provided written, informed consent prior to enrollment.

Anti-HIV-1 broadly neutralizing antibodies (bNAbs) have the dual potential of mediating virus neutralization and antiviral effector functions through their Fab and Fc domains, respectively. So far, bNAbs with enhanced Fc effector functions in vitro have only been tested in NHPs during chronic simian-HIV (SHIV) infection. Here, we investigate the effects of administering in acute SHIV infection either wild-type (WT) bNAbs or bNAbs carrying the S239D/I332E/A330L (DEL) mutation, which increases binding to FcγRs. Emergence of virus in plasma and lymph nodes (LNs) was delayed by bNAb treatment and occurred earlier in monkeys given DEL bNAbs than in those given WT bNAbs, consistent with faster clearance of DEL bNAbs from plasma. DEL bNAb-treated monkeys had higher levels of circulating virus-specific IFNγ single-producing CD8 CD69 T cells than the other groups. In LNs, WT bNAbs were evenly distributed between follicular and extrafollicular areas, but DEL bNAbs predominated in the latter. At week 8 post-challenge, LN monocytes and NK cells from DEL bNAb-treated monkeys upregulated proinflammatory signaling pathways and LN T cells downregulated TNF signaling via NF-κB. Overall, bNAbs with increased affinity to FcγRs shape innate and adaptive cellular immunity, which may be important to consider in future strategies of passive bNAb therapy.

Multisystem Inflammatory Syndrome in Children (MIS-C) is a severe complication of SARS-CoV-2 infection characterized by multi-organ involvement and inflammation. Testing of cellular function ex vivo to understand the aberrant immune response in MIS-C is limited. Despite strong antibody production in MIS-C, SARS-CoV-2 nucleic acid testing can remain positive for 4-6 weeks after infection. Therefore, we hypothesized that dysfunctional cell-mediated antibody responses downstream of antibody production may be responsible for delayed clearance of viral products in MIS-C. In MIS-C, monocytes were hyperfunctional for phagocytosis and cytokine production, while natural killer (NK) cells were hypofunctional for both killing and cytokine production. The decreased NK cell cytotoxicity correlated with an NK exhaustion marker signature and systemic IL-6 levels. Potentially providing a therapeutic option, cellular engagers of CD16 and SARS-CoV-2 proteins were found to rescue NK cell function in vitro. Together, our results reveal dysregulation in antibody-mediated cellular responses unique to MIS-C that likely contribute to the immune pathology of this disease.Multisystem Inflammatory Syndrome in Children (MIS-C) is a severe complication of SARS-CoV-2 infection characterized by multi-organ involvement and inflammation. Testing of cellular function ex vivo to understand the aberrant immune response in MIS-C is limited. Despite strong antibody production in MIS-C, SARS-CoV-2 nucleic acid testing can remain positive for 4-6 weeks after infection. Therefore, we hypothesized that dysfunctional cell-mediated antibody responses downstream of antibody production may be responsible for delayed clearance of viral products in MIS-C. In MIS-C, monocytes were hyperfunctional for phagocytosis and cytokine production, while natural killer (NK) cells were hypofunctional for both killing and cytokine production. The decreased NK cell cytotoxicity correlated with an NK exhaustion marker signature and systemic IL-6 levels. Potentially providing a therapeutic option, cellular engagers of CD16 and SARS-CoV-2 proteins were found to rescue NK cell function in vitro. Together, our results reveal dysregulation in antibody-mediated cellular responses unique to MIS-C that likely contribute to the immune pathology of this disease.