Antibody and T cell responses to COVID-19 vaccination in patients receiving anticancer therapies.

BACKGROUND: Patients with cancer were excluded from phase 3 COVID-19 vaccine trials, and the immunogenicity and side effect profiles of these vaccines in this population is not well understood. Patients with cancer can be immunocompromised from chemotherapy, corticosteroids, or the cancer itself, which may affect cellular and/or humoral responses to vaccination. PD-1 is expressed on T effector cells, T follicular helper cells and B cells, leading us to hypothesize that anti-PD-1 immunotherapies may augment antibody or T cell generation after vaccination. METHODS: Antibodies to the SARS-CoV-2 receptor binding domain (RBD) and spike protein were assessed in patients with cancer (n=118) and healthy donors (HD, n=22) after 1, 2 or 3 mRNA vaccine doses. CD4+ and CD8+ T cell reactivity to wild-type (WT) or B.1.617.2 (delta) spike peptides was measured by intracellular cytokine staining. RESULTS: Oncology patients without prior COVID-19 infections receiving immunotherapy (n=36), chemotherapy (n=15), chemoimmunotherapy (n=6), endocrine or targeted therapies (n=6) and those not on active treatment (n=26) had similar RBD and Spike IgG antibody titers to HDs after two vaccinations. Contrary to our hypothesis, PD-1 blockade did not augment antibody titers or T cell responses. Patients receiving B-cell directed therapies (n=14) including anti-CD20 antibodies and multiple myeloma therapies had decreased antibody titers, and 9/14 of these patients were seronegative for RBD antibodies. No differences were observed in WT spike-reactive CD4+ and CD8+ T cell generation between treatment groups. 11/13 evaluable patients seronegative for RBD had a detectable WT spike-reactive CD4+ T cell response. T cells cross-reactive against the B.1.617.2 variant spike peptides were detected in 31/59 participants. Two patients with prior immune checkpoint inhibitor-related adrenal insufficiency had symptomatic hypoadrenalism after vaccination. CONCLUSIONS: COVID-19 vaccinations are safe and immunogenic in patients with solid tumors, who developed similar antibody and T cell responses compared with HDs. Patients on B-cell directed therapies may fail to generate RBD antibodies after vaccination and should be considered for prophylactic antibody treatments. Many seronegative patients do develop a T cell response, which may have an anti-viral effect. Patients with pre-existing adrenal insufficiency may need to take stress dose steroids during vaccination to avoid adrenal crisis.

Severe COVID-19 infection is associated with aberrant cytokine production by infected lung epithelial cells rather than by systemic immune dysfunction.

The mechanisms explaining progression to severe COVID-19 remain poorly understood. It has been proposed that immune system dysregulation/over-stimulation may be implicated, but it is not clear how such processes would lead to respiratory failure. We performed comprehensive multiparameter immune monitoring in a tightly controlled cohort of 128 COVID-19 patients, and used the ratio of oxygen saturation to fraction of inspired oxygen (SpO2 / FiO2) as a physiologic measure of disease severity. Machine learning algorithms integrating 139 parameters identified IL-6 and CCL2 as two factors predictive of severe disease, consistent with the therapeutic benefit observed with anti-IL6-R antibody treatment. However, transcripts encoding these cytokines were not detected among circulating immune cells. Rather, in situ analysis of lung specimens using RNAscope and immunofluorescent staining revealed that elevated IL-6 and CCL2 were dominantly produced by infected lung type II pneumocytes. Severe disease was not associated with higher viral load, deficient antibody responses, or dysfunctional T cell responses. These results refine our understanding of severe COVID-19 pathophysiology, indicating that aberrant cytokine production by infected lung epithelial cells is a major driver of immunopathology. We propose that these factors cause local immune regulation towards the benefit of the virus.

COVIDOSE: A Phase II Clinical Trial of Low-Dose Tocilizumab in the Treatment of Noncritical COVID-19 Pneumonia.

Interleukin-6 (IL-6)-mediated hyperinflammation may contribute to the mortality of coronavirus disease 2019 (COVID-19). The IL-6 receptor-blocking monoclonal antibody tocilizumab has been repurposed for COVID-19, but prospective trials and dose-finding studies in COVID-19 have not yet fully reported. We conducted a single-arm phase II trial of low-dose tocilizumab in nonintubated hospitalized adult patients with COVID-19, radiographic pulmonary infiltrate, fever, and C-reactive protein (CRP) ≥ 40 mg/L. We hypothesized that doses significantly lower than the emerging standards of 400 mg or 8 mg/kg would resolve clinical and laboratory indicators of hyperinflammation. A dose range from 40 to 200 mg was evaluated, with allowance for one repeat dose at 24 to 48 hours. The primary objective was to assess the relationship of dose to fever resolution and CRP response. Thirty-two patients received low-dose tocilizumab, with the majority experiencing fever resolution (75%) and CRP decline consistent with IL-6 pathway abrogation (86%) in the 24-48 hours following drug administration. There was no evidence of a relationship between dose and fever resolution or CRP decline over the dose range of 40-200 mg. Within the 28-day follow-up, 5 (16%) patients died. For patients who recovered, median time to clinical recovery was 3 days (interquartile range, 2-5). Clinically presumed and/or cultured bacterial superinfections were reported in 5 (16%) patients. Low-dose tocilizumab was associated with rapid improvement in clinical and laboratory measures of hyperinflammation in hospitalized patients with COVID-19. Results of this trial provide rationale for a randomized, controlled trial of low-dose tocilizumab in COVID-19.

COVIDOSE: Low-dose tocilizumab in the treatment of Covid-19.

Background Interleukin-6 (IL-6)-mediated hyperinflammation may contribute to the high mortality of coronavirus disease 2019 (Covid-19). Tocilizumab, an IL-6 receptor blocking monoclonal antibody, has been repurposed for Covid-19, but prospective trials and dose-finding studies in Covid-19 are lacking. Methods We conducted a phase 2 trial of low-dose tocilizumab in hospitalized adult patients with Covid-19, radiographic pulmonary infiltrate, fever, and C-reactive protein (CRP) >= 40 mg/L who did not require mechanical ventilation. Dose cohorts were determined by a trial Operations Committee, stratified by CRP and epidemiologic risk factors. A range of doses from 40 to 200 mg (low-dose tocilizumab) was evaluated, with allowance for one repeat dose at 24-48 hours. The primary objective was to assess the relationship of dose to fever resolution and CRP response. Outcomes were compared with retrospective controls with Covid-19. Correlative studies evaluating host antibody response were performed in parallel. Findings A total of 32 patients received low-dose tocilizumab. This cohort had improved fever resolution (75.0% vs. 34.2%, p = 0.001) and CRP decline (86.2% vs. 14.3%, p < 0.001) in the 24-48 hours following drug administration, as compared to the retrospective controls (N=41). The probabilities of fever resolution or CRP decline did not appear to be dose-related in this small study (p=0.80 and p=0.10, respectively). Within the 28-day follow-up, 5 (15.6%) patients died. For patients who recovered, median time to clinical recovery was 3 days (IQR, 2-5). Clinically presumed and/or cultured bacterial superinfections were reported in 5 (15.6%) patients. Correlative biological studies demonstrated that tocilizumab-treated patients produced anti-SARS-CoV-2 antibodies comparable to controls. Interpretation Low-dose tocilizumab was associated with rapid improvement in clinical and laboratory measures of hyperinflammation in hospitalized patients with Covid-19. Results of this trial and its correlative biological studies provide rationale for a randomized, controlled trial of low-dose tocilizumab in Covid-19.

Tumor heterogeneity and clonal cooperation influence the immune selection of IFN-γ-signaling mutant cancer cells.

PD-1/PD-L1 blockade can promote robust tumor regression yet secondary resistance often occurs as immune selective pressure drives outgrowth of resistant tumor clones. Here using a genome-wide CRISPR screen in B16.SIY melanoma cells, we confirm Ifngr2 and Jak1 as important genes conferring sensitivity to T cell-mediated killing in vitro. However, when implanted into mice, these Ifngr2- and Jak1-deficient tumors paradoxically are better controlled immunologically. This phenotype maps to defective PD-L1 upregulation on mutant tumor cells, which improves anti-tumor efficacy of CD8+ T cells. To reconcile these observations with clinical reports of anti-PD-1 resistance linked to emergence of IFN-γ signaling mutants, we show that when mixed with wild-type tumor cells, IFN-γ-insensitive tumor cells indeed grow out, which depends upon PD-L1 expression by wild-type cells. Our results illustrate the complexity of functions for IFN-γ in anti-tumor immunity and demonstrate that intratumor heterogeneity and clonal cooperation can contribute to immunotherapy resistance.

Influenza Virus Vaccination Elicits Poorly Adapted B Cell Responses in Elderly Individuals.

Influenza is a leading cause of death in the elderly, and the vaccine protects only a fraction of this population. A key aspect of antibody-mediated anti-influenza virus immunity is adaptation to antigenically distinct epitopes on emerging strains. We examined factors contributing to reduced influenza vaccine efficacy in the elderly and uncovered a dramatic reduction in the accumulation of de novo immunoglobulin gene somatic mutations upon vaccination. This reduction is associated with a significant decrease in the capacity of antibodies to target the viral glycoprotein, hemagglutinin (HA), and critical protective epitopes surrounding the HA receptor-binding domain. Immune escape by antigenic drift, in which viruses generate mutations in key antigenic epitopes, becomes highly exaggerated. Because of this reduced adaptability, most B cells activated in the elderly cohort target highly conserved but less potent epitopes. Given these findings, vaccines driving immunoglobulin gene somatic hypermutation should be a priority to protect elderly individuals.

Intratumoral CD8+ T-cell Apoptosis Is a Major Component of T-cell Dysfunction and Impedes Antitumor Immunity.

Subsets of human tumors are infiltrated with tumor antigen-specific CD8+ T cells [tumor-infiltrating lymphocytes (TILs)] despite tumor progression. These TILs are thought to be inactivated by the immunosuppressive tumor microenvironment, through the engagement of inhibitory receptors such as CTLA-4 and PD-1. However, antigen-specific CD8+ TILs are not functionally inert but are undergoing activation in situ Here, we show that antigen-specific CD8+ TILs are actively proliferating, yet also undergo high rates of apoptosis, leading to a vicious cycle of activation and death that limits immune efficacy. Preventing CD8+ TIL apoptosis by Bcl-xL overexpression enabled accumulation and improved tumor control. Effective combination immunotherapy with an agonist 4-1BB mAb plus either CTLA-4 or PD-L1 neutralization led to a marked accumulation of specific CD8+ TILs through decreased apoptosis rather than increased T-cell entry or proliferation. Our data suggest that antigen-driven apoptosis of CD8+ TILs is a barrier to effective spontaneous antitumor immunity and should be considered as a critical factor in the development of cancer immunotherapies. Cancer Immunol Res; 6(1); 14-24. ©2017 AACR.

Inclusion of Strep-tag II in design of antigen receptors for T-cell immunotherapy.

Adoptive immunotherapy with genetically engineered T cells has the potential to treat cancer and other diseases. The introduction of Strep-tag II sequences into specific sites in synthetic chimeric antigen receptors or natural T-cell receptors of diverse specificities provides engineered T cells with a marker for identification and rapid purification, a method for tailoring spacer length of chimeric receptors for optimal function, and a functional element for selective antibody-coated, microbead-driven, large-scale expansion. These receptor designs facilitate cGMP manufacturing of pure populations of engineered T cells for adoptive T-cell therapies and enable in vivo tracking and retrieval of transferred cells for downstream research applications.