Phase 1 study combining elotuzumab with autologous stem cell transplant and lenalidomide for multiple myeloma.

BACKGROUND: Autologous stem cell transplantation (ASCT) after induction therapy improves disease-free survival for patients with multiple myeloma (MM). While the goal of ASCT is to render a minimal disease state, it is also associated with eradication of immunosuppressive cells, and we hypothesize that early introduction of immunotherapy post-ASCT may provide a window of opportunity to boost treatment efficacy. METHODS: We conducted a phase 1 clinical trial to investigate the application of autologous lymphocyte infusion and anti-SLAMF7 monoclonal antibody, elotuzumab, after ASCT in patients with newly diagnosed MM previously treated with induction therapy. In addition to CD34+ stem cells, peripheral blood mononuclear cells were harvested prior to transplant and infused on day 3 after stem cell infusion to accelerate immune reconstitution and provide autologous natural killer (NK) cells that are essential to the mechanism of elotuzumab. Elotuzumab was administered starting on day 4 and then every 28 days after until 1 year post-ASCT. Cycles 4-12 were administered with standard-of-care lenalidomide maintenance. RESULTS: All subjects were evaluated for safety, and 13 of 15 subjects completed the treatment protocol. At 1 year post-ASCT, the disease status of enrolled subjects was as follows: five stringent complete responses, one complete response, six very good partial responses, one partial response, and two progressive diseases. The treatment plan was well tolerated, with most grade 3 and 4 AEs being expected hematologic toxicities associated with ASCT. Correlative analysis of the immune microenvironment demonstrated a trend toward reduced regulatory T cells during the first 3 months post-transplant followed by an increase in NK cells and monocytes in patients achieving a complete remission. CONCLUSIONS: This phase 1 clinical trial demonstrates that early introduction of immunotherapy after ASCT is well tolerated and shows promising disease control in patients with MM, accompanied by favorable changes in the immune microenvironment. TRIAL REGISTRATION NUMBER: NCT02655458.

Host-microbe multiomic profiling reveals age-dependent immune dysregulation associated with COVID-19 immunopathology.

Age is a major risk factor for severe coronavirus disease 2019 (COVID-19), yet the mechanisms behind this relationship have remained incompletely understood. To address this, we evaluated the impact of aging on host immune response in the blood and the upper airway, as well as the nasal microbiome in a prospective, multicenter cohort of 1031 vaccine-naïve patients hospitalized for COVID-19 between 18 and 96 years old. We performed mass cytometry, serum protein profiling, anti-severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) antibody assays, and blood and nasal transcriptomics. We found that older age correlated with increased SARS-CoV-2 viral abundance upon hospital admission, delayed viral clearance, and increased type I interferon gene expression in both the blood and upper airway. We also observed age-dependent up-regulation of innate immune signaling pathways and down-regulation of adaptive immune signaling pathways. Older adults had lower naïve T and B cell populations and higher monocyte populations. Over time, older adults demonstrated a sustained induction of pro-inflammatory genes and serum chemokines compared with younger individuals, suggesting an age-dependent impairment in inflammation resolution. Transcriptional and protein biomarkers of disease severity differed with age, with the oldest adults exhibiting greater expression of pro-inflammatory genes and proteins in severe disease. Together, our study finds that aging is associated with impaired viral clearance, dysregulated immune signaling, and persistent and potentially pathologic activation of pro-inflammatory genes and proteins.

Acute Effects of Growth Hormone on the Cellular Immunologic Landscape in Pediatric Patients.

INTRODUCTION: Growth hormone (GH) and the immune system have multiple bidirectional interactions. Data about the acute effects of GH on the immune system are lacking. The objective of our study was to evaluate the acute effects of GH on the immune system using time-of-flight mass cytometry. METHODS: This was a prospective study of pediatric patients who were being evaluated for short stature and underwent a GH stimulation test at a tertiary care center. Blood samples for immunologic markers, i.e., complete blood count (CBC) and time of flight mass cytometry (CyTOF), were collected at baseline (T0) and over the course of three hours (T3) of the test. Differences in immune profiling in patients by timepoint (T0, T3) and GH response (growth hormone sufficient (GHS) versus growth hormone deficient (GHD)) were calculated using a two-way ANOVA test.  Results: A total of 54 patients (39 boys and 15 girls) aged five to 18 years were recruited. Twenty-two participants tested GHD (peak GH <10 ng/ml). The CyTOF analysis showed a significant increase from T0 to T3 in granulocyte percentage, monocyte count, and dendritic cell (DC) count; in contrast, a significant decrease was seen in T lymphocytes (helper and cytotoxic) and IgD+ B lymphocytes. The CBC analysis supported these findings: an increase in total white blood cell count, absolute neutrophil count, and neutrophil percentage; a decrease in absolute lymphocyte count, lymphocyte percentage, absolute eosinophil count, and absolute monocyte count. No significant differences were found between CBC/CyTOF measurements and GH status at either time. CONCLUSIONS: This study provides the first high-resolution map of acute changes in the immune system with GH stimulation. This implies a key role for GH in immunomodulatory function.

A phase II open-label randomized clinical trial of preoperative durvalumab or durvalumab plus tremelimumab in resectable head and neck squamous cell carcinoma.

PURPOSE: Clinical implications of neoadjuvant immunotherapy in patients with locally advanced but resectable head and neck squamous cell carcinoma (HNSCC) remain largely unexplored. PATIENTS AND METHODS: Patients with resectable HNSCC were randomized to receive a single dose of preoperative durvalumab (D) with or without tremelimumab (T) before resection, followed by postoperative (chemo)radiation based on multidisciplinary discretion and 1-year D treatment. Artificial intelligence (AI)-powered spatial distribution analysis of tumor-infiltrating lymphocytes and high-dimensional profiling of circulating immune cells tracked dynamic intratumoral and systemic immune responses. RESULTS: Of the 48 patients enrolled (D: 24 patients, D+T: 24 patients), 45 underwent surgical resection per protocol (D: 21 patients; D+T: 24 patients). D+/-T had a favorable safety profile and did not delay surgery. Distant recurrence-free survival (DRFS) was significantly better in patients treated with D+T than in those treated with D monotherapy. AI-powered whole-slide image analysis demonstrated that D+T significantly reshaped the tumor microenvironment toward immune-inflamed phenotypes, in contrast to D monotherapy or cytotoxic chemotherapy. High-dimensional profiling of circulating immune cells revealed a significant expansion of T cell subsets characterized by proliferation and activation in response to D+T therapy, which was rare following D monotherapy. Importantly, expansion of specific clusters in CD8+ T cells and non-regulatory CD4+ T cells with activation and exhaustion programs was associated with prolonged DRFS in patients treated with D+T. CONCLUSIONS: Preoperative D+/-T is feasible and may benefit patients with resectable HNSCC. Distinct changes in the tumor microenvironment and circulating immune cells were induced by each treatment regimen, warranting further investigation.

Immune and Neural Response to Acute Social Stress in Adolescent Humans and Rodents.

Studies in adults have linked stress-related activation of the immune system to the manifestation of psychiatric conditions. Using a translational design, this study aimed to examine the impact of social stress on immune activity in adolescents and on neuronal activity in a preclinical mouse model. Participants were 31 adolescents (ages 12-19), including 25 with mood and anxiety symptoms. Whole-blood samples were collected before and after the Trier Social Stress Test (TSST), a stress-inducing public speaking task, then cultured for 6 hours in the presence and absence of the inflammatory endotoxin lipopolysaccharide (LPS). Effects of TSST and LPS on 41 immune biomarkers were examined using repeated-measures analysis of variance. Separately, juvenile (8-week-old) male mice were non-stressed or exposed to reminder social defeat then intraperitoneally injected with saline or LPS (n = 6/group). Brains were perfused and collected for immunohistochemistry and confocal microscopy at 0, 1, 6, and 24 hours post-injection. Activity was determined by the density of cFos-positive neurons in the paraventricular hypothalamus, paraventricular thalamus, and basolateral amygdala, regions known to show sustained activation to immunological challenge. Analyses in the adolescent study indicated a strong effect of LPS but no effects of TSST or TSST×LPS interaction on immune biomarkers. Similarly, reminder social defeat did not induce sustained neuronal activity changes comparable to LPS immunological challenge in juvenile mice. Our convergent findings across species suggest that the acute immune response to stress documented in adults is not present in youth. Thus, aging and chronicity effects may play an important role in the inflammatory response to acute psychosocial stress.

Imprinted immune abnormalities in liver transplant patients cured of hepatitis C with antiviral drugs.

Chronic HCV infection induces interferon and dysregulates immune responses through inflammation and chronic antigenic stimulation. Antiviral drugs can cure HCV, providing a unique opportunity to examine the immunological restoration that does and does not occur when a chronic viral infection is eradicated. We quantified blood cytokines levels and used mass cytometry to immunophenotype peripheral blood mononuclear cells before and after HCV cure in 2 groups of patients and controls. At baseline, serum interferon α and soluble CD163 (a macrophage product) were elevated in both liver transplant and nonliver transplant patients compared to controls; the frequencies of several peripheral blood mononuclear cell populations differed from controls; and programmed death protein 1-positivity was increased in nearly all T cell subsets. Many abnormalities persisted after HCV cure, including elevated programmed death protein 1 expression on CD4 naïve and central memory T cells, elevated soluble CD163, and expansion of the plasmablast/plasma cell compartment. Several myeloid-lineage subsets, including Ag-presenting dendritic cells, remained dysregulated. In mechanistic studies, interferon α treatment increased programmed death protein 1 on human T cells and increased T cell receptor signaling. The data identify immunological abnormalities that persist after curative HCV treatment. Before cure, high levels of interferon α may stimulate programmed death protein 1 expression on human T cells, causing persistent functional changes.

High-dimensional profiling of regulatory T cells in psoriasis reveals an impaired skin-trafficking property.

BACKGROUND: Psoriasis is a chronic inflammatory skin disease with a Th17-skewed immune phenotype. Although it has been generally accepted that regulatory T cells (Tregs) in lesional psoriatic skin have functional impairment due to the local inflammatory microenvironment, the molecular properties of skin-homing psoriatic Tregs have not been well explored. METHODS: We designed an extensive 39 marker mass cytometry (CyTOF) panel to deeply profile the immune landscape of skin-homing Tregs from 31 people with psoriasis stratified by psoriasis area severity index score as mild (n = 15) to moderate-severe (n = 16) and 32 healthy controls. We further validated the findings with an in-vitro chemokine-mediated Treg migration assay, immunofluorescent imaging of normal and psoriatic lesional skin and analysed public single-cell RNA-sequencing datasets to expand upon our findings into the local tissue microenvironments. FINDINGS: We discovered an overall decrease in CLAhi Tregs and specifically, CLAhiCCR5+ Tregs in psoriasis. Functional markers CD39 and FoxP3 were elevated in psoriatic Tregs. However, CCR7 expression was significantly increased while CCR4 and CLA expression was reduced in psoriatic Tregs and CLAhi Tregs, which was associated with disease severity. Moreover, psoriatic Tregs revealed increased migratory capacity towards CCR7's ligands, CCL19/CCL21. Interrogation of public single-cell RNA sequencing data confirmed reduced expression of skin-trafficking markers in lesional-skin Tregs compared to non-lesioned skin, further substantiated by immunofluorescent staining. INTERPRETATION: Psoriatic circulating Tregs showed an impaired skin-trafficking phenotype thus leading to insufficient suppression of ongoing inflammation in the lesional skin, expanding upon our current understanding of the impairment of Treg-mediated immunosuppression in psoriasis. FUNDING: This research was supported by the Basic Science Research Program through the National Research Foundation of Korea funded by the Ministry of Science and Information and Communications Technology (2020R1C1C1014513, 2021R1A4A5032185, 2020R1F1A1073692); and the new faculty research seed money grant of Yonsei University College of Medicine for 2021 (2021-32-0033).

Host-Microbe Multiomic Profiling Reveals Age-Dependent COVID-19 Immunopathology.

Age is a major risk factor for severe coronavirus disease-2019 (COVID-19), yet the mechanisms responsible for this relationship have remained incompletely understood. To address this, we evaluated the impact of aging on host and viral dynamics in a prospective, multicenter cohort of 1,031 patients hospitalized for COVID-19, ranging from 18 to 96 years of age. We performed blood transcriptomics and nasal metatranscriptomics, and measured peripheral blood immune cell populations, inflammatory protein expression, anti-SARS-CoV-2 antibodies, and anti-interferon (IFN) autoantibodies. We found that older age correlated with an increased SARS-CoV-2 viral load at the time of admission, and with delayed viral clearance over 28 days. This contributed to an age-dependent increase in type I IFN gene expression in both the respiratory tract and blood. We also observed age-dependent transcriptional increases in peripheral blood IFN-γ, neutrophil degranulation, and Toll like receptor (TLR) signaling pathways, and decreases in T cell receptor (TCR) and B cell receptor signaling pathways. Over time, older adults exhibited a remarkably sustained induction of proinflammatory genes (e.g., CXCL6) and serum chemokines (e.g., CXCL9) compared to younger individuals, highlighting a striking age-dependent impairment in inflammation resolution. Augmented inflammatory signaling also involved the upper airway, where aging was associated with upregulation of TLR, IL17, type I IFN and IL1 pathways, and downregulation TCR and PD-1 signaling pathways. Metatranscriptomics revealed that the oldest adults exhibited disproportionate reactivation of herpes simplex virus and cytomegalovirus in the upper airway following hospitalization. Mass cytometry demonstrated that aging correlated with reduced naïve T and B cell populations, and increased monocytes and exhausted natural killer cells. Transcriptional and protein biomarkers of disease severity markedly differed with age, with the oldest adults exhibiting greater expression of TLR and inflammasome signaling genes, as well as proinflammatory proteins (e.g., IL6, CXCL8), in severe COVID-19 compared to mild/moderate disease. Anti-IFN autoantibody prevalence correlated with both age and disease severity. Taken together, this work profiles both host and microbe in the blood and airway to provide fresh insights into aging-related immune changes in a large cohort of vaccine-naïve COVID-19 patients. We observed age-dependent immune dysregulation at the transcriptional, protein and cellular levels, manifesting in an imbalance of inflammatory responses over the course of hospitalization, and suggesting potential new therapeutic targets.

Multi-omics Analysis Reveals Immune Features Associated with Immunotherapy Benefit in Patients with Squamous Cell Lung Cancer from Phase III Lung-MAP S1400I Trial.

PURPOSE: Identifying molecular and immune features to guide immune checkpoint inhibitor (ICI)-based regimens remains an unmet clinical need. EXPERIMENTAL DESIGN: Tissue and longitudinal blood specimens from phase III trial S1400I in patients with metastatic squamous non-small cell carcinoma (SqNSCLC) treated with nivolumab monotherapy (nivo) or nivolumab plus ipilimumab (nivo+ipi) were subjected to multi-omics analyses including multiplex immunofluorescence (mIF), nCounter PanCancer Immune Profiling Panel, whole-exome sequencing, and Olink. RESULTS: Higher immune scores from immune gene expression profiling or immune cell infiltration by mIF were associated with response to ICIs and improved survival, except regulatory T cells, which were associated with worse overall survival (OS) for patients receiving nivo+ipi. Immune cell density and closer proximity of CD8+GZB+ T cells to malignant cells were associated with superior progression-free survival and OS. The cold immune landscape of NSCLC was associated with a higher level of chromosomal copy-number variation (CNV) burden. Patients with LRP1B-mutant tumors had a shorter survival than patients with LRP1B-wild-type tumors. Olink assays revealed soluble proteins such as LAMP3 increased in responders while IL6 and CXCL13 increased in nonresponders. Upregulation of serum CXCL13, MMP12, CSF-1, and IL8 were associated with worse survival before radiologic progression. CONCLUSIONS: The frequency, distribution, and clustering of immune cells relative to malignant ones can impact ICI efficacy in patients with SqNSCLC. High CNV burden may contribute to the cold immune microenvironment. Soluble inflammation/immune-related proteins in the blood have the potential to monitor therapeutic benefit from ICI treatment in patients with SqNSCLC.

Features of acute COVID-19 associated with post-acute sequelae of SARS-CoV-2 phenotypes: results from the IMPACC study.

Post-acute sequelae of SARS-CoV-2 (PASC) is a significant public health concern. We describe Patient Reported Outcomes (PROs) on 590 participants prospectively assessed from hospital admission for COVID-19 through one year after discharge. Modeling identified 4 PRO clusters based on reported deficits (minimal, physical, mental/cognitive, and multidomain), supporting heterogenous clinical presentations in PASC, with sub-phenotypes associated with female sex and distinctive comorbidities. During the acute phase of disease, a higher respiratory SARS-CoV-2 viral burden and lower Receptor Binding Domain and Spike antibody titers were associated with both the physical predominant and the multidomain deficit clusters. A lower frequency of circulating B lymphocytes by mass cytometry (CyTOF) was observed in the multidomain deficit cluster. Circulating fibroblast growth factor 21 (FGF21) was significantly elevated in the mental/cognitive predominant and the multidomain clusters. Future efforts to link PASC to acute anti-viral host responses may help to better target treatment and prevention of PASC.

An IL-4 signalling axis in bone marrow drives pro-tumorigenic myelopoiesis.

Myeloid cells are known to suppress antitumour immunity1. However, the molecular drivers of immunosuppressive myeloid cell states are not well defined. Here we used single-cell RNA sequencing of human and mouse non-small cell lung cancer (NSCLC) lesions, and found that in both species the type 2 cytokine interleukin-4 (IL-4) was predicted to be the primary driver of the tumour-infiltrating monocyte-derived macrophage phenotype. Using a panel of conditional knockout mice, we found that only deletion of the IL-4 receptor IL-4Rα in early myeloid progenitors in bone marrow reduced tumour burden, whereas deletion of IL-4Rα in downstream mature myeloid cells had no effect. Mechanistically, IL-4 derived from bone marrow basophils and eosinophils acted on granulocyte-monocyte progenitors to transcriptionally programme the development of immunosuppressive tumour-promoting myeloid cells. Consequentially, depletion of basophils profoundly reduced tumour burden and normalized myelopoiesis. We subsequently initiated a clinical trial of the IL-4Rα blocking antibody dupilumab2-5 given in conjunction with PD-1/PD-L1 checkpoint blockade in patients with relapsed or refractory NSCLC who had progressed on PD-1/PD-L1 blockade alone (ClinicalTrials.gov identifier NCT05013450 ). Dupilumab supplementation reduced circulating monocytes, expanded tumour-infiltrating CD8 T cells, and in one out of six patients, drove a near-complete clinical response two months after treatment. Our study defines a central role for IL-4 in controlling immunosuppressive myelopoiesis in cancer, identifies a novel combination therapy for immune checkpoint blockade in humans, and highlights cancer as a systemic malady that requires therapeutic strategies beyond the primary disease site.

Peripheral Transcriptomics in Acute and Long-Term Kidney Dysfunction in SARS-CoV2 Infection.

Background: Acute kidney injury (AKI) is common in hospitalized patients with SARS-CoV2 infection despite vaccination and leads to long-term kidney dysfunction. However, peripheral blood molecular signatures in AKI from COVID-19 and their association with long-term kidney dysfunction are yet unexplored. Methods: In patients hospitalized with SARS-CoV2, we performed bulk RNA sequencing using peripheral blood mononuclear cells(PBMCs). We applied linear models accounting for technical and biological variability on RNA-Seq data accounting for false discovery rate (FDR) and compared functional enrichment and pathway results to a historical sepsis-AKI cohort. Finally, we evaluated the association of these signatures with long-term trends in kidney function. Results: Of 283 patients, 106 had AKI. After adjustment for sex, age, mechanical ventilation, and chronic kidney disease (CKD), we identified 2635 significant differential gene expressions at FDR<0.05. Top canonical pathways were EIF2 signaling, oxidative phosphorylation, mTOR signaling, and Th17 signaling, indicating mitochondrial dysfunction and endoplasmic reticulum (ER) stress. Comparison with sepsis associated AKI showed considerable overlap of key pathways (48.14%). Using follow-up estimated glomerular filtration rate (eGFR) measurements from 115 patients, we identified 164/2635 (6.2%) of the significantly differentiated genes associated with overall decrease in long-term kidney function. The strongest associations were 'autophagy', 'renal impairment via fibrosis', and 'cardiac structure and function'. Conclusions: We show that AKI in SARS-CoV2 is a multifactorial process with mitochondrial dysfunction driven by ER stress whereas long-term kidney function decline is associated with cardiac structure and function and immune dysregulation. Functional overlap with sepsis-AKI also highlights common signatures, indicating generalizability in therapeutic approaches. SIGNIFICANCE STATEMENT: Peripheral transcriptomic findings in acute and long-term kidney dysfunction after hospitalization for SARS-CoV2 infection are unclear. We evaluated peripheral blood molecular signatures in AKI from COVID-19 (COVID-AKI) and their association with long-term kidney dysfunction using the largest hospitalized cohort with transcriptomic data. Analysis of 283 hospitalized patients of whom 37% had AKI, highlighted the contribution of mitochondrial dysfunction driven by endoplasmic reticulum stress in the acute stages. Subsequently, long-term kidney function decline exhibits significant associations with markers of cardiac structure and function and immune mediated dysregulation. There were similar biomolecular signatures in other inflammatory states, such as sepsis. This enhances the potential for repurposing and generalizability in therapeutic approaches.

Integrated longitudinal multi-omics study identifies immune programs associated with COVID-19 severity and mortality in 1152 hospitalized participants.

Hospitalized COVID-19 patients exhibit diverse clinical outcomes, with some individuals diverging over time even though their initial disease severity appears similar. A systematic evaluation of molecular and cellular profiles over the full disease course can link immune programs and their coordination with progression heterogeneity. In this study, we carried out deep immunophenotyping and conducted longitudinal multi-omics modeling integrating ten distinct assays on a total of 1,152 IMPACC participants and identified several immune cascades that were significant drivers of differential clinical outcomes. Increasing disease severity was driven by a temporal pattern that began with the early upregulation of immunosuppressive metabolites and then elevated levels of inflammatory cytokines, signatures of coagulation, NETosis, and T-cell functional dysregulation. A second immune cascade, predictive of 28-day mortality among critically ill patients, was characterized by reduced total plasma immunoglobulins and B cells, as well as dysregulated IFN responsiveness. We demonstrated that the balance disruption between IFN-stimulated genes and IFN inhibitors is a crucial biomarker of COVID-19 mortality, potentially contributing to the failure of viral clearance in patients with fatal illness. Our longitudinal multi-omics profiling study revealed novel temporal coordination across diverse omics that potentially explain disease progression, providing insights that inform the targeted development of therapies for hospitalized COVID-19 patients, especially those critically ill.

Autonomic Neuropathy is Associated with More Densely Interconnected Cytokine Networks in People with HIV.

The autonomic nervous system (ANS) plays a complex role in the regulation of the immune system, with generally inhibitory effects via activation of ß-adrenergic receptors on immune cells. We hypothesized that HIV-associated autonomic neuropathy (HIV-AN) would result in immune hyperresponsiveness which could be depicted using network analyses. Forty-two adults with well-controlled HIV underwent autonomic testing to yield the Composite Autonomic Severity Score (CASS). The observed range of CASS was 2-5, consistent with normal to moderate HIV-AN. To construct the networks, participants were divided into 4 groups based on the CASS (i.e., 2, 3, 4 or 5). Forty-four blood-based immune markers were included as nodes in all networks and the connections (i.e., edges) between pairs of nodes were determined by their bivariate Spearman's Rank Correlation Coefficient. Four centrality measures (strength, closeness, betweenness and expected influence) were calculated for each node in each network. The median value of each centrality measure across all nodes in each network was calculated as a quantitative representation of network complexity. Graphical representation of the four networks revealed greater complexity with increasing HIV-AN severity. This was confirmed by significant differences in the median value of all four centrality measures across the networks (p ≤ 0.025 for each). Among people with HIV, HIV-AN is associated with stronger and more numerous positive correlations between blood-based immune markers. Findings from this secondary analysis can be used to generate hypotheses for future studies investigating HIV-AN as a mechanism contributing to the chronic immune activation observed in HIV.

Gemcitabine and cisplatin plus nivolumab as organ-sparing treatment for muscle-invasive bladder cancer: a phase 2 trial.

Cystectomy is a standard treatment for muscle-invasive bladder cancer (MIBC), but it is life-altering. We initiated a phase 2 study in which patients with MIBC received four cycles of gemcitabine, cisplatin, plus nivolumab followed by clinical restaging. Patients achieving a clinical complete response (cCR) could proceed without cystectomy. The co-primary objectives were to assess the cCR rate and the positive predictive value of cCR for a composite outcome: 2-year metastasis-free survival in patients forgoing immediate cystectomy or

Immunophenotypic correlates of sustained MRD negativity in patients with multiple myeloma.

The role of the immune microenvironment in maintaining disease remission in patients with multiple myeloma (MM) is not well understood. In this study, we comprehensively profile the immune system in patients with newly diagnosed MM receiving continuous lenalidomide maintenance therapy with the aim of discovering correlates of long-term treatment response. Leveraging single-cell RNA sequencing and T cell receptor ß sequencing of the peripheral blood and CyTOF mass cytometry of the bone marrow, we longitudinally characterize the immune landscape in 23 patients before and one year after lenalidomide exposure. We compare patients achieving sustained minimal residual disease (MRD) negativity to patients who never achieved or were unable to maintain MRD negativity. We observe that the composition of the immune microenvironment in both the blood and the marrow varied substantially according to both MRD negative status and history of autologous stem cell transplant, supporting the hypothesis that the immune microenvironment influences the depth and duration of treatment response.

Understanding the heterogeneity of alloreactive natural killer cell function in kidney transplantation.

Human Natural Killer (NK) cells are heterogeneous lymphocytes regulated by variegated arrays of germline-encoded activating and inhibitory receptors. They acquire the ability to detect polymorphic self-antigen via NKG2A/HLA-E or KIR/HLA-I ligand interactions through an education process. Correlations among HLA/KIR genes, kidney transplantation pathology and outcomes suggest that NK cells participate in allograft injury, but mechanisms linking NK HLA/KIR education to antibody-independent pathological functions remain unclear. We used CyTOF to characterize pre- and post-transplant peripheral blood NK cell phenotypes/functions before and after stimulation with allogeneic donor cells. Unsupervised clustering identified unique NK cell subpopulations present in varying proportions across patients, each of which responded heterogeneously to donor cells based on donor ligand expression patterns. Analyses of pre-transplant blood showed that educated, NKG2A/KIR-expressing NK cells responded greater than non-educated subsets to donor stimulators, and this heightened alloreactivity persisted > 6 months post-transplant despite immunosuppression. In distinct test and validation sets of patients participating in two clinical trials, pre-transplant donor-induced release of NK cell Ksp37, a cytotoxicity mediator, correlated with 2-year and 5-year eGFR. The findings explain previously reported associations between NK cell genotypes and transplant outcomes and suggest that pre-transplant NK cell analysis could function as a risk-assessment biomarker for transplant outcomes.

Antitumor immunity as the basis for durable disease-free treatment-free survival in patients with metastatic urothelial cancer.

Cisplatin-based chemotherapy has been associated with durable disease control in a small subset of patients with metastatic urothelial cancer. However, the mechanistic basis for this phenomenon has remained elusive. Antitumor immunity may underlie these exceptional responders. In a phase II trial evaluating a phased schedule of gemcitabine and cisplatin followed by gemcitabine and cisplatin with ipilimumab for metastatic urothelial cancer, 4 of 36 patients achieved durable disease-free treatment-free survival (DDFTFS) and remain in remission over 5 years after enrolment on the study. We sought to identify the genomic and immunological mechanisms associated with functional cures of such patients. Whole exome sequencing was performed on pretreatment archival tumor tissue. Neoantigen prediction and ranking were performed using a novel pipeline. For a subset of patients with available biospecimens, selected peptides were tested for neoantigen-specific T cell reactivity in peripheral blood CD4+ and CD8+ T cells cultured with autologous antigen-presenting cells at baseline, postchemotherapy, and postchemotherapy and ipilimumab timepoints. Multiplex assays of serum protein analytes were also assessed at each time point. Serum proteomic analysis revealed that pretreatment, patients achieving DDFTFS demonstrated an immune activated phenotype with elevations in TH1 adaptive immunity, costimulatory molecules, and immune checkpoint markers. After combination cisplatin-based chemotherapy and ipilimumab treatment, DDFTFS patients again displayed enrichment for markers of adaptive immunity, as well as T cell cytotoxicity. CD27 was uniquely enriched in DDFTFS patients at all timepoints. Neoantigen reactivity was not detected in any patient at baseline or post two cycles of chemotherapy. Both CD4+ and CD8+ neoantigen-specific T cell reactivity was detected in two of two DDFTFS patients in comparison to zero of five non-DDFTFS patients after combination cisplatin-based chemotherapy and ipilimumab treatment. Antitumor immunity may underlie functional cures achieved in patients with metastatic urothelial cancer treated with cisplatin-based chemotherapy and immune checkpoint blockade. Probing the mechanistic basis for DDFTFS may facilitate the identification of biomarkers, therapeutic components, and optimal treatment sequences necessary to extend this ultimate goal to a larger subset of patients.