Affinity purification mass spectrometry analysis of PD-1 uncovers SAP as a new checkpoint inhibitor.

Programmed cell death-1 (PD-1) is an essential inhibitory receptor in T cells. Antibodies targeting PD-1 elicit durable clinical responses in patients with multiple tumor indications. Nevertheless, a significant proportion of patients do not respond to anti-PD-1 treatment, and a better understanding of the signaling pathways downstream of PD-1 could provide biomarkers for those whose tumors respond and new therapeutic approaches for those whose tumors do not. We used affinity purification mass spectrometry to uncover multiple proteins associated with PD-1. Among these proteins, signaling lymphocytic activation molecule-associated protein (SAP) was functionally and mechanistically analyzed for its contribution to PD-1 inhibitory responses. Silencing of SAP augmented and overexpression blocked PD-1 function. T cells from patients with X-linked lymphoproliferative disease (XLP), who lack functional SAP, were hyperresponsive to PD-1 signaling, confirming its inhibitory role downstream of PD-1. Strikingly, signaling downstream of PD-1 in purified T cell subsets did not correlate with PD-1 surface expression but was inversely correlated with intracellular SAP levels. Mechanistically, SAP opposed PD-1 function by acting as a molecular shield of key tyrosine residues that are targets for the tyrosine phosphatase SHP2, which mediates PD-1 inhibitory properties. Our results identify SAP as an inhibitor of PD-1 function and SHP2 as a potential therapeutic target in patients with XLP.

Risk of Toxicity After Initiating Immune Checkpoint Inhibitor Treatment in Patients With Rheumatoid Arthritis.

INTRODUCTION: Immune checkpoint inhibitors (ICIs) are increasingly used to treat advanced cancer. Rheumatoid arthritis (RA) is associated with an increased risk of malignancies; however, patients with RA have been excluded from ICI trials. In this study, we evaluated risk of toxicity after initiation of ICI treatment in RA patients. METHODS: We conducted a single-institution, medical records review analysis to assess the incidence of immune-related adverse events (irAEs) and autoimmune disease (AID) flares among patients with AIDs treated with ICIs from 2011 to 2018. A subgroup analysis for RA patients was performed with frequencies of irAEs and AID flares reported. RESULTS: Twenty-two patients with RA who were treated with ICI for malignancy were identified. At the time of ICI initiation, 86% had inactive RA disease activity. Immune-related adverse events occurred in 7 (32%) of patients, with 2 (9%) developing grade 3 (i.e., severe) irAEs. Immune checkpoint inhibitors were temporarily discontinued because of irAEs in 5 patients (23%), and permanently in 1 patient. Rheumatoid arthritis flares occurred in 12 patients (55%). Of those, 10 (83%) received oral corticosteroids with an adequate treatment response. CONCLUSIONS: Our analysis suggests that irAEs following ICI treatment are not increased among RA patients compared with other cancer patients. Heightened RA disease activity during ICI treatment is common, but most adverse events are manageable with oral corticosteroids, and few require permanent ICI discontinuation. A close collaboration between the oncologist and rheumatologist is advisable when considering ICIs in patients with RA.

SAP interacts with CD28 to inhibit PD-1 signaling in T lymphocytes.

T cell co-stimulation is important for the maintenance of immunologic tolerance. Co-inhibitory receptors including programmed cell death-1 (PD-1) confer peripheral tolerance to prevent autoimmunity. SAP (SH2D1A) is an adaptor molecule that is important in T cell signaling and has been shown to interact with signaling lymphocytic activation molecule (SLAM) family receptors also in the context of self-tolerance. We recently reported that SAP interferes with PD-1 function. In the current study, we investigated the levels of SAP and PD-1 in patients with rheumatoid arthritis (RA) to further understand what role they play in disease activity. We observed increased SAP levels in lymphocytes of RA patients and found that PD-1 levels correlated positively with RA disease activity. Additionally, we found that SAP interacts with CD28 to inhibit T cell signaling in vitro. This work demonstrates a putative molecular mechanism for SAP mediated PD-1 inhibition.

Immune checkpoint inhibitors and vasculitis.

PURPOSE OF REVIEW: Clinical use of immune checkpoint inhibitor (ICI) therapy has revolutionized the therapeutic landscape of cancer. By activating the immune system using monoclonal anti-CTLA-4 and PD(L)-1 antibodies, remission can be induced in previously terminal cancers. However, these breakthroughs come at a price. Multiple de-novo autoimmune illnesses, termed immune-related adverse events (irAEs), have been reported with patients increasingly being referred to rheumatologists with varying diagnoses. Among these are vasculitic syndromes, which may be limited to an organ or systemic and potentially-life threatening. Relatively little is known about the prevalence, mechanisms, and phenotypes of vasculitis occurring in response to ICIs. Here, we review the literature and describe the frequency and patterns of presentation. RECENT FINDINGS: Vasculitis, while infrequent, has been described as an irAE in patients treated with ICI therapy with resultant morbidity and mortality. SUMMARY: Recognizing the risk and management of immune checkpoint inhibitor induced vasculitis in patients with cancer is important in the daily practice of rheumatology.

Immune-Related Adverse Events in Cancer Patients Treated With Immune Checkpoint Inhibitors.

PURPOSE OF REVIEW: With the advent of cancer immunotherapy and immune checkpoint inhibitors, patients with malignancies can now achieve durable remissions for conditions previously described as terminal. However, immune-related adverse events (irAEs) associated with cancer immunotherapy have become an anticipated consequence of enhanced T cell activation. Through an extensive literature review, we assess the most recent clinical and basic research data concerning immune checkpoint blockade and describe the spectrum of associated irAEs as well as their management. RECENT FINDINGS: Anti-PD-1, anti-PD-L1, and anti-CTLA-4 antibodies are widely used in the management of an array of tumors with incredible clinical remissions. However, irAEs cause significant morbidity and mortality and in some cases, result in withdrawal of cancer therapy and initiation of immunosuppression. While this is an exciting time in oncology, irAEs are a barrier to adequate care and therefore deserve close attention and improved capacity to predict and prevent toxicity. Rheumatologists should be familiar with these topics in the eventuality of patient evaluation and management.

Safety and effectiveness of combination versus monotherapy with immune checkpoint inhibitors in patients with preexisting autoimmune diseases.

Patients with preexisting autoimmune disease (pAID) are generally excluded from clinical trials for immune checkpoint inhibitors (ICIs) for cancer due to concern of flaring pAID. In this multi-center, retrospective observational study, we compared safety of ICI combination (two ICI agents) versus monotherapy in cancer patients with pAIDs. The primary outcome was time to AEs (immune-related adverse events (irAEs) and/or pAID flares), with progression-free survival (PFS) and overall survival as secondary outcomes. Sixty-four of 133 patients (48%) received ICI combination and 69 (52%) monotherapy. Most had melanoma (32%) and lung cancer (31%). Most common pAIDs were rheumatic (28%) and dermatologic (23%). Over a median follow-up of 15 months (95%CI, 11-18 mo), the cumulative incidence of any-grade irAEs was higher for combination compared to monotherapy (subdistribution hazard ratio (sHR) 2.27, 95%CI 1.35-3.82). No statistically significant difference was observed in high-grade irAEs (sHR 2.31 (0.95-5.66), P = .054) or the cumulative incidence of pAID flares. There was no statistically significant difference for melanoma PFS between combination versus monotherapy (23.2 vs. 17.1mo, P = .53). The combination group was more likely to discontinue or hold ICI, but > 50% of the combination group was still able to continue ICI therapy. No treatment-related deaths occurred. In our cohort with pAIDs, patients had a tolerable toxicity profile with ICI combination therapy. Our results support the use of ICI combination if deemed necessary for cancer therapy in patients with pAIDs, since the ICI toxicities were comparable to monotherapy, able to be effectively managed and mostly did not require ICI interruption.

Selective immune suppression using interleukin-6 receptor inhibitors for management of immune-related adverse events.

BACKGROUND: Management of immune-related adverse events (irAEs) is important as they cause treatment interruption or discontinuation, more often seen with combination immune checkpoint inhibitor (ICI) therapy. Here, we retrospectively evaluated the safety and effectiveness of anti-interleukin-6 receptor (anti-IL-6R) as therapy for irAEs. METHODS: We performed a retrospective multicenter study evaluating patients diagnosed with de novo irAEs or flare of pre-existing autoimmune disease following ICI and were treated with anti-IL-6R. Our objectives were to assess the improvement of irAEs as well as the overall tumor response rate (ORR) before and after anti-IL-6R treatment. RESULTS: We identified a total of 92 patients who received therapeutic anti-IL-6R antibodies (tocilizumab or sarilumab). Median age was 61 years, 63% were men, 69% received anti-programmed cell death protein-1 (PD-1) antibodies alone, and 26% patients were treated with the combination of anti-cytotoxic T lymphocyte antigen-4 and anti-PD-1 antibodies. Cancer types were primarily melanoma (46%), genitourinary cancer (35%), and lung cancer (8%). Indications for using anti-IL-6R antibodies included inflammatory arthritis (73%), hepatitis/cholangitis (7%), myositis/myocarditis/myasthenia gravis (5%), polymyalgia rheumatica (4%), and one patient each with autoimmune scleroderma, nephritis, colitis, pneumonitis and central nervous system vasculitis. Notably, 88% of patients had received corticosteroids, and 36% received other disease-modifying antirheumatic drugs (DMARDs) as first-line therapies, but without adequate improvement. After initiation of anti-IL-6R (as first-line or post-corticosteroids and DMARDs), 73% of patients showed resolution or change to ≤grade 1 of irAEs after a median of 2.0 months from initiation of anti-IL-6R therapy. Six patients (7%) stopped anti-IL-6R due to adverse events. Of 70 evaluable patients by RECIST (Response Evaluation Criteria in Solid Tumors) V.1.1 criteria; the ORR was 66% prior versus 66% after anti-IL-6R (95% CI, 54% to 77%), with 8% higher complete response rate. Of 34 evaluable patients with melanoma, the ORR was 56% prior and increased to 68% after anti-IL-6R (p=0.04). CONCLUSION: Targeting IL-6R could be an effective approach to treat several irAE types without hindering antitumor immunity. This study supports ongoing clinical trials evaluating the safety and efficacy of tocilizumab (anti-IL-6R antibody) in combination with ICIs (NCT04940299, NCT03999749).

Immune checkpoint inhibition in patients with inactive pre-existing neuromuscular autoimmune diseases.

OBJECTIVES: To evaluate the safety of immune checkpoint inhibitor use in patients with pre-existing neurological autoimmune diseases. METHODS: In this retrospective case-series, we examined exacerbations of underlying disease and the occurrence of immune-related adverse events in 5 patients who had been diagnosed with a neurological autoimmune disease prior to receiving immune checkpoint inhibitor therapy for advanced malignancy. RESULTS: Two patients had a prior diagnosis of myasthenia gravis, two had Guillain-Barré syndrome, and one had chronic idiopathic demyelinating polyneuropathy. Only one patient experienced a flare of neurological autoimmune disease. Four of the five patients experienced immune-related adverse events unrelated to their neurological disease. CONCLUSIONS: In this case-series, exacerbations of neurological autoimmune disease were less common and less severe than expected. Further research is needed to determine which individuals are at greatest risk of neurological autoimmune disease complication while receiving immune checkpoint inhibitor therapy.

Toxicity and Cosmetic Outcome of Breast Irradiation in Women with Breast Cancer and Autoimmune Connective Tissue Disease: The Role of Fraction and Field Size.

PURPOSE: Hypofractionation has historically been underused among breast cancer patients with connective tissue diseases given a theoretical risk of increased toxicity and their overall underrepresentation in clinical trials that established hypofractionation as standard of care. We aim to compare the rates of toxicity in patients with autoimmune connective tissue diseases treated with conventionally fractioned radiation therapy (CF-RT) and hypofractionated radiation therapy (HF-RT) including accelerated partial breast irradiation. METHODS: A total of 1983 patients treated with breast conservation between 2012 and 2016 were reviewed for diagnosis of autoimmune disease. Univariate analysis using binary logistic regression was performed to evaluate the effect of disease and treatment variables on acute and late toxicity. Multivariate analyses using Cox regression models were used to evaluate the independent associations between covariates and the primary end points. Odds ratios (ORs) and 95% confidence intervals (CIs) were calculated for reach risk group. RESULTS: Ninety-two patients with autoimmune disease were identified. Median follow-up was 59 months. Of the patients 35% received CF-RT and 65% received HF-RT, of whom 70% received whole breast radiation (WBI) without regional nodal irradiation, 12% received WBI with regional nodal irradiation, and 18% received accelerated partial breast radiation. Patients who received CF-RT were significantly more likely to have autoimmune disease (AD) symptoms (78% vs 37%, P <.001), to be managed on disease-modifying antirheumatic drugs (DMARDs; 41% vs 15%, P = .013), and to have active autoimmune disease (84% vs 43%, P <.001). On multivariate analysis, HF-RT was associated with a significantly decreased odds of acute and late grade 2/3 toxicity compared with CF-RT fractionation (acute: OR 0.200, 95% CI 0.064-0.622, P = .005; late: OR 0.127, 95% CI 0.031-0.546, P = .005). CONCLUSIONS: Hypofractionation including accelerated partial-breast irradiation is associated with less acute or late grade 2/3 toxicity in this population.

Role of IVIG in the Treatment of Autoimmune Conditions With Concurrent Immune Checkpoint Inhibitors for Metastatic Cancer.

Immune checkpoint inhibitors (ICIs) are a class of medications targeting mostly the PD-1/PD-L1 and CTLA-4 immune pathways in the treatment of many cancers. Despite the encouraging success of ICIs, they are associated with immune-related adverse events as well as exacerbation of underlying autoimmune conditions. The treatment of these conditions often involves discontinuation of ICI in addition to the utilization of immunomodulatory agents. In this report, we discuss a case in which a patient with metastatic renal cell carcinoma experienced exacerbation of underlying paraneoplastic dermatomyositis after treatment with ICI. He was successfully continued on ICI with the use of intravenous immunoglobulin. The patient experienced adequate control of his myositis but also experienced deepening of his antitumor response.

SHP2 Targets ITK Downstream of PD-1 to Inhibit T Cell Function.

PD-1 is a critical therapeutic target in cancer immunotherapy and antibodies blocking PD-1 are approved for multiple types of malignancies. The phosphatase SHP2 is the main effector mediating PD-1 downstream signaling and accordingly attempts have been made to target this enzyme as an alternative approach to treat immunogenic tumors. Unfortunately, small molecule inhibitors of SHP2 do not work as expected, suggesting that the role of SHP2 in T cells is more complex than initially hypothesized. To better understand the perplexing role of SHP2 in T cells, we performed interactome mapping of SAP, an adapter protein that is associated with SHP2 downstream signaling. Using genetic and pharmacological approaches, we discovered that SHP2 dephosphorylates ITK specifically downstream of PD-1 and that this event was associated with PD-1 inhibitory cellular functions. This study suggests that ITK is a unique target in this pathway, and since ITK is a SHP2-dependent specific mediator of PD-1 signaling, the combination of ITK inhibitors with PD-1 blockade may improve upon PD-1 monotherapy in the treatment of cancer.

Breast Conservation in Women with Autoimmune Disease: The Role of Active Autoimmune Disease and Hypofractionation on Acute and Late Toxicity in a Case-Controlled Series.

PURPOSE: Autoimmune connective tissue disease (CTD) has historically represented a relative contraindication to breast conservation (BC) among patients with early-stage breast cancer. Controversy exists regarding the use of hypofractionated radiation therapy (RT) among patients with CTDs. We evaluated acute and late toxicity in patients with breast cancer and CTD treated with BC. METHODS AND MATERIALS: Of 1983 patients treated with BC from 2012 to 2016, we identified 91 patients with an autoimmune disease (AD). Each patient was matched to a control without AD based on age, RT field, and fractionation. RT toxicity and clinician-rated cosmesis were compared between cases and controls. Overall survival, disease-free survival, and local recurrence-free survival were estimated using the Kaplan-Meier method. RESULTS: The median follow-up was 49.9 months for cases and 53.0 months for controls, and 67% of cases and controls were treated with hypofractionated RT. There was no difference in grade 2/3 acute toxicity between cases and controls (26.4% vs. 16.5%, respectively; P = .148). There was a significantly higher rate of grade 2/3 late toxicity among cases (25.8% vs 12.1% among controls; P = .049). Active AD at the time of RT increased the rate of grade 2/3 late toxicity compared with controls (41.7% in cases vs. 11.4% in controls; P = .018). Among patients treated with hypofractionated RT, there was no difference in acute or late grade 2/3 toxicity between cases and controls (acute: 13.1% in cases vs. 11.5% in controls [P > 0.9]; late: 11.9% in cases vs. 13.1% in controls [P > 0.9]). The rates of good/excellent clinician-rated cosmesis were similar between groups (92.9% in cases vs. 98.9% in controls; P = .142). CONCLUSIONS: In the largest matched case-control study of patients with CTD treated with conventional and hypofractionated RT, we demonstrate low rates of radiation toxicity, with good to excellent clinician-rated cosmesis. There was increased late toxicity in cases, especially in patients with active AD at time of RT. There was no increase in acute or late toxicity in the patients treated with hypofractionation.

Immunotherapy to treat malignancy in patients with pre-existing autoimmunity.

In the past 10 years, immune checkpoint inhibitors (ICIs) have become an additional pillar of cancer therapy by activating the immune system to treat a number of different malignancies. Many patients receiving ICIs develop immune-related adverse events (irAEs) that mimic some features of classical autoimmune diseases. Unfortunately, patients with underlying autoimmune conditions, many of whom have an increased risk for malignancy, have been excluded from clinical trials of ICIs due to a concern that they will have an increased risk of irAEs. Retrospective data from patients with autoimmune diseases and concomitant malignancy treated with ICIs are encouraging and suggest that ICIs may be tolerated safely in patients with specific autoimmune diseases, but there are no prospective data to guide management. In this manuscript, we review the relationship between pre-existing autoimmune disease and irAEs from checkpoint inhibitors. In addition, we assess the likelihood of autoimmune disease exacerbations in patients with pre-existing autoimmunity receiving ICI.

Borrelia burgdorferi vlsE antigenic variation is not mediated by RecA.

RecA is a key protein linking genetic recombination to DNA replication and repair in bacteria. Previous functional characterization of Borrelia burgdorferi RecA indicated that the protein is mainly involved in genetic recombination rather than DNA repair. Genetic recombination may play a role in B. burgdorferi persistence by generation of antigenic variation. We report here the isolation of a recA null mutant in an infectious B. burgdorferi strain. Comparison of the in vitro growth characteristics of the mutant with those of the wild-type strain under various conditions showed no significant differences. While the RecA mutant was moderately more sensitive to UV irradiation and mitomycin C than the wild-type strain, the lack of RecA abolished allelic exchange in the mutant. Absence of RecA did not affect the ability of the mutant to infect mice. However, the RecA mutant was attenuated for joint infection in competitive-infection assays with the wild-type strain. vlsE sequence variation in mice was observed in both wild-type and RecA mutant spirochetes, indicating that the mechanism of antigenic variation is not homologous genetic recombination.

Borrelia burgdorferi sensu stricto is clonal in patients with early Lyme borreliosis.

Lyme borreliosis, the most commonly reported vector-borne disease in North America, is caused by the spirochete Borrelia burgdorferi. Given the extensive genetic polymorphism of B. burgdorferi, elucidation of the population genetic structure of the bacterium in clinical samples may be relevant for understanding disease pathogenesis and may have applicability for the development of diagnostic tests and vaccine preparations. In this investigation, the genetic polymorphism of the 16S-23S rRNA (rrs-rrlA) intergenic spacer and ospC was investigated at the sequence level in 127 clinical isolates obtained from patients with early Lyme borreliosis evaluated in suburban New York City. Sixteen distinct rrs-rrlA and 16 distinct ospC alleles were identified, representing virtually all of the genotypes previously found in questing Ixodes scapularis nymphs in this region. In addition, a new ospC group was identified in a single patient. The strong linkage observed between the chromosome-located rrs-rrlA and plasmid-borne ospC genes suggests a clonal structure of B. burgdorferi in these isolates, despite evidence of recombination at ospC.

Targeting the programmed cell death-1 pathway in rheumatoid arthritis.

Since the introduction of TNF-α inhibitors and other biologic agents, the clinical outcome for many treated rheumatoid arthritis patients has significantly improved. However, there are still a substantial proportion of patients that are intolerant, or have inadequate responses, with current agents that have become the standards of care. While the majority of these agents are designed to affect the inflammatory features of the disease, there are also agents in the clinic that instead target lymphocyte subsets (e.g., rituximab) or interfere with lymphocyte co-receptor signaling pathways (e.g., abatacept). Due in part to their ability to orchestrate downstream inflammatory responses that lead to joint damage and disease progression, pathogenic expansions of T and B lymphocytes are appreciated to play key roles in the pathogenesis of rheumatoid arthritis. New insights into immune regulation have suggested novel approaches for the pharmacotherapeutic targeting of lymphocytes. In this review, we discuss deepening insights into human genetics and our understanding of the interface with rheumatoid arthritis pathogenesis providing a strong rationale for exploiting the co-inhibitory receptor programmed cell death-1 signaling pathway as a better approach for the treatment of this chronic, often progressive destructive joint disease.

The Potential Role of Inhibitory Receptors in the Treatment of Psoriasis.

Psoriasis is a common autoimmune disorder that affects the skin. Approximately 30% of individuals with psoriasis will develop inflammatory arthritis, often in the setting of human leukocyte antigen B27. Both forms of disease are thought to be the result of prolonged inflammation mediated by T lymphocytes, dendritic cells, and keratinocytes. While there are treatments aimed at immunomodulation, targeting T cell co-inhibitory receptors signaling pathways may provide therapeutic benefit. This review will discuss in detail four T cell co-inhibitory receptors and their potential application for the treatment of psoriasis and psoriatic arthritis.

Multiple uracil-DNA glycosylase activities in Deinococcus radiodurans.

The extremely radiation resistant bacterium, Deinococcus radiodurans, contains a spectrum of genes that encode for multiple activities that repair DNA damage. We have cloned and expressed the product of three predicted uracil-DNA glycosylases to determine their biochemical function. DR0689 is a homologue of the Escherichia coli uracil-DNA glycosylase, the product of the ung gene; this activity is able to remove uracil from a U : G and U : A base pair in double-stranded DNA and uracil from single-stranded DNA and is inhibited by the Ugi peptide. DR1751 is a member of the class 4 family of uracil-DNA glycosylases such as those found in the thermophiles Thermotoga maritima and Archaeoglobus fulgidus. DR1751 is also able to remove uracil from a U : G and U : A base pair; however, it is considerably more active on single-stranded DNA. Unlike its thermophilic relatives, the enzyme is not heat stable. Another putative enzyme, DR0022, did not demonstrate any appreciable uracil-DNA glycosylase activity. DR0689 appears to be the major activity in the organism based on inhibition studies with D. radiodurans crude cell extracts utilizing the Ugi peptide. The implications for D. radiodurans having multiple uracil-DNA glycosylase activities and other possible roles for these enzymes are discussed.

The propensity of different Borrelia burgdorferi sensu stricto genotypes to cause disseminated infections in humans.

Lineages of Borrelia burgdorferi, the bacterium that causes Lyme disease, can be characterized by distinct alleles at the outer surface protein C (ospC) locus. The lineages marked by ospC genotypes have been shown to be differentially invasive in different species of mammals, including humans; genotypes A, B, I, and K effectively disseminate to human blood and cerebrospinal fluid. In this report, we extend the sample of genotypes isolated from human blood to include genotypes N, H, C, M, and D, and rank each by their probability of disseminating from ticks to the blood of humans. Our results demonstrate that only some genotypes of B. burgdorferi present in ticks have a high propensity to disseminate in humans.