Background Immune checkpoint inhibitors (ICI) have revolutionized the treatment landscape of advanced malignancies, but come with a diverse spectrum of immune-related adverse events (irAEs). Studies into irAE mechanisms are needed to make a transition from expert-opinion to evidence-based irAE treatment strategies. Methods We aimed to longitudinally characterize peripheral blood T and B cell dynamics in ICI-treated patients developing irAEs and remaining irAE-free. PBMCs were immunophenotyped and functionally assessed with multicolor flow cytometry at baseline, after ±3 weeks and ±6 weeks or upon clinically relevant irAEs. Additionally, serum cytokine concentrations for 23 analytes were measured by multiplex immunoassay at the same timepoints. Results We analyzed samples from 44 ICI-treated patients (24 anti-PD-1 monotherapy, 20 combined anti-PD-1 and anti-CTLA-4; cICI), of whom 22 developed clinically relevant irAEs, and 10 healthy donors. IrAEs after cICI were characterized by significantly enhanced proliferation of Th1-associated, mainly effector memory T cells, as well as Th17-associated and possibly antibody-mediated immune responses. The latter response was reflected in cICI toxicity by rising CXCL13 and IL-21 levels, but without changes in CD21lo, memory, class-switched or newly activated B cell subsets. Anti-PD-1 monotherapy-inflicted irAEs were associated with a modestly enhanced Th1-associated response compared to irAE-free patients, only reflected by increasing serum CXCL9 and CXCL10, but without cellular changes in Th1/Tc1 subsets. PD-1+LAG-3+double-positive (DP) CD8+T cells retained their proinflammatory potential in all treatment groups, regardless of irAE development. Especially DP CD8+T cells showed enhanced cytotoxic capacity in patients with irAEs after cICI. In all patients with irAEs, highly cytotoxic CD57+CD8+T cells were abundant. Conclusions Peripheral blood immune responses are substantially different between cICI and anti-PD-1 monotherapy-treated patients. ICI-induced toxicity is clearly dominated by an enhanced Th1-associated response, but in cICI we also found evidence for Th17-associated and possibly antibody-mediated responses. CD4+effector memory T cells were the principal cycling cells in irAEs after cICI. Together, our data add to the growing body of evidence that irAEs may be antibody-mediated and driven by newly activated CD4+helper T cells, specifically after cICI.

Immune checkpoint inhibition (ICI) can induce durable responses in patients with advanced malignancies. Three cases of hematological neoplasia following ICI for solid tumors have been reported to date. We present five patients treated at our tertiary referral center between 2017 and 2021 who developed chronic myeloid leukemia (two patients), acute myeloid leukemia, myelodysplastic syndrome and chronic eosinophilic leukemia during or after anti-PD-1-based treatment. Molecular analyses were performed on pre-ICI samples to identify baseline variants in myeloid genes. We hypothesize that PD-1 blockade might accelerate progression to overt myeloid malignancies and discuss potential underlying mechanisms.

Immune checkpoint inhibitors (ICIs) have changed perspectives for patients with cancer, but come with severe immune-related adverse events (irAEs). To prevent fatality or chronicity, these irAEs are often promptly treated with high-dose immunosuppressants. Until recently, evidence on the effects of irAE management on ICI efficacy has been sparse. As a result, algorithms for irAE management are mostly expert-opinion based and barely consider possible detrimental effects of immunosuppressants on ICI efficacy. However, recent growing evidence suggests that vigorous immunosuppressive management of irAEs comes with unfavourable effects on ICI efficacy and survival. With expansion of the indications of ICIs, evidence-based treatment of irAEs without hampering tumour control becomes more and more important. In this review, we discuss novel evidence from pre-clinical and clinical studies on the effects of different irAE management regimens including corticosteroids, TNF inhibition and tocilizumab on cancer control and survival. We provide recommendations for pre-clinical research, cohort studies and clinical trials that can help clinicians in tailored irAE management, minimising patients’ burden while maintaining ICI efficacy.

Since the mid-1980s magnetic resonance imaging (MRI) has been investigated as a non- or minimally invasive tool to probe kidney allograft function. Despite this long-standing interest, MRI still plays a subordinate role in daily practice of transplantation nephrology. With the introduction of new functional MRI techniques, administration of exogenous gadolinium-based contrast agents has often become unnecessary and true non-invasive assessment of allograft function has become possible. This raises the question why application of MRI in the follow-up of kidney transplantation remains restricted, despite promising results. Current literature on kidney allograft MRI is mainly focused on assessment of (sub) acute kidney injury after transplantation. The aim of this review is to survey whether MRI can provide valuable diagnostic information beyond 1 year after kidney transplantation from a mechanistic point of view. The driving force behind chronic allograft nephropathy is believed to be chronic hypoxia. Based on this, techniques that visualize kidney perfusion and oxygenation, scarring, and parenchymal inflammation deserve special interest. We propose that functional MRI mechanistically provides tools for diagnostic work-up in long-term follow-up of kidney allografts.

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Introduction Within the general population over 50 years, 3% is affected with a monoclonal gammopathy of undetermined significance (MGUS). Although MGUS generally demands no treatment, interest in treating premalignant stages to prevent progression to overt malignancies has grown. Better insight in processes underlying progression of MGUS is therefore warranted. Platelet RNA profiles have been shown to reflect information on the topical tumor environment in various cancer types, including multiple myeloma (MM). Our aim was to investigate whether IgM MGUS patients can be distinguished from healthy controls (HCs) using platelet RNA splicing profiles, and to relate IgM MGUS splicing profiles to genes associated with Waldenström's Macroglobulinemia (WM). Methods Twenty-four IgM MGUS patients with presence of a paraprotein confirmed through immunofixation consented to participation. The thromboSeq protocol for platelet RNA sequencing (RNA-Seq) based classification (Best et al. Nat Protoc 2019) was used to acquire platelet RNA profiles from all MGUS subjects. RNA was extracted from platelets isolated from whole blood. After reverse transcription and SMARTer amplification, 100 bp single-read sequencing was performed on the Illumina Hiseq 2500 platform. Raw sequencing data were processed with Trimmomatic, aligned to a reference genome with STAR and mapped with HTSeq. Mapping data of 29 age and blood storage time matched HCs were also included in the analysis. Differential expression of spliced RNAs expressed as log2 fold change (logFC) was assessed with a likelihood-ratio ANOVA and visualized in a heatmap constructed with differentially spliced RNAs that passed the particle swarm optimized (PSO) false discovery rate (FDR) threshold. RNA profiles were further explored and related to WM and MM by DAVID gene ontology enrichment analyses. Results Mean age in the MGUS and HC groups was 67.2 (SD 7.7) and 66.9 (SD 6.7) years. Thirteen of 24 MGUS patients suffered from IgM mediated anti-myelin associated glycoprotein (MAG) neuropathy; the other 11 were asymptomatic. Of 3,426 high-abundant spliced RNAs detected, 1,371 demonstrated significant differential expression (FDR <0.05) and allowed perfect distinction of MGUS from HCs after PSO (Fig. 1; P <0.0001). The two distinct MGUS phenotypes (neuropathy vs. asymptomatic) could not be distinguished. Top 50 overexpressed RNA isoforms were mainly involved in apoptotic processes, transcription regulation, cellular defense response and immune response regulation. WM associated genes (logFC in parentheses) BCL7C (1.35), CASP8 (1.75), CD79A (2.65), CD81 (1.58), DUSP1 (2.32), HLA-DRB1 (1.83), JAK3 (0.65), NFKBIA (3.59), NFKB2 (1.63), REL (1.35) and TNFRSF14 (0.75), as well as MM associated JUND (4.61) and CCNL1 (3.11), were overexpressed in MGUS. TNFAIP3, negatively regulating the NF-κB pathway, was highly overexpressed (logFC 4.07). JAK1 and JAK2 mRNAs were mildly though significantly depleted in the MGUS group. We also found overexpression of CCNL2 (1.81), MS4A1 (2.17) and WNK1 (0.79), which have been shown to be enriched in WM relative to IgM MGUS. Top 50 depleted RNAs were mainly involved in protein phosphorylation, intracellular signal transduction and response to tumor necrosis factor. CASP3 (-1.43), CASP4 (-0.62) and TNFSF4 (-0.99) mRNAs, of which overexpression is associated with WM, were depleted in MGUS. Conclusions For the first time, we have demonstrated widespread differential expression of spliced RNAs in platelets of MGUS patients as compared to healthy controls. We found evidence of overexpression of various genes that have been associated with WM. The differences were mainly in pathways related to apoptosis, signaling and immune response regulation. This is in line with findings that balanced elevation of free light chains, possibly due to a chronic inflammatory state, increases risk of developing a monoclonal gammopathy (Kumar et al. Blood Cancer J 2019). Overexpression of NFKBIA, NFKB2 andTNFAIP3 in MGUS possiblysuggests activation of the NF-κB pathway, a key step in MYD88 mediated progression to WM. JAK subtype expression levels may reflect that the JAK/STAT pathway, also important for progression to WM, has not (yet) been aberrantly activated in our MGUS subjects. Future research could be focused on studying the development of MGUS and MGUS progression to WM and MM using differentially expressed splice junctions in platelets. Disclosures Minnema: Amgen: Honoraria; Celgene Corporation: Honoraria, Research Funding; Gilead: Honoraria; Jansen Cilag: Honoraria; Servier: Honoraria.

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Colitis is a prevalent adverse event associated with immune checkpoint inhibitor (ICI) therapy with similarities to inflammatory bowel disease. Incomplete mechanistic understanding of ICI-colitis curtails evidence-based treatment. Given the often-overlooked connection between tissue architecture and mucosal immune cell function, we here applied imaging mass cytometry (IMC) to gain spatial proteomic insight in ICI-colitis in comparison to ulcerative colitis (UC). Using a cell segmentation pipeline that simultaneously utilizes high-resolution nuclear imaging and high-multiplexity IMC, we show that CD8+T cells are significantly more abundant (and dominant) in anti-PD-1 +/- anti-CTLA-4-induced colitis compared to anti-CTLA-4-induced colitis and UC. We identified activated, cycling CD8+tissue-resident memory T (TRM) cells at the lamina propria-epithelial interface as drivers of cytotoxicity in ICI-colitis and UC. Moreover, we found that combined ICI-induced colitis featured highest granzyme B levels both in tissue and serum. Together, these data reinforce CD8+TRMcells as potentially targetable drivers of ICI-colitis.