KMT2D regulates activation, localization, and integrin expression by T-cells.

Individuals with Kabuki syndrome present with immunodeficiency; however, how pathogenic variants in the gene encoding the histone-modifying enzyme lysine methyltransferase 2D (KMT2D) lead to immune alterations remain poorly understood. Following up on our prior report of KMT2D-altered integrin expression in B-cells, we performed targeted analyses of KMT2D's influence on integrin expression in T-cells throughout development (thymocytes through peripheral T-cells) in murine cells with constitutive- and conditional-targeted Kmt2d deletion. Using high-throughput RNA-sequencing and flow cytometry, we reveal decreased expression (both at the transcriptional and translational levels) of a cluster of leukocyte-specific integrins, which perturb aspects of T-cell activation, maturation, adhesion/localization, and effector function. H3K4me3 ChIP-PCR suggests that these evolutionary similar integrins are under direct control of KMT2D. KMT2D loss also alters multiple downstream programming/signaling pathways, including integrin-based localization, which can influence T-cell populations. We further demonstrated that KMT2D deficiency is associated with the accumulation of murine CD8+ single-positive (SP) thymocytes and shifts in both human and murine peripheral T-cell populations, including the reduction of the CD4+ recent thymic emigrant (RTE) population. Together, these data show that the targeted loss of Kmt2d in the T-cell lineage recapitulates several distinct features of Kabuki syndrome-associated immune deficiency and implicates epigenetic mechanisms in the regulation of integrin signaling.

MET receptor serves as a promising target in melanoma brain metastases.

The development of brain metastases hallmarks disease progression in 20-40% of melanoma patients and is a serious obstacle to therapy. Understanding the processes involved in the development and maintenance of melanoma brain metastases (MBM) is critical for the discovery of novel therapeutic strategies. Here, we generated transcriptome and methylome profiles of MBM showing high or low abundance of infiltrated Iba1high tumor-associated microglia and macrophages (TAMs). Our survey identified potential prognostic markers of favorable disease course and response to immune checkpoint inhibitor (ICi) therapy, among them APBB1IP and the interferon-responsive gene ITGB7. In MBM with high ITGB7/APBB1IP levels, the accumulation of TAMs correlated significantly with the immune score. Signature-based deconvolution of MBM via single sample GSEA revealed enrichment of interferon-response and immune signatures and revealed inflammation, stress and MET receptor signaling. MET receptor phosphorylation/activation maybe elicited by inflammatory processes in brain metastatic melanoma cells via stroma cell-released HGF. We found phospho-METY1234/1235 in a subset of MBM and observed a marked response of brain metastasis-derived cell lines (BMCs) that lacked druggable BRAF mutations or developed resistance to BRAF inhibitors (BRAFi) in vivo to MET inhibitors PHA-665752 and ARQ197 (tivantinib). In summary, the activation of MET receptor in brain colonizing melanoma cells by stromal cell-released HGF may promote tumor self-maintenance and expansion and might counteract ICi therapy. Therefore, therapeutic targeting of MET possibly serves as a promising strategy to control intracranial progressive disease and improve patient survival.

Expression of integrin ß-7 is epigenetically enhanced in multiple myeloma subgroups with high-risk cytogenetics.

BACKGROUND: Oncogenic overexpression of integrin-ß7 (ITGB7) in cases of high-risk multiple myeloma (MM) was reported to promote enhanced interactions between neoplastic plasma-B cells and stromal cells to develop cell-adhesion mediated drug resistance. METHODS: Expression profiles of adhesion related genes were analyzed in a cohort of MM patients containing major IgH translocations or hyperdiploidies (HY), diagnosed at the premalignant monoclonal gammopathy of undetermined significance (MGUS; n = 103), smoldering multiple myeloma; (SMM; n = 190) or MM (MM; n = 53) stage. Differential expression was integrated with loci-specific alterations in DNA-methylation and chromatin marks in MM patients. A CRISPR-based targeted induction of DNA-methylation at the ITGB7 super-enhancer (SE) in MM.1S cells was employed to intersect the impact of cis-regulatory elements on ITGB7 expression. RESULTS: ITGB7 was significantly (p < 0.05) upregulated in patients with t(14;16) and t(14;20) subgroups in all MGUS, SMM and MM stages, but sporadically upregulated in t(4;14) subgroup at the MM stage. We demonstrate a predetermined enhancer state on ITGB7 in primary-B cells that is maintained under bivalent chromatin, which undergoes a process of chromatin-state alterations and develops into an active enhancer in cases of the t(4;14) subgroup or SE in cases of the t(14;16) subgroup. We also demonstrate that while targeted induction of DNA-methylation at the ITGB7-SE further upregulated the gene, inhibition of ITGB7-SE-associated transcription factor bromodomain-4 downregulated expression of the gene. CONCLUSIONS: Our findings suggest an epigenetic regulation of oncogenic overexpression of ITGB7 in MM cells, which could be critical in MM progression and an attractive therapeutic target.

EccDNA-oriented ITGB7 expression in breast cancer.

Background: Extrachromosomal circular DNA (eccDNA) is omnipresent in cancers and related to the progression of tumors and oncogene amplification. However, its function in breast cancer (BC) is unclear. Methods: After constructing the DNA library, CLeavage Effects by Circularization for In vitro Reporting of sequencing was performed for eccDNA detection using 1 BC tissue sample. Fastqc was used to evaluate the quality of the original data. Burrows-Wheeler-Alignment Tool was used to compare the original data to the reference genome. A Circle-MAP was subsequently performed to detect eccDNA, and Bedtools was used to annotate the eccDNA genes. Gene Ontology and Kyoto Encyclopedia of Genes and Genomes enrichment analyses were conducted by ClusterProfiler. The Genotype-Tissue Expression and the Cancer Genome Atlas databases were used to collect the ribonucleic acid-sequencing data of the BC and normal samples. A Gene Expression Profiling Interactive Analysis, the University of Alabama at Birmingham CANcer data analysis Portal, and Kaplan-Meier survival curves were used to analyze the Cancer Genome Atlas data. Results: A total of 200 eccDNA genes, including IGTB7, were obtained. About the biological processes (BPs), these 200 genes were mainly enriched in actin cytoskeleton reorganization and axon guidance. Concerning the molecular functions (MFs), these 200 genes were mainly enriched in sodium ion transmembrane transporter activity and metal ion transmembrane transporter activity. As for cellular components (CCs), these 200 genes were mainly enriched in the transcription regulator complex and focal adhesion. ITGB7 was significantly enriched in cell-matrix adhesion and localization within the membrane in the BPs, integrin binding in the MFs, and cell-substrate junction and focal adhesion in the CCs. The 200 eccDNA genes were mainly enriched in the PI3K-Akt signaling pathway and focal adhesion. Notably, ITGB7 was enriched in focal adhesion, ECM-receptor interaction, the PI3K-Akt signaling pathway, and human papillomavirus infection. Besides, ITGB7 was significantly upregulated in BC patients and was associated with the menopause status of the BC patients. Conclusions: ITGB7 might serve as a prognostic marker for BC patients. ITGB7 has important implications for the individualized clinical treatment of BC patients.

Circulating MAdCAM-1 and ITGB7 in Patients with Plaque Psoriasis and Eruptive Lichen Planus-Preliminary Data.

Plaque psoriasis (PSO) and lichen planus (LP) are skin diseases with some similarities in pathogenesis, comorbidities, and clinical presentation. Mucosal addressin cell adhesion molecule-1 (MAdCAM-1) and its ligand, α4ß7 integrin, are involved in inflammatory bowel diseases and liver dysfunctions, which occur more frequently in PSO and LP. Serum MAdCAM-1 and ITGB7 levels in patients with plaque PSO and eruptive LP have never been studied before. The study included 42 patients with PSO, 13 with LP, and 23 controls. Serum molecules levels were evaluated using the immune-enzymatic method. ITGB7 concentration was not statistically different, both in patients with PSO and LP, compared to controls (both p > 0.05). MAdCAM-1 level was significantly lower in PSO subjects than in controls (p = 0.041), whereas in the LP group, a downward trend was observed (p = 0.088) with p = 0.0455 in ANOVA. Multiple linear regression revealed independent associations between ITGB7 and HDL and BMI and RBC in the LP group. In psoriatic patients with elevated CRP, there was an upward trend for MAdCAM-1, and also a positive correlation between MAdCAM-1 and WBC. ITGB7 and MAdCAM-1 cannot serve as markers of disease activity or liver pathology neither in patients with PSO nor LP. MAdCAM-1 might play a role as an inflammation indicator in PSO and a beneficial influence on the lipid profile in LP.

Genetics of fulminant type 1 diabetes.

Since fulminant type 1 diabetes was reported as a distinct subtype of type 1 diabetes in 2000, the Committee on Type 1 diabetes, Japan Diabetes Society has continuously recruited patients and conducted genomic research to elucidate the genetic basis of fulminant type 1 diabetes. The contribution of the human leukocyte antigen complex (HLA) to genetic susceptibility to fulminant type 1 diabetes was compared with that of other subtypes in 2009. The alleles and haplotypes associated with fulminant type 1 diabetes were found to be different from acute-onset and slowly progressive type 1 diabetes. DRB1*15:01-DQB1*06:02, a protective haplotype against acute-onset type 1 diabetes, does not provide protection against fulminant type 1 diabetes and DRB1*08:02-DQB1*03:02, a susceptible haplotype to acute-onset type 1 diabetes, does not confer susceptibility to fulminant type 1 diabetes. Recently, the first genome-wide association study (GWAS) of fulminant type 1 diabetes was performed in Japanese individuals. A strong association was observed with multiple single nucleotide polymorphisms (SNPs) in the HLA region, and the strongest association was observed with rs9268853 in the class II DR region. In addition, 11 SNPs outside the HLA region showed some evidence of association with the disease. In particular, rs11170445 in CSAD/lnc-ITGB7-1 on chromosome 12q13.13 showed an association at a genome-wide significance level. Fine mapping revealed that rs3782151 in CSAD/lnc-ITGB7-1 showed the lowest P value. CSAD/lnc-ITGB7-1 was found to be strongly associated with susceptibility to fulminant, but not classical, autoimmune type 1 diabetes, implicating this locus in the distinct phenotype of fulminant type 1 diabetes.

Abnormal Peyer patch development and B-cell gut homing drive IgA deficiency in Kabuki syndrome.

BACKGROUND: Kabuki syndrome (KS) is commonly caused by mutations in the histone-modifying enzyme lysine methyltransferase 2D (KMT2D). Immune dysfunction is frequently observed in individuals with KS, but the role of KMT2D in immune system function has not been identified. OBJECTIVE: We sought to understand the mechanisms driving KS-associated immune deficiency (hypogammaglobulinemia [low IgA], splenomegaly, and diminished immunization responses). METHODS: We performed a comprehensive evaluation of humoral immunity and secondary lymphoid tissues in an established KS (Kmt2d+/ßGeo) mouse model and validated select findings in a patient with KS. RESULTS: Compared with wild-type littermates, Kmt2d+/ßGeo mice demonstrated deficiencies in multiple B-cell lineages and reduced serum IgA and elevated IgM levels across multiple ages. The bone marrow, spleen, and intestine of Kmt2d+/ßGeo mice contained diminished numbers of IgA-secreting cells, while elevated germinal center B cells were found in the mesenteric lymph node and Peyer patches. Kmt2d+/ßGeo mice have decreased size and numbers of Peyer patches, a finding confirmed in human samples. We identified deficiency of Itgb7 RNA and protein expression, a gene encoding an adhesion protein that mediates intestinal homing, and we demonstrated KMT2D-dependent control of ITGB7 expression in a human cell line. CONCLUSIONS: Kmt2d haploinsufficiency has broad deleterious effects on B-cell differentiation, specifically hampering gut lymphocyte homing and IgA+ plasma cell differentiation. Intestinal lymphoid defects caused by ITGB7 deficiency have not previously been recognized in KS, and these results provide new mechanistic insights into the pathogenesis of KS-associated immune deficiency.