Identification of a Six-Gene SLC Family Signature With Prognostic Value in Patients With Lung Adenocarcinoma

Given the importance of solute carrier (SLC) proteins in maintaining cellular metabolic homeostasis and that their dysregulation contributes to cancer progression, here we constructed a robust SLC family signature for lung adenocarcinoma (LUAD) patient stratification. Transcriptomic profiles and relevant clinical information of LUAD patients were downloaded from the TCGA and GEO databases. SLC family genes differentially expressed between LUAD tissues and adjacent normal tissues were identified using limma in R. Of these, prognosis-related SLC family genes were further screened out and used to construct a novel SLC family-based signature in the training cohort. The accuracy of the prognostic signature was assessed in the testing cohort, the entire cohort, and the external GSE72094 cohort. Correlations between the prognostic signature and the tumor immune microenvironment and immune cell infiltrates were further explored. We found that seventy percent of SLC family genes (279/397) were differentially expressed between LUAC tissues and adjacent normal. Twenty-six genes with p-values < 0.05 in univariate Cox regression analysis and Kaplan-Meier survival analysis were regarded as prognosis-related SLC family genes, six of which were used to construct a prognostic signature for patient classification into high- and low-risk groups. Kaplan-Meier survival analysis in all internal and external cohorts revealed a better overall survival for patients in the low-risk group than those in the high-risk group. Univariate and multivariate Cox regression analyses indicated that the derived risk score was an independent prognostic factor for LUAD patients. Moreover, a nomogram based on the six-gene signature and clinicopathological factors was developed for clinical application. High-risk patients had lower stromal, immune, and ESTIMATE scores and higher tumor purities than those in the low-risk group. The proportions of infiltrating naive CD4 T cells, activated memory CD4 T cells, M0 macrophages, resting dendritic cells, resting mast cells, activated mast cells, and eosinophils were significantly different between the high- and low-risk prognostic groups. In all, the six-gene SLC family signature is of satisfactory accuracy and generalizability for predicting overall survival in patients with LUAD. Furthermore, this prognostics signature is related to tumor immune status and distinct immune cell infiltrates in the tumor microenvironment.

IMMU-38. DEEP IMMUNOPROFILING OF HUMAN GLIOBLASTOMA (GBM) REVEALS DIFFERENCES IN THE TUMOR IMMUNE CELL INFILTRATE IN PATIENTS WITH HIGH VS. LOW PLASMA CELL-FREE DNA (CFDNA)

BACKGROUND We have previously demonstrated that high baseline plasma cfDNA concentration is associated with poor survival in patients with newly diagnosed GBM. The mechanism of this association remains unknown. To explore whether differences in the immune landscape between high- vs. low-cfDNA patients may play a role in their divergent clinical outcomes, we phenotyped tumors from patients with high vs. low cfDNA using mass cytometry by time of flight (CyTOF). METHODS We performed CyTOF on frozen tumor infiltrate suspension from a pilot cohort of patients with previously untreated GBM with known baseline plasma cfDNA concentration (Bagley, Clin Cancer Res 2020). CyTOF was used to simultaneously measure expression of 39 molecules related to immune cell lineage, differentiation state, and function. Differences in immune cell infiltrates between high- and low-cfDNA patients were assessed using Mann-Whitney U tests. RESULTS Four patients with high cfDNA (median 57, range 33-90 ng/mL) were compared to six patients with low cfDNA (median 12, range 7-16 ng/mL). Immune cell infiltrates with increased adaptive cells (high monocytes and T cells, p=0.05) were present in high-cfDNA compared to low-cfDNA patients. While > 70% of the infiltrating T cells were exhausted in both groups, the pattern of exhaustion was significantly different in high- vs. low-cfDNA patients, with less CXCR5+CD69+ and more CXCR5-CD69- (p=0.008) progenitor exhausted T cells in cfDNA-high patients. CONCLUSIONS In this GBM pilot study, we demonstrated differences in the tumor immune infiltrate in patients with high vs. low baseline plasma cfDNA concentration. Preclinical studies will be needed to determine if this explains the association between high plasma cfDNA and poor outcomes previously observed in patients. Our results may have implications for the use of cfDNA concentration as a predictive biomarker for immunotherapy, as tumors with more intermediate progenitor (CXCR5-CD69-) exhausted T cells may respond better to PD-1 checkpoint blockade.

Comparative Histological Subtyping of Immune Cell Infiltrates in MPO-ANCA and PR3-ANCA Glomerulonephritis

BackgroundAcute kidney injury (AKI) is a common and severe complication of anti-neutrophil cytoplasmic antibodies (ANCA)-associated vasculitis (AAV), potentially leading to chronic kidney disease (CKD), end-stage renal disease (ESRD), or death. Pathogenic ANCAs, in particular proteinase 3 (PR3) and myeloperoxidase (MPO), trigger a deleterious immune response with intrarenal immune cell infiltration resulting in a pauci-immune necrotizing and crescentic glomerulonephritis (GN). However, a systematic analysis of intrarenal immune cell subtypes concerning neutrophils, eosinophils, plasma cells, and mononuclear cell infiltrates (macrophages, lymphocytes) in ANCA GN remains elusive. Therefore, we aimed to compare distinct immune cell infiltrates in association with clinicopathological findings in ANCA GN.MethodsA total of 53 kidney biopsies with ANCA GN at the University Medical Center Göttingen were retrospectively analyzed. Histological infiltrates of neutrophils, eosinophils, plasma cells, and mononucleated cells (macrophages, lymphocytes) were quantified as a fraction of the total area of inflammation.ResultsNeutrophilic infiltrates were associated with glomerular necrosis and severe kidney injury in ANCA GN. Among tubulointerstitial lesions, intrarenal neutrophils correlated with interstitial inflammation, tubulitis, and inflammation in areas of interstitial fibrosis/tubular atrophy (IFTA), representing active inflammatory lesions. Concerning eosinophils, infiltrates were associated with severe kidney injury, interstitial inflammation, and cellular casts independent of glomerular lesions, implicating a distinct role in inflammation and damage in ANCA GN. Plasma cell infiltrates correlated with tubulitis and interstitial fibrosis and were associated with renal replacement therapy during the short-term disease course. Finally, mononuclear cell infiltrates correlated with severe kidney injury and active histopathological lesions (glomerular crescents, interstitial inflammation, tubulitis, inflammation, and tubulitis in areas of IFTA) besides chronic lesions (interstitial fibrosis and tubular atrophy) in ANCA GN. Interestingly, intrarenal subtypes of immune cell infiltrates differed in MPO-ANCA versus PR3-ANCA GN and were associated with distinct glomerular and tubulointerstitial lesions, implicating different pathogenic mechanisms of kidney injury in ANCA subtypes.ConclusionOur observations imply distinct pathomechanisms contributing to inflammation and renal injury in MPO vs. PR3-associated ANCA GN and potentially contribute to new therapeutic targets in specific ANCA subtypes.

Deconvolution of sarcoma methylomes reveals varying degrees of immune cell infiltrates with association to genomic aberrations

Background Soft-tissue sarcomas (STS) are a heterogeneous group of mesenchymal tumors for which response to immunotherapies is not well established. Therefore, it is important to risk-stratify and identify STS patients who will most likely benefit from these treatments. Results To reveal shared and distinct methylation signatures present in STS, we performed unsupervised deconvolution of DNA methylation data from the TCGA sarcoma and an independent validation cohort. We showed that leiomyosarcoma can be subclassified into three distinct methylation groups. More importantly, we identified a component associated with tumor-infiltrating leukocytes, which suggests varying degrees of immune cell infiltration in STS subtypes and an association with prognosis. We further investigated the genomic alterations that may influence tumor infiltration by leukocytes including RB1 loss in undifferentiated pleomorphic sarcomas and ELK3 amplification in dedifferentiated liposarcomas. Conclusions In summary, we have leveraged unsupervised methylation-based deconvolution to characterize the immune compartment and molecularly stratify subtypes in STS, which may benefit precision medicine in the future.

Shaoyao Decoction Inhibits Inflammation and Improves Intestinal Barrier Function in Mice With Dextran Sulfate Sodium-Induced Colitis

Shaoyao decoction (SYD), a classical traditional Chinese medicine formula, is effective for the treatment of inflammatory bowel disease (IBD). This study was designed to investigate the therapeutic effects of SYD on IBD and possible mechanisms. Dextran sulfate sodium (DSS, 3.5%) was used to induce colitis in C57BL/6 mice. Disease phenotypes were investigated based on disease activity index (DAI), colon length, and microscopic and macroscopic scores. Additionally, the presence of proinflammatory cytokines, immune cell infiltrates, intestinal cell proliferation, apoptosis, epithelial permeability, signal transducer and activator of transcription 3 (STAT3), and nuclear factor-κB (NF-κB) signaling, as well as the intestinal mucosal barrier function, were investigated. The administration of SYD significantly ameliorated the clinical signs, suppressed the levels of proinflammatory cytokines, and reduced immune cell infiltrates into colonic tissues of DSS-induced colitis model mice. SYD also significantly reduced the DSS-induced activation of STAT3 and NF-κB signaling. Furthermore, SYD promoted epithelial integrity by regulating epithelial cell apoptosis and epithelial permeability. Finally, we demonstrated that SYD protected the intestinal barrier function by significantly regulating the mucus layer genes Muc1, Muc2, Muc4, and Tff3, as well as the epithelial barrier genes Z O -1 and Occludin. Our results indicate that SYD has a protective effect on DSS-induced colitis, which is attributable to its anti-inflammatory activity and intestinal barrier function-enhancing effects. These results provide valuable insights into the pharmacological actions of SYD for the treatment of IBD.

Placental Malaria: Histological Findings and Immune Cell Infiltrates in Submicroscopic Infections

Most research on placental malaria is focused on microscopic infection by Plasmodium falciparum; there are very few studies on submicroscopic infection. This study aimed to assess alterations of placental tissue associated with placental malaria, to describe the immune cell populations in the placental tissue, and to explore the relationships between the histopathological changes and cell infiltrates. A descriptive, prospective and cross-sectional study was carried out. Women were recruited at hospital obstetric facilities in three municipalities in Northwest Colombia. The histopathological analysis was performed in a total of 132 placentas including 66 placentas with submicroscopic plasmodial infection and 66 that were negatives. Immunohistochemistry was performed on a subset of 75 placentas to determine the distribution of immune cells. Based on histology, there were more immune cells in placentas with submicroscopic plasmodial infection compared with those without infection. The quantity of syncytial knots and calcifications was greater with submicroscopic plasmodial infection, but the quantity of abruption and thrombi was greater in placentas without infection. By immunohistochemistry, we observed a significant increase of CD56+ and CD68+ cells in the infected placentas. Submicroscopic plasmodial infection in the placenta causes tissue alterations and increased immune cell infiltrates. Submicroscopic plasmodial infection is very common in Colombia and can represent a serious threat to mothers and newborns.

MYC oncogene is associated with suppression of tumor immunity and targeting Myc induces tumor cell immunogenicity for therapeutic whole cell vaccination

BackgroundMYC oncogene is deregulated in 70% of all human cancers and is associated with multiple oncogenic functions including immunosuppression in the tumor microenvironment. The role of MYC in the immune microenvironment of neuroblastoma and melanoma is investigated and the effect of targeting Myc on immunogenicity of cancer cells is evaluated.MethodsImmune cell infiltrates and immunogenic pathway signatures in the context of MYCN amplification were analyzed in human neuroblastoma tumors and in metastatic melanoma. Dose response and cell susceptibility to MYC inhibitors (I-BET726 and JQ1) were determined in mouse cell lines. The influence of downregulating Myc in tumor cells was characterized by immunogenic pathway signatures and functional assays. Myc-suppressed tumor cells were used as whole cell vaccines in preclinical neuroblastoma and melanoma models.ResultsAnalysis of immune phenotype in human neuroblastoma and melanoma tumors revealed that MYCN or c-MYC amplified tumors respectively are associated with suppressed immune cell infiltrates and functional pathways. Targeting Myc in cancer cells with I-BET726 and JQ1 results in cell cycle arrest and induces cell immunogenicity. Combining vaccination of Myc-inhibited tumor cells with checkpoint inhibition induced robust antitumor immunity and resulted in therapeutic cancer vaccine therapy in mouse neuroblastoma tumors. Despite vigorous antitumor immunity in the mouse melanoma model, upregulation of immunosuppressive pathways enabled tumor escape.ConclusionsThis study demonstrates that the Myc oncogene is an appropriate target for inducing tumor cell immunogenicity and suggests that Myc-suppressed whole tumor cells combined with checkpoint therapy could be used for formulating a personalized therapeutic tumor vaccine.