Untargeted Metabolomic Profiling Reveals Differentially Expressed Serum Metabolites and Pathways in Type 2 Diabetes Patients with and without Cognitive Decline: A Cross-Sectional Study.

Diabetes is recognized as a risk factor for cognitive decline, but the underlying mechanisms remain elusive. We aimed to identify the metabolic pathways altered in diabetes-associated cognitive decline (DACD) using untargeted metabolomics. We conducted liquid chromatography-mass spectrometry-based untargeted metabolomics to profile serum metabolite levels in 100 patients with type 2 diabetes (T2D) (54 without and 46 with DACD). Multivariate statistical tools were used to identify the differentially expressed metabolites (DEMs), and enrichment and pathways analyses were used to identify the signaling pathways associated with the DEMs. The receiver operating characteristic (ROC) analysis was employed to assess the diagnostic accuracy of a set of metabolites. We identified twenty DEMs, seven up- and thirteen downregulated in the DACD vs. DM group. Chemometric analysis revealed distinct clustering between the two groups. Metabolite set enrichment analysis found significant enrichment in various metabolite sets, including galactose metabolism, arginine and unsaturated fatty acid biosynthesis, citrate cycle, fructose and mannose, alanine, aspartate, and glutamate metabolism. Pathway analysis identified six significantly altered pathways, including arginine and unsaturated fatty acid biosynthesis, and the metabolism of the citrate cycle, alanine, aspartate, glutamate, a-linolenic acid, and glycerophospholipids. Classifier models with AUC-ROC > 90% were developed using individual metabolites or a combination of individual metabolites and metabolite ratios. Our study provides evidence of perturbations in multiple metabolic pathways in patients with DACD. The distinct DEMs identified in this study hold promise as diagnostic biomarkers for DACD patients.

Uncovering a neurological protein signature for severe COVID-19.

Coronavirus disease of 2019 (COVID-19), caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has sparked a global pandemic with severe complications and high morbidity rate. Neurological symptoms in COVID-19 patients, and neurological sequelae post COVID-19 recovery have been extensively reported. Yet, neurological molecular signature and signaling pathways that are affected in the central nervous system (CNS) of COVID-19 severe patients remain still unknown and need to be identified. Plasma samples from 49 severe COVID-19 patients, 50 mild COVID-19 patients, and 40 healthy controls were subjected to Olink proteomics analysis of 184 CNS-enriched proteins. By using a multi-approach bioinformatics analysis, we identified a 34-neurological protein signature for COVID-19 severity and unveiled dysregulated neurological pathways in severe cases. Here, we identified a new neurological protein signature for severe COVID-19 that was validated in different independent cohorts using blood and postmortem brain samples and shown to correlate with neurological diseases and pharmacological drugs. This protein signature could potentially aid the development of prognostic and diagnostic tools for neurological complications in post-COVID-19 convalescent patients with long term neurological sequelae.

Modulation of SLFN11 induces changes in DNA Damage response in breast cancer.

BACKGROUND: Lack of Schlafen family member 11 (SLFN11) expression has been recently identified as a dominant genomic determinant of response to DNA damaging agents in numerous cancer types. Thus, several strategies aimed at increasing SLFN11 are explored to restore chemosensitivity of refractory cancers. In this study, we examined various approaches to elevate SLFN11 expression in breast cancer cellular models and confirmed a corresponding increase in chemosensitivity with using the most successful efficient one. As oncogenic transcriptomic downregulation is often driven by methylation of the promotor region, we explore the demethylation effect of 5-aza-2'-deoxycytidine (decitabine), on the SLFN11 gene. Since SLFN11 has been reported as an interferon inducible gene, and interferon is secreted during an active anti-tumor immune response, we investigated the in vitro effect of IFN-γ on SLFN11 expression in breast cancer cell lines. As a secondary approach to pick up cross talk between immune cells and SLFN11 expression we used indirect co-culture of breast cancer cells with activated PBMCs and evaluated if this can drive SLFN11 upregulation. Finally, as a definitive and specific way to modulate SLFN11 expression we implemented SLFN11 dCas9 (dead CRISPR associated protein 9) systems to specifically increase or decrease SLFN11 expression. RESULTS: After confirming the previously reported correlation between methylation of SLFN11 promoter and its expression across multiple cell lines, we showed in-vitro that decitabine and IFN-γ could increase moderately the expression of SLFN11 in both BT-549 and T47D cell lines. The use of a CRISPR-dCas9 UNISAM and KRAB system could increase or decrease SLFN11 expression significantly (up to fivefold), stably and specifically in BT-549 and T47D cancer cell lines. We then used the modified cell lines to quantify the alteration in chemo sensitivity of those cells to treatment with DNA Damaging Agents (DDAs) such as Cisplatin and Epirubicin or DNA Damage Response (DDRs) drugs like Olaparib. RNAseq was used to elucidate the mechanisms of action affected by the alteration in SLFN11 expression. In cell lines with robust SLFN11 promoter methylation such as MDA-MB-231, no SLFN11 expression could be induced by any approach. CONCLUSION: To our knowledge this is the first report of the stable non-lethal increase of SLFN11 expression in a cancer cell line. Our results show that induction of SLFN11 expression can enhance DDA and DDR sensitivity in breast cancer cells and dCas9 systems may represent a novel approach to increase SLFN11 and achieve higher sensitivity to chemotherapeutic agents, improving outcome or decreasing required drug concentrations. SLFN11-targeting therapies might be explored pre-clinically to develop personalized approaches.

Blood Proteomics Analysis Reveals Potential Biomarkers and Convergent Dysregulated Pathways in Autism Spectrum Disorder: A Pilot Study.

Autism spectrum disorder (ASD) is an umbrella term that encompasses several disabling neurodevelopmental conditions. These conditions are characterized by impaired manifestation in social and communication skills with repetitive and restrictive behaviors or interests. Thus far, there are no approved biomarkers for ASD screening and diagnosis; also, the current diagnosis depends heavily on a physician's assessment and family's awareness of ASD symptoms. Identifying blood proteomic biomarkers and performing deep blood proteome profiling could highlight common underlying dysfunctions between cases of ASD, given its heterogeneous nature, thus laying the foundation for large-scale blood-based biomarker discovery studies. This study measured the expression of 1196 serum proteins using proximity extension assay (PEA) technology. The screened serum samples included ASD cases (n = 91) and healthy controls (n = 30) between 6 and 15 years of age. Our findings revealed 251 differentially expressed proteins between ASD and healthy controls, of which 237 proteins were significantly upregulated and 14 proteins were significantly downregulated. Machine learning analysis identified 15 proteins that could be biomarkers for ASD with an area under the curve (AUC) = 0.876 using support vector machine (SVM). Gene Ontology (GO) analysis of the top differentially expressed proteins (TopDE) and weighted gene co-expression analysis (WGCNA) revealed dysregulation of SNARE vesicular transport and ErbB pathways in ASD cases. Furthermore, correlation analysis showed that proteins from those pathways correlate with ASD severity. Further validation and verification of the identified biomarkers and pathways are warranted.

Blood-Based Proteomic Profiling Identifies Potential Biomarker Candidates and Pathogenic Pathways in Dementia.

Dementia is a progressive and debilitating neurological disease that affects millions of people worldwide. Identifying the minimally invasive biomarkers associated with dementia that could provide insights into the disease pathogenesis, improve early diagnosis, and facilitate the development of effective treatments is pressing. Proteomic studies have emerged as a promising approach for identifying the protein biomarkers associated with dementia. This pilot study aimed to investigate the plasma proteome profile and identify a panel of various protein biomarkers for dementia. We used a high-throughput proximity extension immunoassay to quantify 1090 proteins in 122 participants (22 with dementia, 64 with mild cognitive impairment (MCI), and 36 controls with normal cognitive function). Limma-based differential expression analysis reported the dysregulation of 61 proteins in the plasma of those with dementia compared with controls, and machine learning algorithms identified 17 stable diagnostic biomarkers that differentiated individuals with AUC = 0.98 ± 0.02. There was also the dysregulation of 153 plasma proteins in individuals with dementia compared with those with MCI, and machine learning algorithms identified 8 biomarkers that classified dementia from MCI with an AUC of 0.87 ± 0.07. Moreover, multiple proteins selected in both diagnostic panels such as NEFL, IL17D, WNT9A, and PGF were negatively correlated with cognitive performance, with a correlation coefficient (r2) ≤ -0.47. Gene Ontology (GO) and pathway analysis of dementia-associated proteins implicated immune response, vascular injury, and extracellular matrix organization pathways in dementia pathogenesis. In conclusion, the combination of high-throughput proteomics and machine learning enabled us to identify a blood-based protein signature capable of potentially differentiating dementia from MCI and cognitively normal controls. Further research is required to validate these biomarkers and investigate the potential underlying mechanisms for the development of dementia.

Immune-related 3-lncRNA signature with prognostic connotation in a multi-cancer setting.

BACKGROUND: Advances in our understanding of the tumor microenvironment have radically changed the cancer field, highlighting the emerging need for biomarkers of an active, favorable tumor immune phenotype to aid treatment stratification and clinical prognostication. Numerous immune-related gene signatures have been defined; however, their prognostic value is often limited to one or few cancer types. Moreover, the area of non-coding RNA as biomarkers remains largely unexplored although their number and biological roles are rapidly expanding. METHODS: We developed a multi-step process to identify immune-related long non-coding RNA signatures with prognostic connotation in multiple TCGA solid cancer datasets. RESULTS: Using the breast cancer dataset as a discovery cohort we found 2988 differentially expressed lncRNAs between immune favorable and unfavorable tumors, as defined by the immunologic constant of rejection (ICR) gene signature. Mapping of the lncRNAs to a coding-non-coding network identified 127 proxy protein-coding genes that are enriched in immune-related diseases and functions. Next, we defined two distinct 20-lncRNA prognostic signatures that show a stronger effect on overall survival than the ICR signature in multiple solid cancers. Furthermore, we found a 3 lncRNA signature that demonstrated prognostic significance across 5 solid cancer types with a stronger association with clinical outcome than ICR. Moreover, this 3 lncRNA signature showed additional prognostic significance in uterine corpus endometrial carcinoma and cervical squamous cell carcinoma and endocervical adenocarcinoma as compared to ICR. CONCLUSION: We identified an immune-related 3-lncRNA signature with prognostic connotation in multiple solid cancer types which performed equally well and in some cases better than the 20-gene ICR signature, indicating that it could be used as a minimal informative signature for clinical implementation.

Immune checkpoints in the tumor microenvironment.

Interactions between immune checkpoints (ICs) and their ligands negatively regulate T cell activation pathways involved in physiological immune responses against specific antigens. ICs and their ligands are frequently upregulated in the tumor microenvironment (TME) of various malignancies, and they represent significant barriers for induction of effective anti-tumor immune responses. Several IC inhibitors (ICIs) have been developed, with some currently in clinical trials and others have been approved for the treatment of different cancers. However, tumor cells are able to counteract the activity of ICIs and can commission additional inhibitory pathways via expression of other ICs/ligands within the TME. This review discusses the expression of various ICs/ligands in the TME and their impact on tumor immune evasion. Additionally, we discuss various regulatory mechanisms, including genetic and epigenetic, and other modulatory factors including hypoxia and the presence of immunosuppressive populations in the TME, which result in upregulation of ICs in various cancers. Moreover, we discuss the prognostic significance of ICs and their ligands, and the potential strategies to enhance treatment responses to ICIs. This review aims to advance our current knowledge on the role of ICs in the TME and the clinical benefits of targeting them.

Cancer testis antigen PRAME: An anti-cancer target with immunomodulatory potential.

PReferentially expressed Antigen in Melanoma (PRAME) is a cancer testis antigen with restricted expression in somatic tissues and re-expression in poor prognostic solid tumours. PRAME has been extensively investigated as a target for immunotherapy, however, its role in modulating the anti-tumour immune response remains largely unknown. Here, we show that PRAME tumour expression is associated with worse survival in the TCGA breast cancer cohort, particularly in immune-unfavourable tumours. Using direct and indirect co-culture models, we found that PRAME overexpressing MDA-MB-468 breast cancer cells inhibit T cell activation and cytolytic potential, which could be partly restored by silencing of PRAME. Furthermore, silencing of PRAME reduced expression of several immune checkpoints and their ligands, including PD-1, LAG3, PD-L1, CD86, Gal-9 and VISTA. Interestingly, silencing of PRAME induced cancer cell killing to levels similar to anti-PD-L1 atezolizumab treatment. Comprehensive analysis of soluble inflammatory mediators and cancer cell expression of immune-related genes showed that PRAME tumour expression can suppress the expression and secretion of multiple pro-inflammatory cytokines, and mediators of T cell activation, differentiation and cytolysis. Together, our data indicate that targeting of PRAME offers a potential, novel dual therapeutic approach to specifically target tumour cells and regulate immune activation in the tumour microenvironment.

Identification of two HLA-A*0201 immunogenic epitopes of lactate dehydrogenase C (LDHC): potential novel targets for cancer immunotherapy.

Lactate dehydrogenase C (LDHC) is an archetypical cancer testis antigen with limited expression in adult tissues and re-expression in tumors. This restricted expression pattern together with the important role of LDHC in cancer metabolism renders LDHC a potential target for immunotherapy. This study is the first to investigate the immunogenicity of LDHC using T cells from healthy individuals. LDHC-specific T cell responses were induced by in vitro stimulation with synthetic peptides, or by priming with autologous peptide-pulsed dendritic cells. We evaluated T cell activation by IFN-γ ELISpot and determined cytolytic activity of HLA-A*0201-restricted T cells in breast cancer cell co-cultures. In vitro T cell stimulation induced IFN-γ secretion in response to numerous LDHC-derived peptides. Analysis of HLA-A*0201 responses revealed a significant T cell activation after stimulation with peptide pools 2 (PP2) and 8 (PP8). The PP2- and PP8-specific T cells displayed cytolytic activity against breast cancer cells with endogenous LDHC expression within a HLA-A*0201 context. We identified peptides LDHC41-55 and LDHC288-303 from PP2 and PP8 to elicit a functional cellular immune response. More specifically, we found an increase in IFN-γ secretion by CD8 + T cells and cancer-cell-killing of HLA-A*0201/LDHC positive breast cancer cells by LDHC41-55- and LDHC288-303-induced T cells, albeit with a possible antigen recognition threshold. The majority of induced T cells displayed an effector memory phenotype. To conclude, our findings support the rationale to assess LDHC as a targetable cancer testis antigen for immunotherapy, and in particular the HLA-A*0201 restricted LDHC41-55 and LDHC288-303 peptides within LDHC.

PRAME promotes epithelial-to-mesenchymal transition in triple negative breast cancer.

BACKGROUND: The triple negative breast cancer (TNBC) paradox marks a major challenge in the treatment-decision making process. TNBC patients generally respond better to neoadjuvant chemotherapy compared to other breast cancer patients; however, they have a substantial higher risk of disease recurrence. We evaluated the expression of the tumor-associated antigen PReferentially Antigen expressed in MElanoma (PRAME) as a prognostic biomarker in breast cancer and explored its role in cell migration and invasion, key hallmarks of progressive and metastatic disease. METHODS: TCGA and GTeX datasets were interrogated to assess the expression of PRAME in relation to overall and disease-free survival. The role of PRAME in cell migration and invasion was investigated using gain- and loss-of-function TNBC cell line models. RESULTS: We show that PRAME promotes migration and invasion of TNBC cells through changes in expression of E-cadherin, N-cadherin, vimentin and ZEB1, core markers of an epithelial-to-mesenchymal transition. Mechanistic analysis of PRAME-overexpressing cells showed an upregulation of 11 genes (SNAI1, TCF4, TWIST1, FOXC2, IL1RN, MMP2, SOX10, WNT11, MMP3, PDGFRB, and JAG1) and downregulation of 2 genes (BMP7 and TSPAN13). Gene ontology analyses revealed enrichment of genes that are dysregulated in ovarian and esophageal cancer and are involved in transcription and apoptosis. In line with this, interrogation of TCGA and GTEx data demonstrated an increased PRAME expression in ovarian and esophageal tumor tissues in addition to breast tumors where it is associated with worse survival. CONCLUSIONS: Our findings indicate that PRAME plays a tumor-promoting role in triple negative breast cancer by increasing cancer cell motility through EMT-gene reprogramming. Therefore, PRAME could serve as a prognostic biomarker and/or therapeutic target in TNBC.

Loss of MMP-8 in ductal carcinoma in situ (DCIS)-associated myoepithelial cells contributes to tumour promotion through altered adhesive and proteolytic function.

BACKGROUND: Normal myoepithelial cells (MECs) play an important tumour-suppressor role in the breast but display an altered phenotype in ductal carcinoma in situ (DCIS), gaining tumour-promoter functions. Matrix metalloproteinase-8 (MMP-8) is expressed by normal MECs but is lost in DCIS. This study investigated the function of MMP-8 in MECs and the impact of its loss in DCIS. METHODS: Primary normal and DCIS-associated MECs, and normal (N-1089) and DCIS-modified myoepithelial (ß6-1089) cell lines, were used to assess MMP-8 expression and function. ß6-1089 lacking MMP-8 were transfected with MMP-8 WT and catalytically inactive MMP-8 EA, and MMP-8 in N-1089 MEC was knocked down with siRNA. The effect on adhesion and migration to extracellular matrix (ECM), localisation of α6ß4 integrin to hemidesmosomes (HD), TGF-ß signalling and gelatinase activity was measured. The effect of altering MEC MMP-8 expression on tumour cell invasion was investigated in 2D and 3D organotypic models. RESULTS: Assessment of primary cells and MEC lines confirmed expression of MMP-8 in normal MEC and its loss in DCIS-MEC. Over-expression of MMP-8 WT but not MMP-8 EA in ß6-1089 cells increased adhesion to ECM proteins and reduced migration. Conversely, knock-down of MMP-8 in N-1089 reduced adhesion and increased migration. Expression of MMP-8 WT in ß6-1089 led to greater localisation of α6ß4 to HD and reduced retraction fibre formation, this being reversed by MMP-8 knock-down in N-1089. Over-expression of MMP-8 WT reduced TGF-ß signalling and gelatinolytic activity. MMP-8 knock-down enhanced TGF-ß signalling and gelatinolytic activity, which was reversed by blocking MMP-9 by knock-down or an inhibitor. MMP-8 WT but not MMP-8 EA over-expression in ß6-1089 reduced breast cancer cell invasion in 2D and 3D invasion assays, while MMP-8 knock-down in N-1089 enhanced cancer cell invasion. Staining of breast cancer cases for MMP-8 revealed a statistically significant loss of MMP-8 expression in DCIS with invasion versus pure DCIS (p = 0.001). CONCLUSIONS: These data indicate MMP-8 is a vital component of the myoepithelial tumour-suppressor function. It restores MEC interaction with the matrix, opposes TGF-ß signalling and MMP-9 proteolysis, which contributes to inhibition of tumour cell invasion. Assessment of MMP-8 expression may help to determine risk of DCIS progression.

Immunogenomic Classification of Colorectal Cancer and Therapeutic Implications.

The immune system has a substantial effect on colorectal cancer (CRC) progression. Additionally, the response to immunotherapeutics and conventional treatment options (e.g., chemotherapy, radiotherapy and targeted therapies) is influenced by the immune system. The molecular characterization of colorectal cancer (CRC) has led to the identification of favorable and unfavorable immunological attributes linked to clinical outcome. With the definition of consensus molecular subtypes (CMSs) based on transcriptomic profiles, multiple characteristics have been proposed to be responsible for the development of the tumor immune microenvironment and corresponding mechanisms of immune escape. In this review, a detailed description of proposed immune phenotypes as well as their interaction with different therapeutic modalities will be provided. Finally, possible strategies to shift the CRC immune phenotype towards a reactive, anti-tumor orientation are proposed per CMS.

Pleiotropic functions of the tumor- and metastasis-suppressing matrix metalloproteinase-8 in mammary cancer in MMTV-PyMT transgenic mice.

INTRODUCTION: Matrix metalloproteinase-8 (MMP-8; neutrophil collagenase) is an important regulator of innate immunity that has oncosuppressive actions in numerous tumor types. METHODS: We have intercrossed Mmp8-null mice with the Polyoma virus middle T oncogene-driven (MMTV-PyMT) mouse model of mammary cancer to explore the effects of loss of MMP-8 on the incidence and progression of mammary carcinomas. RESULTS: In this aggressive mouse model of breast cancer, loss of MMP-8 accelerated tumor onset even further, such that 90% of MMTV-PyMT; Mmp8-null female mice were tumor-bearing at the time of weaning. Throughout the 14 weeks of the model, tumor burden increased in homozygous Mmp8-null mice compared to Mmp8-wild-type and -heterozygote animals. Likewise, lung metastasis dramatically increased in the MMTV-PyMT; Mmp8-null mice. Immunohistochemistry revealed that tumors in wild-type, Mmp8-heterozygotes and -null animals had similar vascular density at 8 weeks, but at 10 weeks Mmp8-wild-type tumors had a lower vascularity than their heterozygote and null counterparts. No differences in macrophage infiltration were apparent throughout primary tumor development, though at 10 weeks a drop in neutrophil infiltrates was observed in Mmp8-wild-type tumors. Using quantitative real-time RT-PCR, we tracked the expression of the entire Mmp and Timp gene families, observing a significant decrease in Mmp3 expression in Mmp8-null tumors compared to wild-type and heterozygotes throughout the time course of the model, which was confirmed at the protein level. CONCLUSIONS: These findings provide novel insight into the suppressive action of MMP-8 on mammary tumorigenesis and metastasis, and indicate that the loss of MMP-8 likely has pleiotropic effects on innate immunity and angiogenesis that are reflected in changes in the protease web.

Metalloproteinase-dependent and -independent processes contribute to inhibition of breast cancer cell migration, angiogenesis and liver metastasis by a disintegrin and metalloproteinase with thrombospondin motifs-15.

The ADAMTS proteinases are a family of secreted, matrix-associated enzymes that have diverse roles in the regulation of tissue organization and vascular homeostasis. Several of the 19 human family members have been identified as having either tumor promoting or suppressing roles. We previously demonstrated that decreased ADAMTS15 expression correlated with a worse clinical outcome in mammary carcinoma (e.g., Porter et al., Int J Cancer 2006;118:1241-7). We have explored the effects of A Disintegrin and Metalloproteinase with Thrombospondin motifs-15 (ADAMTS-15) on the behavior of MDA-MB-231 and MCF-7 breast cancer cells by stable expression of either a wild-type (wt) or metalloproteinase-inactive (E362A) protein. No effects on mammary cancer cell proliferation or apoptosis were observed for either form of ADAMTS-15. However, both forms reduced cell migration on fibronectin or laminin matrices, though motility on a Type I collagen matrix was unimpaired. Knockdown of syndecan-4 attenuated the inhibitory effects of ADAMTS-15 on cell migration. In contrast to its effects on cell migration, wt ADAMTS-15 but not the E362A inactive mutant inhibited endothelial tubulogenesis in 3D collagen gels and angiogenesis in the aortic ring assay. In experimental metastasis assays in nude mice, MDA-MB-231 cells expressing either form of ADAMTS-15 showed reduced spread to the liver, though lung colonization was enhanced for cells expressing wt ADAMTS-15. These studies indicate that extracellular ADAMTS-15 has multiple actions on tumor pathophysiology. Via modulation of cell-ECM interactions, which likely involve syndecan-4, it attenuates mammary cancer cell migration independent of its metalloproteinase activity; however, its antiangiogenic action requires catalytic functionality, and its effects on metastasis in vivo are tissue niche-dependent.

Matrix metalloproteinase 8 (collagenase 2) induces the expression of interleukins 6 and 8 in breast cancer cells.

Matrix metalloproteinase 8 (MMP-8) is a tumor-suppressive protease that cleaves numerous substrates, including matrix proteins and chemokines. In particular, MMP-8 proteolytically activates IL-8 and, thereby, regulates neutrophil chemotaxis in vivo. We explored the effects of expression of either a WT or catalytically inactive (E198A) mutant version of MMP-8 in human breast cancer cell lines. Analysis of serum-free conditioned media from three breast cancer cell lines (MCF-7, SK-BR-3, and MDA-MB-231) expressing WT MMP-8 revealed elevated levels of IL-6 and IL-8. This increase was mirrored at the mRNA level and was dependent on MMP-8 catalytic activity. However, sustained expression of WT MMP-8 by breast cancer cells was non-permissive for long-term growth, as shown by reduced colony formation compared with cells expressing either control vector or E198A mutant MMP-8. In long-term culture of transfected MDA-MB-231 cells, expression of WT but not E198A mutant MMP-8 was lost, with IL-6 and IL-8 levels returning to base line. Rare clonal isolates of MDA-MB-231 cells expressing WT MMP-8 were generated, and these showed constitutively high levels of IL-6 and IL-8, although production of the interleukins was no longer dependent upon MMP-8 activity. These studies support a causal connection between MMP-8 activity and the IL-6/IL-8 network, with an acute response to MMP-8 involving induction of the proinflammatory mediators, which may in part serve to compensate for the deleterious effects of MMP-8 on breast cancer cell growth. This axis may be relevant to the recognized ability of MMP-8 to orchestrate the innate immune system in inflammation in vivo.

Cancer testis antigens: Emerging therapeutic targets leveraging genomic instability in cancer.

Cancer care has witnessed remarkable progress in recent decades, with a wide array of targeted therapies and immune-based interventions being added to the traditional treatment options such as surgery, chemotherapy, and radiotherapy. However, despite these advancements, the challenge of achieving high tumor specificity while minimizing adverse side effects continues to dictate the benefit-risk balance of cancer therapy, guiding clinical decision making. As such, the targeting of cancer testis antigens (CTAs) offers exciting new opportunities for therapeutic intervention of cancer since they display highly tumor specific expression patterns, natural immunogenicity and play pivotal roles in various biological processes that are critical for tumor cellular fitness. In this review, we delve deeper into how CTAs contribute to the regulation and maintenance of genomic integrity in cancer, and how these mechanisms can be exploited to specifically target and eradicate tumor cells. We review the current clinical trials targeting aforementioned CTAs, highlight promising pre-clinical data and discuss current challenges and future perspectives for future development of CTA-based strategies that exploit tumor genomic instability.

Transcriptome profiling and network enrichment analyses identify subtype-specific therapeutic gene targets for breast cancer and their microRNA regulatory networks.

Previous studies have suggested that breast cancer (BC) from the Middle East and North Africa (MENA) is presented at younger age with advanced tumor stage, indicating underlying biological differences. Given the scant transcriptomic data on BC from the MENA region and to better understand the biology of this disease, we performed mRNA and microRNA (miRNA) transcriptomic profiling on a local cohort of BC (n = 96) from Qatar. Our data revealed the differentially expressed genes and miRNAs as function of BC molecular subtypes (HR+, HER2+, HER2+HR+, and TNBC), tumor grade (GIII vs GI-II), patients' age (young (≤40) vs old (>40)), and ethnicity (MENA vs non-MENA). Our profiling data revealed close similarity between TNBC and HER2+, while the transcriptome of HER2+HR+ tumor was resemblant of that from HR+ tumors. Network analysis identified complex miRNA-mRNA regulatory networks in each BC molecular subtype, in high vs low grade tumors, in tumors from young vs old patients, and in tumors from MENA vs non-MENA, thus implicating miRNA-mediated gene regulation as an essential mechanism in shaping the transcriptome of BC. Integration of our transcriptomic data with CRISPR-Cas9 functional screen data and the OncoKB database identified numerous dependencies and therapeutic vulnerabilities in each BC molecular subtype, while CDC123 was functionally validated as potential therapeutic target for TNBC. Cox regression survival analyses identified mRNA and miRNA-based signatures predicative of worse and better relapse free survival (RFS), which were validated in larger BC cohorts. Our data provides comprehensive transcriptomic profiling and unraveled the miRNA-mRNA regulatory networks in BC patients from the region and identified novel actionable gene targets, employing integrated approach. Findings from the current study have potential implications to improve the current standard-of-care for BC from the MENA as well as patients from other ethnicities.

High-throughput autoantibody screening identifies differentially abundant autoantibodies in autism spectrum disorder.

Introduction: Autism spectrum disorder (ASD) is a neurodevelopmental condition characterized by defects in two core domains, social/communication skills and restricted/repetitive behaviors or interests. There is no approved biomarker for ASD diagnosis, and the current diagnostic method is based on clinical manifestation, which tends to vary vastly between the affected individuals due to the heterogeneous nature of ASD. There is emerging evidence that supports the implication of the immune system in ASD, specifically autoimmunity; however, the role of autoantibodies in ASD children is not yet fully understood. Materials and methods: In this study, we screened serum samples from 93 cases with ASD and 28 healthy controls utilizing high-throughput KoRectly Expressed (KREX) i-Ome protein-array technology. Our goal was to identify autoantibodies with differential expressions in ASD and to gain insights into the biological significance of these autoantibodies in the context of ASD pathogenesis. Result: Our autoantibody expression analysis identified 29 differential autoantibodies in ASD, 4 of which were upregulated and 25 downregulated. Subsequently, gene ontology (GO) and network analysis showed that the proteins of these autoantibodies are expressed in the brain and involved in axonal guidance, chromatin binding, and multiple metabolic pathways. Correlation analysis revealed that these autoantibodies negatively correlate with the age of ASD subjects. Conclusion: This study explored autoantibody reactivity against self-antigens in ASD individuals' serum using a high-throughput assay. The identified autoantibodies were reactive against proteins involved in axonal guidance, synaptic function, amino acid metabolism, fatty acid metabolism, and chromatin binding.

Profiling the autoantibody repertoire reveals autoantibodies associated with mild cognitive impairment and dementia.

Background: Dementia is a debilitating neurological disease affecting millions of people worldwide. The exact mechanisms underlying the initiation and progression of the disease remain to be fully defined. There is an increasing body of evidence for the role of immune dysregulation in the pathogenesis of dementia, where blood-borne autoimmune antibodies have been studied as potential markers associated with pathological mechanisms of dementia. Methods: This study included plasma from 50 cognitively normal individuals, 55 subjects with MCI (mild cognitive impairment), and 22 subjects with dementia. Autoantibody profiling for more than 1,600 antigens was performed using a high throughput microarray platform to identify differentially expressed autoantibodies in MCI and dementia. Results: The differential expression analysis identified 33 significantly altered autoantibodies in the plasma of patients with dementia compared to cognitively normal subjects, and 38 significantly altered autoantibodies in the plasma of patients with dementia compared to subjects with MCI. And 20 proteins had significantly altered autoantibody responses in MCI compared to cognitively normal individuals. Five autoantibodies were commonly dysregulated in both dementia and MCI, including anti-CAMK2A, CKS1B, ETS2, MAP4, and NUDT2. Plasma levels of anti-ODF3, E6, S100P, and ARHGDIG correlated negatively with the cognitive performance scores (MoCA) (r2 -0.56 to -0.42, value of p < 0.001). Additionally, several proteins targeted by autoantibodies dysregulated in dementia were significantly enriched in the neurotrophin signaling pathway, axon guidance, cholinergic synapse, long-term potentiation, apoptosis, glycolysis and gluconeogenesis. Conclusion: We have shown multiple dysregulated autoantibodies in the plasma of subjects with MCI and dementia. The corresponding proteins for these autoantibodies are involved in neurodegenerative pathways, suggesting a potential impact of autoimmunity on the etiology of dementia and the possible benefit for future therapeutic approaches. Further investigations are warranted to validate our findings.

Auto-immunoproteomics analysis of COVID-19 ICU patients revealed increased levels of autoantibodies related to the male reproductive system.

Background: Coronavirus disease (COVID-19) manifests many clinical symptoms, including an exacerbated immune response and cytokine storm. Autoantibodies in COVID-19 may have severe prodromal effects that are poorly understood. The interaction between these autoantibodies and self-antigens can result in systemic inflammation and organ dysfunction. However, the role of autoantibodies in COVID-19 complications has yet to be fully understood. Methods: The current investigation screened two independent cohorts of 97 COVID-19 patients [discovery (Disc) cohort from Qatar (case = 49 vs. control = 48) and replication (Rep) cohort from New York (case = 48 vs. control = 28)] utilizing high-throughput KoRectly Expressed (KREX) Immunome protein-array technology. Total IgG autoantibody responses were evaluated against 1,318 correctly folded and full-length human proteins. Samples were randomly applied on the precoated microarray slides for 2 h. Cy3-labeled secondary antibodies were used to detect IgG autoantibody response. Slides were scanned at a fixed gain setting using the Agilent fluorescence microarray scanner, generating a 16-bit TIFF file. Group comparisons were performed using a linear model and Fisher's exact test. Differentially expressed proteins were used for KEGG and WIKIpathway annotation to determine pathways in which the proteins of interest were significantly over-represented. Results and conclusion: Autoantibody responses to 57 proteins were significantly altered in the COVID-19 Disc cohort compared to healthy controls (p ≤ 0.05). The Rep cohort had altered autoantibody responses against 26 proteins compared to non-COVID-19 ICU patients who served as controls. Both cohorts showed substantial similarities (r 2 = 0.73) and exhibited higher autoantibody responses to numerous transcription factors, immunomodulatory proteins, and human disease markers. Analysis of the combined cohorts revealed elevated autoantibody responses against SPANXN4, STK25, ATF4, PRKD2, and CHMP3 proteins in COVID-19 patients. The sequences for SPANXN4 and STK25 were cross-validated using sequence alignment tools. ELISA and Western blot further verified the autoantigen-autoantibody response of SPANXN4. SPANXN4 is essential for spermiogenesis and male fertility, which may predict a potential role for this protein in COVID-19-associated male reproductive tract complications, and warrants further research.