Laboratory Investigation最新文献

{"title":"Leptin modulates ovarian granulosa cell apoptosis by regulating telomerase activity and telomere length in polycystic ovary syndrome.","authors":"Feijing Zhou, Zhimin Sun, Luyao Cheng, Yuezhi Dong","doi":"10.1016/j.labinv.2024.102169","DOIUrl":"https://doi.org/10.1016/j.labinv.2024.102169","url":null,"abstract":"<p><p>Leptin (LEP) is implicated in the pathogenesis of polycystic ovary syndrome (PCOS). This study investigates the mechanism of LEP in PCOS. The baseline information of 80 PCOS patients and matched controls was analyzed, with serum and follicular fluid (FF) LEP and LEP receptor (LEPR) levels, telomerase activity, and relative telomere length (TL) measured. The correlation of FF LEP with telomerase activity and TL was analyzed. The viability and apoptosis of KGN cells (the ovarian granulosa cells) treated with gradient LEP were assessed. LEP-LEPR interaction was examined. LEPR, c-MYC, and TERT levels and c-MYC protein expression in the TERT promoter region were determined. Nuclear c-MYC translocation was detected. LEP was upregulated in sera and FF of PCOS patients. FF LEP positively-correlated with telomerase activity and TL. Low-concentration LEP facilitated KGN cell proliferation and high-concentration LEP dose-dependently suppressed cell proliferation, promoted apoptosis, upregulated LEPR and increased telomerase activity and relative TL. LEP-LEPR interaction upregulated c-MYC and facilitated its nuclear accumulation. c-MYC enrichment in the TERT promoter region upregulated TERT, altering telomerase activity and TL and inducing cell apoptosis. Briefly, LEP/LEPR activate c-MYC, modulate TERT expression, and increase telomerase activity and TL, thus inducing ovarian granulosa cell apoptosis and participating in PCOS.</p>","PeriodicalId":17930,"journal":{"name":"Laboratory Investigation","volume":null,"pages":null},"PeriodicalIF":5.1,"publicationDate":"2024-11-02","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142567318","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Deciphering the intricate relationship between macrophages, pigmentation, and prognosis in uveal melanoma.","authors":"Jayanti Jha, Mithalesh Kumar Singh, Lata Singh, Neelam Pushker, Aanchal Kakkar, Rachna Meel, Neiwete Lomi, Sameer Bakhshi, Tapas Chandra Nag, Chanda Panwar, Seema Sen, Seema Kashyap","doi":"10.1016/j.labinv.2024.102167","DOIUrl":"https://doi.org/10.1016/j.labinv.2024.102167","url":null,"abstract":"<p><p>High pigmentation and the abundance of M2 macrophages have been identified as negative predictors in uveal melanoma (UM). Risk factors associated with UM that are prevalent in high-risk white populations are still present, though less common, in relatively low-risk Asian population. Research shows that proangiogenic M2 macrophages and monosomy 3 play a significant role in UM progression. Our aim is to investigate the impact of tumor-associated macrophages in UM and examine their correlation with monosomy 3 & pigmentation. TEM was used to analyze the morphology of macrophages in UM. Forty UM samples underwent FISH for monosomy 3 identification. Immunohistochemistry was done to assess M2/M1 macrophages on 82 UM tissue samples. IL-10 and IL-12 expression was quantified in UM serum samples by ELISA. Expression of all markers was correlated with pigmentation markers (TYRP1, TYRP2, SILV & MITF). Prognostic outcomes were determined using the Cox proportional hazard model & log-rank test. Increased expression of M2/M1 macrophages was observed in 31 UM cases, which correlated with high expression of pigmentation markers. IL-10 concentration was high in UM cases. Monosomy 3 was evident in 50% of UM cases and significantly associated with increased immunoexpression of M2/M1 macrophages and pigmentation markers. Reduced MFS was observed in UM patients with high M2/M1 macrophage expression (p=0.001). High pigmentation and increased M2 macrophage density could impact the tumor microenvironment in UM. This could contribute to ineffective antitumor immune responses in UM patients. Our findings suggest avenues for developing novel therapeutic approaches to counteract these immunosuppressive effects in UM.</p>","PeriodicalId":17930,"journal":{"name":"Laboratory Investigation","volume":null,"pages":null},"PeriodicalIF":5.1,"publicationDate":"2024-11-02","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142567313","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Nup210 Promotes Colorectal Cancer Progression by Regulating Nuclear Plasma Transport","authors":"","doi":"10.1016/j.labinv.2024.102149","DOIUrl":"10.1016/j.labinv.2024.102149","url":null,"abstract":"<div><div>The nuclear pore complex (NPC) regulates nucleoplasmic transport, transcription, and genomic integrity in eukaryotic cells. However, little is known about how NPC works in cancer. In this study, we investigated the role of the nuclear pore protein 210 (Nucleoporin 210, Nup210) in colorectal cancer (CRC). Bioinformatics analysis revealed that the expression of Nup210 was increased in CRC and was associated with poor patient prognosis, but it was not a statistically significant independent prognostic factor. Moreover, knockdown of Nup210 in CRC cells inhibited the proliferation, invasion, and metastasis of CRC cells in vivo and in vitro. Additionally, nuclear size and nuclear plasma material transport capacity decreased along with the number and density of NPCs on the surface of CRC cells when Nup210 expression was inhibited. Furthermore, Nup210 required nuclear localization sequences (NLS) to localize to the nuclear membrane surface and interact with importin-α/β, which in turn affected the transit of nuclear plasma material. Importazole, a small molecule inhibitor of importin, along with therapy that targets the Nup210 protein is anticipated to be a novel strategy for CRC treatment. Their combination may be able to more effectively lower CRC tumor load. In conclusion, Nup210 modulates cellular nucleoplasmic transport capability and cell surface NPC density via NLS, thus promoting CRC progression. This discovery validates the molecular function of NPC in the development of CRC and provides a theoretical foundation for NPC-regulated nuclear import targeting as a therapeutic strategy for CRC.</div></div>","PeriodicalId":17930,"journal":{"name":"Laboratory Investigation","volume":null,"pages":null},"PeriodicalIF":5.1,"publicationDate":"2024-11-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142406581","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Epigenomic and Transcriptomic Profiling of Solitary Fibrous Tumors Identifies Site-Specific Patterns and Candidate Genes Regulated by DNA Methylation","authors":"","doi":"10.1016/j.labinv.2024.102146","DOIUrl":"10.1016/j.labinv.2024.102146","url":null,"abstract":"<div><div>A solitary fibrous tumor (SFT) is a rare mesenchymal neoplasm that can arise at any anatomical site and is characterized by recurrent <em>NAB2::STAT6</em> fusions and metastatic progression in 10% to 30%. The cell of origin has not been identified. Despite some progress in understanding the contribution of heterogeneous fusion types and secondary mutations to SFT biology, epigenetic alterations in extrameningeal SFT remain largely unexplored, and most sarcoma research to date has focused on the use of methylation profiling for tumor classification. We interrogated genome-wide DNA methylation in 79 SFTs to identify informative epigenetic changes. RNA-seq data from targeted panels and data from the Cancer Genome Atlas (TCGA) were used for orthogonal validation of selected findings. In unsupervised clustering analysis, the top 500 most variable cytosine-guanine sites segregated SFTs by primary anatomical site. Differentially methylated genes associated with the primary SFT site included <em>EGFR</em>; <em>TBX15</em>; multiple <em>HOX</em> genes; and their cofactors <em>EBF1</em>, <em>EBF3</em>, and <em>PBX1</em>; as well as <em>RUNX1</em> and <em>MEIS1</em>. Of the 20 DMGs interrogated on the RNA-seq panel, 12 were significantly differentially expressed according to site. However, except <em>TBX15</em>, most of these also showed differential expression according to <em>NAB2::STAT6</em> fusion type, suggesting that the fusion oncogene contributes to the transcriptional regulation of these genes. Transcriptomic data confirmed an inverse correlation between gene methylation and the expression of <em>TBX15</em> in both SFT and TCGA sarcomas<em>. TBX15</em> also showed differential mRNA expression and 5′ UTR methylation between tumors in different anatomical sites in TCGA data. In all analyses, <em>TBX15</em> methylation and mRNA expression retained the strongest association with tissue of origin in SFT and other sarcomas, suggesting a possible marker to distinguish metastatic tumors from new primaries without genomic profiling. Epigenetic signatures may further help to identify SFT progenitor cells at different anatomical sites.</div></div>","PeriodicalId":17930,"journal":{"name":"Laboratory Investigation","volume":null,"pages":null},"PeriodicalIF":5.1,"publicationDate":"2024-11-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142365739","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Highly-Multiplexed Immunofluorescence PhenoCycler Panel for Murine FFPE Yields Insight into Tumor Microenvironment Immunoengineering.","authors":"Sachin S Surwase, Xin Ming M Zhou, Kathryn M Luly, Qingfeng Zhu, Robert A Anders, Jordan J Green, Stephany Y Tzeng, Joel C Sunshine","doi":"10.1016/j.labinv.2024.102165","DOIUrl":"https://doi.org/10.1016/j.labinv.2024.102165","url":null,"abstract":"<p><p>Spatial proteomics profiling is an emerging set of technologies that has the potential to elucidate the cell types, interactions, and molecular signatures that make up complex tissue microenvironments, with applications in the study of cancer, immunity, and much more. An emerging technique in the field is Co-Detection-by-indEXing (CODEX), recently renamed as the PhenoCycler system. This is a highly-multiplexed immunofluorescence imaging technology that relies on oligonucleotide-barcoded antibodies and cyclic immunofluorescence to visualize many antibody markers in a single specimen while preserving tissue architecture. Existing PhenoCycler panels are primarily designed for fresh-frozen tissues. Formalin-fixed paraffin-embedded (FFPE) blocks offer several advantages in preclinical research, but few antibody clones have been identified in this setting for PhenoCycler imaging. Here, we present a novel PhenoCycler panel of 28 validated antibodies for murine FFPE tissues. We describe our workflow for selecting and validating clones, barcoding antibodies, designing our panel, and performing multiplex imaging. We further detail our analysis pipeline for comparing marker expressions, clustering and phenotyping single-cell proteomics data, and quantifying spatial relationships. We then apply our panel and analysis protocol to profile the effects of three gene-delivery nanoparticle formulations, in combination with systemic anti-PD1, on the murine melanoma tumor immune microenvironment. Intralesional delivery of genes expressing the costimulatory molecule 4-1BBL and the cytokine IL-12 led to a shift towards intratumoral M1 macrophage polarization and promoted closer associations between intratumoral CD8 T cells and macrophages. Delivery of IFNγ, in addition to 4-1BBL and IL-12, further increased markers of antigen presentation on tumor cells and intratumoral antigen-presenting cells but also promoted greater expression of checkpoint marker PD-L1 and closer associations between intratumoral CD8 T cells and PD-L1-expressing tumor cells. These findings help to explain the benefits of 4-1BBL and IL-12 delivery while offering additional mechanistic insights into the limitations of IFNγ therapeutic efficacy.</p>","PeriodicalId":17930,"journal":{"name":"Laboratory Investigation","volume":null,"pages":null},"PeriodicalIF":5.1,"publicationDate":"2024-10-25","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142567314","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Concordance of whole slide imaging and conventional light microscopy for assessment of pathologic response following neoadjuvant therapy for lung cancer.","authors":"Julie Stein Deutsch, Daphne Wang, Krista Y Chen, Ashley Cimino-Mathews, Elizabeth D Thompson, Jaroslaw Jedrych, Robert A Anders, Edward Gabrielson, Peter B Illei, Sonali Uttam, Alexa Fiorante, Emily Cohen, Michael Fotheringham, Logan L Engle, Joel C Sunshine, Hao Wang, Dimple Pandya, Vipul Baxi, Joseph Fiore, Kurex Sidik, James Pratt, Alexander S Baras, Tricia R Cottrell, Janis M Taube","doi":"10.1016/j.labinv.2024.102166","DOIUrl":"https://doi.org/10.1016/j.labinv.2024.102166","url":null,"abstract":"<p><p>Pathologic response is an endpoint in many ongoing clinical trials for neoadjuvant regimens, including immune checkpoint blockade and chemotherapy. Whole slide scanning of glass slides generates high resolution digital images and allows for remote review and potential measurement with image analysis tools, but concordance of pathologic response assessment on digital scans compared to glass slides has yet to be evaluated. Such a validation goes beyond previous concordance studies which focused on establishing surgical pathology diagnoses, as it requires quantitative assessment of tumor, necrosis, and regression. Further, as pathologic response assessment is being used as an endpoint, such concordance studies have regulatory implications. The purpose of this study was two fold: firstly, to determine the concordance between pathologic response assessed on glass slides and on digital scans; and secondly, to determine if pathologists benefited from using measurement tools when determining pathologic response. To that end, H&E-stained glass slides from 64 non-small cell lung carcinoma specimens were visually assessed for percent residual viable tumor (%RVT). The sensitivity and specificity for digital vs. glass reads of complete pathologic response (pCR, 0% RVT) and major pathologic response (MPR, ≤10% RVT) were all >95%. When %RVT was considered as a continuous variable, intraclass correlation coefficient of digital vs. glass reads was 0.94. The visual assessments of pathologic response were supported by pathologist annotations of residual tumor and tumor bed areas. In a separate subset of H&E-stained glass slides, several measurement approaches to quantifying %RVT were performed. Pathologist estimates strongly reflected measured %RVT. This study demonstrates the high level of concordance between glass slides evaluated using light microscopy and digital whole slide images for pathologic response assessments. Pathologists did not require measurement tools to generate robust %RVT values from slide annotations. These findings have broad implications for improving clinical workflows and multisite clinical trials.</p>","PeriodicalId":17930,"journal":{"name":"Laboratory Investigation","volume":null,"pages":null},"PeriodicalIF":5.1,"publicationDate":"2024-10-24","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142503126","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Tumor necrosis factor-α-dependent inflammation upregulates high mobility group box 1 to induce tumor promotion and anti-programmed cell death protein-1 immunotherapy resistance in lung adenocarcinoma.","authors":"Lifei Kang, Jingjing Cao, Wenli Guo, Xiaohui Cui, Yangxuan Wei, Jiayu Zhang, Feiran Liu, Chenyang Duan, Qiang Lin, Ping Lv, Zhiyu Ni, Jing Zuo, Haitao Shen","doi":"10.1016/j.labinv.2024.102164","DOIUrl":"https://doi.org/10.1016/j.labinv.2024.102164","url":null,"abstract":"<p><p>Tumor-associated chronic lung inflammation depends on tumor necrosis factor (TNF)-α to activate several cytokines as part of an inflammatory loop, which plays a critical role in tumor progression in lung adenocarcinoma. High mobility group box 1 (HMGB1) is a cytokine that mediates inflammation. Whether TNF-α-induced inflammation regulates HMGB1 to contribute to tumor progression and promotion in lung adenocarcinoma remains unclear. Thus, human samples and a urethane-induced inflammation-driven lung adenocarcinoma (IDLA) mouse model were used to explore the involvement of HMGB1 in tumorigenesis, tumor progression, and efficacy of anti-programmed cell death protein (PD)-1 immunotherapy. High levels of HMGB1 were observed in human lung adenocarcinoma associated with poor overall survival in patients. HMGB1 upregulation was positively correlated with TNF-α-related inflammation and TIM3⁺ infiltration. TNF-α upregulated intracellular and extracellular HMGB1 expression to contribute to tumor promotion in A549 cells in vitro. Using a urethane-induced IDLA mouse model, we found HMGB1 upregulation was associated with increased TIM3⁺ T cell infiltration. Blocking TNF-α-dependent inflammation downregulated HMGB1 expression and inhibited tumorigenesis in the IDLA. Anti-PD-1 treatment alone did not inhibit tumor growth in the TNF-α-dependent IDLA, whereas anti-PD-1 combined with TNF-α blockade overcame anti-PD-1 immunotherapy resistance. Furthermore, anti-PD-1 combined with anti-HMGB1 also inhibited tumor growth in IDLA, suggesting increased HMGB1 release by TNF-α contributes to the resistance of anti-PD-1 immunotherapy in IDLA. Thus, tumor-associated TNF-α-dependent inflammation upregulated intracellular and extracellular HMGB1 expression in an inflammatory loop, contributing to tumor promotion and anti-PD-1 immunotherapy resistance in lung adenocarcinoma.</p>","PeriodicalId":17930,"journal":{"name":"Laboratory Investigation","volume":null,"pages":null},"PeriodicalIF":5.1,"publicationDate":"2024-10-24","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142503128","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Real-world performance of integrative clinical genomics in pediatric precision oncology.","authors":"Petra Pokorna, Hana Palova, Sona Adamcova, Robin Jugas, Dagmar Al Tukmachi, Michal Kyr, Dana Knoflickova, Katerina Kozelkova, Vojtech Bystry, Sona Mejstrikova, Tomas Merta, Karolina Trachtova, Eliska Podlipna, Peter Mudry, Zdenek Pavelka, Viera Bajciova, Pavel Tinka, Marie Jarosova, Tina Catela Ivkovic, Sibylle Madlener, Karol Pal, Natalia Stepien, Lisa Mayr, Boris Tichy, Klara Drabova, Marta Jezova, Sarka Kozakova, Jitka Vanackova, Lenka Radova, Karin Steininger, Christine Haberler, Johannes Gojo, Jaroslav Sterba, Ondrej Slaby","doi":"10.1016/j.labinv.2024.102161","DOIUrl":"https://doi.org/10.1016/j.labinv.2024.102161","url":null,"abstract":"<p><p>Despite significant improvement in the survival of pediatric cancer patients, treatment outcomes for high-risk, relapsed, and refractory cancers remain unsatisfactory. Moreover, prolonged survival is frequently associated with long-term adverse effects due to intensive multimodal treatments. Accelerating the progress of pediatric oncology requires both therapeutic advances and strategies to mitigate the long-term cytotoxic side effects, potentially through targeting specific molecular drivers of pediatric malignancies. In this report, we present the results of integrative genomic and transcriptomic profiling of 230 patients with malignant solid tumors (the \"primary cohort\") and 18 patients with recurrent or otherwise difficult-to-treat nonmalignant conditions (the \"secondary cohort\"). The integrative workflow for the primary cohort enabled the identification of clinically significant single-nucleotide variants, small insertions/deletions, and fusion genes, which were found in 55% and 28% of patients, respectively. For 38% of patients, molecularly informed treatment recommendations were made. In the secondary cohort, known or potentially driving alteration was detected in 89% of cases, including a suspected novel causal gene for patients with inclusion body infantile digital fibromatosis. Furthermore, 47% of findings also brought therapeutic implications for subsequent management. Across both cohorts, changes or refinements to the original histopathological diagnoses were achieved in 4% of cases. Our study demonstrates the efficacy of integrating advanced genomic and transcriptomic analyses to identify therapeutic targets, refine diagnoses, and optimize treatment strategies for challenging pediatric and young adult malignancies and underscores the need for broad implementation of precision oncology in clinical settings.</p>","PeriodicalId":17930,"journal":{"name":"Laboratory Investigation","volume":null,"pages":null},"PeriodicalIF":5.1,"publicationDate":"2024-10-21","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142503127","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Analysis of gliomas DNA methylation: Assessment of pre-analytical variables.","authors":"Karol Bomsztyk, Daniel Mar, Oleg Denisenko, Suzanne Powell, Monika Vishnoi, Zheng Yin, Jennifer Delegard, Caroline Hadley, Nitin Tandon, Akash Patel, Anoop Patel, Richard G Ellenbogen, Rohan Ramakrishna, Robert Rostomily","doi":"10.1016/j.labinv.2024.102160","DOIUrl":"https://doi.org/10.1016/j.labinv.2024.102160","url":null,"abstract":"<p><p>Precision oncology is driven by biomarkers. For glioblastoma multiforme (GBM), the most common malignant adult primary brain tumor, O^6-methylguanine-DNA methyltransferase (MGMT) gene promoter methylation is an important prognostic and treatment clinical biomarker. Time consuming pre-analytical steps such as biospecimen storage, fixation, sampling, and processing are sources of data irreproducibility, and all these pre-analytical variables are confounded by intratumor heterogeneity of MGMT promoter methylation. To assess the effect of pre-analytical variables on GBM DNA methylation, tissue storage/sampling (CryoGrid), sample preparation multi-sonicator (PIXUL), and 5-methylcytosine (5mC) DNA immunoprecipitation (Matrix MeDIP-qPCR/seq) platforms were used. MGMT promoter methylation status assayed by MeDIP-qPCR was validated with methylation specific PCR (MS-PCR). MGMT promoter methylation levels in frozen and formalin fixed paraffin embedded (FFPE) sample pairs were not statistically different, confirming reliability of FFPEs for MGMT promoter methylation analysis. Warm ex-vivo ischemia (up to 4hrs at 37^oC) and 3 cycles of repeated sample thawing and freezing did not statistically impact 5mC at MGMT promoter, exon, and enhancer regions, indicating the resistance of DNA methylation to common variations in sample processing conditions that might be encountered in research and clinical settings. 26-34% of specimens exhibited intratumor heterogeneity in the MGMT DNA promoter methylation. These data demonstrate that variations in sample fixation, ischemia duration and temperature, and DNA methylation assay technique do not have a statistically significant impact on MGMT promoter methylation assessment. However, intratumor methylation heterogeneity underscores the value of multiple biopsies at different GBM geographic tumor sites in the evaluation of MGMT promoter methylation status. Matrix-MeDIP-seq analysis revealed that MGMT promoter methylation status clustered with other differentially methylated genomic loci (e.g. HOXA and lncRNAs) that are resilient to variation in the above pre-analytical conditions. These observations offer new opportunities to develop more granular data-based epigenetic GBM biomarkers. In this regard, the high throughput CryoGrid-PIXUL-Matrix toolbox could be useful.</p>","PeriodicalId":17930,"journal":{"name":"Laboratory Investigation","volume":null,"pages":null},"PeriodicalIF":5.1,"publicationDate":"2024-10-17","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142468980","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Immunoproteomics reveal different characteristics for the prognostic markers of intratumoral-infiltrating CD3+ T lymphocytes and immunoscore in colorectal cancer.","authors":"Saiyan Ji, Huanying Fang, Jingjie Guan, Kun He, Qingyuan Yang","doi":"10.1016/j.labinv.2024.102159","DOIUrl":"https://doi.org/10.1016/j.labinv.2024.102159","url":null,"abstract":"<p><p>Tumor-infiltrating lymphocytes (TILs) and immunoscoring based on densities of CD3+ and CD8+ TILs are both favorable prognostic markers in colorectal cancer (CRC). However, determination of the molecular features of TILs, particularly their immunoproteomic signatures would require the development of large-scale in situ spatiotemporal technologies. Recently, a multiplex in situ digital spatial proteomic profiling (DSP) tool GeoMx DSP has been applied to identify biomarkers predictive of therapeutic responses and to understand disease mechanisms and progression. Taking advantage of this tool, we simultaneously characterized the spatial distribution and interactions of 42 immune proteins in tumor cells (TC), CD3+ T stromal TILs (sTILs), and CD20+ B sTILs using tissue microarrays, and further studied their associations with CD3+ T TILs and immunoscores in CRC. First, our data showed that well-known immune checkpoints, such as PD-L1, PD-L2, and LAG3, were expressed at low levels, whereas some other immune proteins, such as CD11c, CD68, STING, and CD44, were highly expressed. Second, eight spatial interactions were identified, including five interactions between TC and CD20+ B sTILs, two interactions between CD3+ T sTILs and CD20+ B sTILs, and one interaction among TC, CD3+ T sTILs, and CD20+ B sTILs. Third, the differential immune microlandscape in the spatial compartments was identified in tissues with positive CD3+ T intratumoral TILs and high immunoscores. Collectively, our study is the first to provide in situ spatial immune characteristics at the proteomic level. Moreover, our findings provide direct evidence supporting the infiltration of CD3+ T sTILs from stoma to TC and shed important insights into better understanding and treating CRC patients related to different immune prognostic markers.</p>","PeriodicalId":17930,"journal":{"name":"Laboratory Investigation","volume":null,"pages":null},"PeriodicalIF":5.1,"publicationDate":"2024-10-15","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142468981","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}