bluuebunny/crossref_metadata_2025_split huggingface dataset:
Clinical significance and biomarker potential of MGMT protein measurement in colorectal cancer.
Authors: Zhang Janie Yue , Cecchini Michael , Desai Shruti S. , Schalper Kurt A. 
Published: January 2022

136 Background: O6-methylguanine DNA methyltransferase (MGMT) is the principal repair mechanism of DNA damage created by alkylating agents. Approximately 40% of colorectal cancers (CRC) exhibit MGMT promoter hypermethylation, expected to result in gene silencing. The role of MGMT as a biomarker is poorly understood, and quantitative measurement of MGMT protein has not been performed in CRC. We hypothesized that altered tumor MGMT would affect DNA damage response, modulate adaptive anti-tumor immune responses, and influence survival in CRC. Methods: We used multiplexed quantitative immunofluorescence (QIF) to study 4 clinically annotated retrospective patient cohorts treated from 2000 to 2017 at Yale, consisting of 400 formalin-fixed paraffin-embedded CRC cases represented in tissue microarray format. These included paired tumors and adjacent non-tumor colonic mucosa (n = 112 pairs in Cohort 1), paired primary CRC and lymph node metastases (n = 31 pairs in Cohort 2), and 2 cohorts with primary stage I-IV tumors (n = 250 in Cohort 3; n = 150 in Cohort 4). We established a QIF panel for simultaneous, localized measurement of DAPI (all cells), cytokeratin (CK; tumor epithelial cells), MGMT, γH2AX (DNA damage response), and CD8. The measurement of fluorescent signals in CK-positive tumor cells or CK-negative stromal cells was achieved using co-localization strategies and the AQUA QIF platform. Results: We identified lower MGMT protein levels in CRC cells than in non-tumor colonic epithelium (p < 0.0001), and in lymph node metastases compared to paired primary CRC tumors (p = 0.0411). Using the visual detection threshold, tumor selective MGMT protein downregulation was identified in 20% of cases in Cohorts 3 and 4. Microsatellite instability-high (MSI-H) cases showed decreased tumoral MGMT protein compared to microsatellite stable (MSS) cases (p = 0.002). In Cohorts 3 and 4, low tumoral MGMT was consistently associated with increased CD8+ tumor infiltrating lymphocytes (p = 0.0472; p = 0.0002). The association between MGMT and γH2AX was inconsistent. The association between MGMT and CD8 was significant only in MSS cases (p = 0.0001) but not in MSI-H cases (p = 0.0979), supporting that the effect is not driven by MSI-H status. Tumor-selective MGMT downregulation evidenced by a low tumor-to-stroma ratio was associated with improved progression-free survival and overall survival in both Cohorts 3 and 4, but statistical significance was only seen in Cohort 3. Conclusions: MGMT protein downregulation occurs in 20% of CRCs and is associated with increased adaptive anti-tumor immune responses, mismatch repair (MMR) deficiency and better prognosis. MGMT deficiency may alter DNA repair in tumor cells and mediate the accumulation of antigenic mutations or neopeptides, independent from MMR status. Our results support a biomarker role of MGMT protein and suggest a role for immunotherapy combinations in MGMT deficient tumors.
Clinical significance of Bacteroides fragilis as potential prognostic factor in colorectal cancer patients.
Authors: Matsumiya Yuriko , Suenaga Mitsukuni , Ishikawa Toshiaki , Hanaoka Marie , Iwata Noriko , Masuda Taiki , Yamauchi Shinichi , Tokunaga Masanori , Kinugasa Yusuke 
Published: January 2022

137 Background: Bacteroides fragilis ( B. fragilis) is an obligate anaerobe and generally acts as anti-inflammatory manner on the intestinal tract. Enterotoxigenic Bacteroides fragilis (ETBF), a subtype of B. fragilis, produces Bacteroides fragilis toxin (BFT) leading to either asymptomatic chronic colonic inflammation or colonic tumor formation. However, the impact of B. fragilis and ETBF on colorectal cancer (CRC) prognoses still remains unclear. We tested whether the presence of B. fragilis and ETBF affect clinical outcome in CRC patients who underwent curative surgery. Methods: We obtained 197 pairs of matched FFPE samples from colorectal cancerous and adjusted non-cancerous tissues of patients with stage II and III CRC who underwent curative resection between 2014 and 2016. Quantitative analyses of B. fragilis and ETBF in the colon tissues were performed using quantitative PCR with primers, 16S rRNA for B. fragilis and bft DNA, respectively. Recurrence-free survival (RFS) and overall survival (OS) were analyzed using Kaplan-Meier curves, log-rank test, and Cox proportional hazards regression. Results: Among 197 patients, 16S rRNA for B. fragilis DNA and bft DNA were detected in 120 patients (60.9%) and 12 patients (6.1%), respectively. B. fragilis-positive patients had better RFS (5-y RFS rate: 81.4% vs. 73.4%, HR0.59, 95% CI: 0.31-1.12, p =0.10) and OS (5-y OS rate: 88.9% vs 78.3%, HR0.53, 95% CI: 0.26-1.11, p =0.091) compared with B. fragilis-negative patients though statistically not significant. In multivariable analysis for RFS, B. fragilis-positive remained as an independent prognostic factor (HR0.53, 95% CI: 0.28-0.99, p =0.049) along with tumor depth T4 and Stage III, while there was no significance in OS. No significant differences were observed between ETBF and nontoxigenic B. fragilis in patients' characteristics and clinical outcomes. Conclusions: Our findings suggest that the presence of B. fragilis may predict outcome especially RFS in patients with curatively resected stage II and III CRC. Further research are warranted to explore whether B. fragilis status could be involved in a novel prediction model for outcome in early-stage CRC and develop probiotics treatments to prevent recurrence.
Deep learning analysis of the adipose tissue and the prediction of prognosis in colorectal cancer.
Authors: Lin Anqi , Zhang Jian , Luo Peng 
Published: January 2022

139 Background: Colorectal cancer (CRC) is the third most common type of cancer and has a poor prognosis and high recurrence rate. Research has shown that the lipid microenvironment surrounding tumors is closely associated with the occurrence, development, and metastasis of CRC. Recently, advances in artificial intelligence have greatly improved the accuracy of models for CRC prognosis and survival analysis. Methods: According to pathological images from the National Center for Tumor diseases(NCT), the University Medical Center Mannheim(UMM) database and the ImageNet data set, a model called VGG19 was pre-trained. A deep convolutional neural network(CNN), VGG19CRC, was trained by the migration learning method. According to the VGG19CRC model, adipose tissue scores were calculated for TCGA-CRC hematoxylin and eosin(H&E) images and images from patients at Zhujiang Hospital of Southern Medical University and First People’s Hospital of Chenzhou. Kaplan-Meier(KM) analysis was used to compare the overall survival(OS) of patients. The XCell and MCP-Counter algorithms were used to evaluate the immune cell scores of the patients. Gene set enrichment analysis(GSEA) and single-sample GSEA(ssGSEA) were used to analyze upregulated and downregulated pathways. Results: In TCGA-CRC, patients with high-adipocytes(high-ADI) CRC had significantly shorter OS times than those with low-ADI CRC. In a validation queue from Zhujiang Hospital of Southern Medical University(Local-CRC1), patients with high-ADI had worse OS than CRC patients with low-ADI. In another validation queue from First People's Hospital of Chenzhou(Local-CRC2), patients with low-ADI CRC had significantly longer OS than patients with high-ADI CRC. In subgroup analysis, ADI could be used as a prognostic marker for patients with colon adenocarcinoma(COAD) and rectum adenocarcinoma(READ), as well as male and female CRC patients. Among these subgroups, patients with lower ADI also had significantly improved OS. Compared with the low-ADI group, high-ADI patients had significantly decreased CD8+ T cells, T cells,and monocytes in the tumor immune microenvironment(TIME), while M2 macrophages were significantly increased. According to the GSEA and ssGSEA analyses, pathways mediating anti-tumor immunity were significantly downregulated in the high-ADI group, while some oncogenic signaling pathways were significantly upregulated. Conclusions: We developed a deep convolution network to segment various tissues from pathological H&E images of CRC and automatically quantify ADI. This allowed us to further analyze and predict the survival of CRC patients according to information from their segmented pathological tissue images, such as tissue components and the tumor microenvironment. Furthermore, we found that ADI may also predict OS in CRC patients and among the subgroups.
The role of germline polymorphisms in genes involved in the antioxidant system to predict the efficacy of cetuximab for patients with metastatic colorectal cancer (mCRC) enrolled in FIRE-3 trial.
Authors: Arai Hiroyuki , Millstein Joshua , Stintzing Sebastian , Wang Jingyuan , Battaglin Francesca , Kawanishi Natsuko , Jayachandran Priya , Soni Shivani , Wu Zhang , Heinemann Volker , Lenz Heinz-Josef 
Published: January 2022

143 Background: Reactive oxygen species activate RAS/MAPK signaling either through inactivation of phosphatases or by direct oxidation of kinases. We hypothesized that functional genetic variants in genes involved in the antioxidant system may be associated with the efficacy of cetuximab in mCRC patients. Methods: We analyzed genomic and clinical data from FIRE-3, a phase III trial which compared cetuximab and bevacizumab in combination with FOLFIRI in untreated mCRC patients. Genomic DNA extracted from blood samples was genotyped using an OncoArray (Illumina, Inc., San Diego, CA, USA). Candidate 13 functional single nucleotide polymorphisms (SNPs) ( TXN rs2301242, TXN rs2301241, TXN2 rs4821494, TXN2 rs9619611, TXN2 rs59841625, CAT rs7943316, CAT rs564250, CAT rs11604331, CAT rs1001179, CAT rs769217, GPX4 rs757229, GPX4 rs4807542, and GPX4 rs713041) were tested for association with progression-free survival and overall survival (OS), using log-rank test and Cox proportional hazards model. To confirm the predictive value, the treatment-by-SNP interaction was tested. Results: A total of 236 patients were available for the SNP analyses (cetuximab arm, n = 129; bevacizumab arm, n = 107). In the cetuximab arm, two SNPs were significantly associated with clinical outcomes in univariate analyses: TXN2 rs4821494 (T/T vs any G allele, hazard ratio [HR] on OS = 2.17, 95% confidence interval [CI] = 1.04–4.56, log-rank p = 0.04) and GPX4 rs4807542 (G/G vs any A allele, HR on OS = 2.04, 95% CI = 1.05–3.98, log-rank p = 0.03). Multivariate analysis confirmed the significance in TXN2 rs4821494 (T/T vs any G allele, HR on OS = 2.47, 95% CI = 1.06–5.72, p = 0.03). In the bevacizumab arm, univariate analyses did not detect any significant associations between the SNPs and clinical outcomes. Treatment-by-SNP interaction test showed the significant predictive value of TXN2 rs4821494 in terms of OS ( p = 0.03). Conclusions: TXN2 rs4821494 involved in the antioxidant system may predict the efficacy of cetuximab, in comparing with bevacizumab, in the first-line treatment of mCRC. Our novel findings warrant further validation studies.
Encorafenib, cetuximab, and cytotoxic chemotherapy combinations in BRAFV600E CRC murine models.
Authors: Woods Melanie Nicole , De Falco Vincenzo , Prisco Claudia , Martini Giulia , Guerrera Luigi Pio , Belli Valentina , Troiani Teresa , Morris Van K. , Shen John Paul Y.C. , Lee Hey Min , Villarreal Oscar , Sorokin Alexey , Kanikarla Marie Preeti , Ciardiello Fortunato , Kopetz Scott , Napolitano Stefania 
Published: January 2022

145 Background: Based on promising results from latest trials, a crucial point is to evaluate whether encorafenib (E) plus cetuximab (C), alone or in combination with chemotherapy, can improve clinical outcomes relative to current standard of care in previously untreated BRAFV600E mutant mCRC. Considering the high number of BRAFV600E mutant mCRC patients who will never receive a second-line treatment, the rationale of this strategy is to maximize treatment outcome within the first-line setting. Methods: We performed an in vivo study using human BRAFV600E CRC cell line-derived xenografts in nude mice. We evaluated the efficacy of encorafenib (E) + cetuximab (C), FOLFOX, and FOLFIRI, both as individual regimens and in combinations. Mice were treated for 3 weeks and followed for an additional 8 weeks to evaluate durability of tumor control. Additionally, we validated our findings using 3 BRAFV600E mutated patient derived xenografts. Tumors progressing on single agent and combined treatment were profiled by RNA sequencing, protein extraction for RPPA/Western blot, and establishment of in vitro primary cell cultures for further analyses. Results: Our study showed across all 4 models both FOLFOX and FOLFIRI, each in combination with encorafenib plus cetuximab, having greater efficacy than encorafenib plus cetuximab or either chemotherapy alone. No significant change in toxicity was seen with the addition of chemotherapy. Interestingly, in the one model with long term treatment, FOLFOX + E +/- C performed greatest over the long-term, with significant endpoint separation against all other treatment arms (P < 0.05). Conclusions: Taken together, results from our study suggest that the addition of chemotherapy to BRAF+EGFR targeted therapy can further increase the magnitude of response in BRAFV600E mCRC and is a promising combination now being explored clinically. Additionally, this research will substantially contribute to our understanding of the genetic and molecular bases of resistance to target therapies and chemo-based approach in BRAFV600Econtext.
Gene expression-based personalized prescription of targeted drugs in colorectal cancer.
Authors: Poddubskaya Elena , Sorokin Maxim , Zolotovskaya Marianna , Suntsova Maria , Garazha Andrew , Moisseev Alexey , Bondarenko Alexey , Skrijalin Vasiliy , Baranova Madina , Sekacheva Marina , Seryakov Alexander , Buzdin Anton 
Published: January 2022

151 Background: Colorectal cancer (CRC) is the fourth most common cancer worldwide with relatively poor patient survival. Transcriptome assay could be used to personalize CRC treatment thus complementing standard mutation analysis. Methods: We performed retrospective hybrid experimental and meta-analysis of CRC patient gene expression data with available progression-free survival (PFS) information and/or targeted drug response status. In total we analyzed 243 gene expression profiles from four publicly available (TCGA and three datasets from Gene Expression Omnibus GSE19860, GSE19862, GSE104645), and one experimental (PRJNA663280) patient cohorts. Each gene expression profile was analyzed using bioinformatic second-opinion platform Oncobox to calculate balanced drug efficiency scores (BES) to build personalized ratings of potentially effective targeted drugs. Area under the ROC curve (AUC) metric and Cox regression analysis were used to assess Oncobox capacity to predict tumor response and PFS, respectively. Results: Patients from GSE19860 (n = 12), GSE19862 (n = 14), GSE104645 (n = 81) received bevacizumab as monotherapy or in combination with chemotherapy as the nearest line of treatment after biopsy collection. Oncobox correctly classified treatment responders vs non-responders with AUC 0.94, 0.90 and 0.84, respectively. BES value was strongly associated with PFS (HR = 0.53, CI 0.33-0.84, log-rank test p-value 0.0057) in the GSE104645 cohort. However, BES was ineffective for predicting response and PFS after second-line (after biopsy collection) treatment with cetuximab. BES also predicted treatment response with AUC 0.74 in the TCGA cohort (n = 17) treated with 4 different targeted drugs. Thirty clinical outcomes were collected for 14 patients from our experimental cohort PRJNA663280. Patients were treated with 10 different targeted drugs. BES was an effective biomarker that could predict treatment outcomes with AUC 0.74 for all lines of therapy and 0.94 for the first line therapy (after biopsy), and could predict PFS after first-line treatment (HR 0.14, CI 0.027-0.73, log-rank test p-value 0.0091). Conclusions: Our results suggest that RNA profiling in tumor samples may be helpful for personalizing prescriptions of targeted therapeutics in CRC. Using recent biopsies is essential to obtain robust estimates of targeted drugs efficacy.
Transcriptomic profiling to identify subsets of immune hot locally advanced rectal adenocarcinomas with favorable outcomes after neoadjuvant treatment.
Authors: Sanchez Vega Francisco , Chatila Walid Khaled , Kim Jin Ki , Walch Henry S. , Marco Michael , Chen Ching-Tung , Wu Fan , Khalil Danny , Ganesh Karuna , Qu Xuan , Luthra Anisha , Choi Seo-Hyun , Ho Yu-jui , Omer Dana , Shia Jinru , Romesser Paul Bernard , Schultz Nikolaus , Yaeger Rona , Smith Jesse Joshua , Garcia-Aguilar Julio 
Published: January 2022

155 Background: Understanding the role of the tumor microenvironment in the response to chemotherapy and radiation in patients with locally advanced rectal cancer (LARC, stage II-III) can lead to the identification of novel immunologic biomarkers to preselect patients who can avoid surgery and benefit from watch-and-wait strategies. Methods: We performed DNA and RNA sequencing of pre-treatment biopsies from 89 LARC patients who received neoadjuvant therapy, including 5 microsatellite unstable (MSI) and 84 microsatellite stable (MSS) patients. We computed single-sample gene set enrichment analysis (ssGSEA) scores for immune infiltrates and signaling pathways implicated in tumor progression. Immunofluorescence and hematoxylin-eosin staining of tumor slides was performed to confirm significant correlations with RNA-Seq estimates of immune markers. Other genomic variables were also included in the analysis, such as tumor mutational burden (TMB), fraction of genome altered by copy number changes, whole genome duplication events and somatic mutations in rectal cancer driver genes and pathways. Results were largely replicated using an independent cohort of 42 LARC samples with publicly available data from The Cancer Genome Atlas (TCGA). Results: Since MSI tumors are known to have a distinct immunologic profile, we separated them into their own group and performed unsupervised hierarchical clustering on the MSS tumors. We identified a set of immune hot MSS tumors (n = 7) with extensive immune infiltration. These tumors had low TMB and were predominantly classified as CMS4 (5/7). None of the 12 patients in the combined MSI and immune hot MSS groups recurred during the length of our study and they had response rates > 50% (vs. < 25% in the rest of MSS patients). MSI and immune hot MSS tumors had lower frequency of TP53 and APC mutations, and they exhibited increased levels of T cell infiltration. In particular, we observed overexpression of markers for Th1 cells, which produce inflammatory cytokines (e.g., IFN-gamma) and are associated with antitumor immunity. Genes encoding protein targets of immune checkpoint blockade, such as PDCD1 (PD-1), CD274 (PD-L1), CTLA4, HAVCR2 (TIM3) and LAG3, were also overexpressed in the immune hot MSS and - to a lesser extent – the MSI tumors, suggesting that these patients might benefit from the use of immune checkpoint inhibitors. Conclusions: Our results uncover a unique LARC tumor immune profile evident in the pre-treatment setting that could be used to better prognosticate rectal cancer patients and develop novel therapeutic strategies.
CLK2 upregulation and outcomes for males with colorectal adenocarcinoma.
Authors: Siddiqui Zain I. , Sun Chenyu , Tuason John Pocholo W. , Lising Jannell F. , Paudel Arpana , Ahmed Marwan K. , Kim Na Hyun , Bhan Chandur , Memon Burhan , Mohammed Abdul Humaed , Prasad Apurwa 
Published: January 2022

159 Background: Colorectal adenocarcinoma (COAD) is a common cancer in gastrointestinal tract. CDC-like kinase (CLK) family, containing four characterized isoforms (CLK1-4), have been reported for their roles in precursor-mRNA splicing. However, studies on their roles in COAD are limited. Thus, this study was performed to explore the roles of CLK family members in COAD. Methods: RNA-sequencing FPKM data and corresponding clinical information of 41 normal tissues and 480 tumor tissues of COAD were obtained from The Cancer Genome Atlas (TCGA). Then expression differences of CLK family between the normal tissues and COAD cancer tissues were compared with Wilcoxon rank-sum test via ‘limma’ package. Overall survival (OS) of upregulated CLK family members was analyzed by Kaplan–Meier (K–M) method via ‘survminer’ package. Subgroup analyses of different genders were also conducted. The best discriminate cut-off point between the high and low expression groups was assessed by the receiver operating characteristic (ROC) curve and area under the curve (AUC) values for upregulated CLK family members via ‘pROC’ package to assess their diagnostic values. Results: The expression of CLK1 and CLK2 were upregulated in tumors compared with normal tissues (p < 0.001), while CLK3 were downregulated in tumor tissues (p = 0.02) and no statically significant difference was found regarding CLK4 (p = 0.25). Higher expression of CLK1 was not associated with a change of OS (HR: 1.24, 95%CI: 0.84-1.83, p = 0.279), while higher expression of CLK2 is associated with worse OS (HR: 2.03, 95%CI: 1.34-3.06, p = 0.001). Subgroup analysis found that higher expression of CLK2 was associated with worse OS (HR: 2.28, 95%CI 1.27-4.08, p = 0.006) in males, but not in females (HR: 1.72, 95%CI 0.95-3.10, P = 0.072). Further analysis of ROC curve shown that AUC of CLK1 and CLK2 were 0.748 (0.687-0.809) and 0.884 (0.840-0.928), respectively. Conclusions: CLK1 and CLK2 are upregulated in COAD tumor samples, and higher expression of CLK2 in male COAD patients is associated worse OS. This demonstrated the potential therapeutic value of CLK2 in male patients with COAD. ROC curve indicated the potential diagnostic value of CLK1 and CLK2.