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Ovarian cancer is a common gynecologic malignancy with a high fatality rate. Intraperitoneal chemotherapy has been proved as an efficient clinical treatment for disseminated ovarian cancer. However, there are limitations for conventional small molecule drugs to achieve an ideal therapeutic effect. Herein, a synergistic treatment for intraperitoneally disseminated ovarian cancer was achieved by Arg-Gly-Asp (RGD)-modified amorphous calcium phosphate loading with doxorubicin (designated as RGD-CaPO/DOX). The engineered calcium-involved nanomedicine augmented the therapeutic effect of DOX by aggravating endoplasmic reticulum stress, calcium overload, and mitochondrial dysfunction, ultimately triggering mitochondrial apoptosis in the SKOV3 (human ovarian cancer) cell line. In an intraperitoneally disseminated tumor model, RGD modification and the weak negative surface potential of the NPs were beneficial for intraperitoneal retention and tumor targeting. Moreover, intraperitoneal injection of RGD-CaPO/DOX NPs resulted in a favorable antitumor effect. The mean survival time of SKOV3-bearing mice was significantly extended from 29 to 59 days with negligible toxicity. Therefore, this study has been designed to provide an effective chemotherapeutic-augmented treatment for intraperitoneally disseminated ovarian cancer.

PDF file 1227K, This file contains Supplemental Figures 1-7, which provide additional supporting information for each of the main figures

Spearman rank-correlation coefficient (r) between the densities of T-cells

MIR21 expression after adjusting for cellularity in colorectal cancer tissue and statistical analysis

Supplementary Table S22: Segmented b-allele deficits

Ordinal logistic regression analysis to assess the association of tumor PDCD1LG2 (PD-L2) expression (predictor) with the overall lymphocytic reaction score (outcome)

Supplementary Tables S16-S19, describing IHC results on NHS/HPFS data

Supplementary Figure S1. Flow diagram of the study population in the Nurses'' Health Study and the Health Professionals Follow-Up Study. Supplementary Figure S2. Immunohistochemistry of a colorectal carcinoma case using anti-PDCD1LG2 (PD-L2) antibody (A) and its isotype control (B). Supplementary Figure S3. Kaplan-Meier curves for colorectal cancer-specific mortality (A) and overall mortality (B) according to tumor PDCD1LG2 (PD-L2) expression.

The complex interactions between colorectal neoplasia and immune cells in the tumor microenvironment remain to be elucidated. Experimental evidence suggests that microRNA MIR21 (miR-21) suppresses antitumor T-cell–mediated immunity. Thus, we hypothesized that tumor MIR21 expression might be inversely associated with T-cell density in colorectal carcinoma tissue. Using 538 rectal and colon cancer cases from the Nurses' Health Study and the Health Professionals Follow-up Study, we measured tumor MIR21 expression by a quantitative reverse-transcription PCR assay. Densities of CD3+, CD8+, CD45RO (PTPRC)+, and FOXP3+ cells in tumor tissue were determined by tissue microarray immunohistochemistry and computer-assisted image analysis. Ordinal logistic regression analysis was conducted to assess the association of MIR21 expression (ordinal quartiles as a predictor variable) with T-cell density (ordinal quartiles as an outcome variable), adjusting for tumor molecular features, including microsatellite instability; CpG island methylator phenotype; KRAS, BRAF, and PIK3CA mutations; and LINE-1 methylation. We adjusted the two-sided α level to 0.012 for multiple hypothesis testing. Tumor MIR21 expression was inversely associated with densities of CD3+ and CD45RO+ cells (Ptrend < 0.0005). The multivariate odds ratio of the highest versus lowest quartile of MIR21 for a unit increase in quartile categories of CD3+ or CD45RO+ cells was 0.44 [95% confidence interval (CI), 0.28 to 0.68] or 0.41 (95% CI, 0.26–0.64), respectively. Our data support a possible role of tumor epigenetic deregulation by noncoding RNA in suppressing the antitumor T-cell–mediated adaptive immune response and suggest MIR21 as a potential target for immunotherapy and prevention in colorectal cancer. Cancer Immunol Res; 4(1); 33–40. ©2015 AACR.

Figure S1, Table S1-7

Calcium intake has been associated with a lower risk of colorectal cancer. Calcium signaling may enhance T-cell proliferation and differentiation, and contribute to T-cell–mediated antitumor immunity. In this prospective cohort study, we investigated the association between calcium intake and colorectal cancer risk according to tumor immunity status to provide additional insights into the role of calcium in colorectal carcinogenesis. The densities of tumor-infiltrating T-cell subsets [CD3+, CD8+, CD45RO (PTPRC)+, or FOXP3+ cell] were assessed using IHC and computer-assisted image analysis in 736 cancer cases that developed among 136,249 individuals in two cohorts. HRs and 95% confidence intervals (CI) were calculated using Cox proportional hazards regression. Total calcium intake was associated with a multivariable HR of 0.55 (comparing ≥1,200 vs. Ptrend = 0.002) for CD8+ T-cell–low but not for CD8+ T-cell–high tumors (HR = 1.02; 95% CI, 0.67–1.55; Ptrend = 0.47). Similarly, the corresponding HRs (95% CIs) for calcium for low versus high T-cell–infiltrated tumors were 0.63 (0.42–0.94; Ptrend = 0.01) and 0.89 (0.58–1.35; Ptrend = 0.20) for CD3+; 0.58 (0.39–0.87; Ptrend = 0.006) and 1.04 (0.69–1.58; Ptrend = 0.54) for CD45RO+; and 0.56 (0.36–0.85; Ptrend = 0.006) and 1.10 (0.72–1.67; Ptrend = 0.47) for FOXP3+, although the differences by subtypes defined by T-cell density were not statistically significant. These potential differential associations generally appeared consistent regardless of sex, source of calcium intake, tumor location, and tumor microsatellite instability status. Our findings suggest a possible role of calcium in cancer immunoprevention via modulation of T-cell function.

Experimental evidence suggests that the let-7 family of noncoding RNAs suppresses adaptive immune responses, contributing to immune evasion by the tumor. We hypothesized that the amount of let-7a and let-7b expression in colorectal carcinoma might be associated with limited T-lymphocyte infiltrates in the tumor microenvironment and worse clinical outcome. Utilizing the molecular pathological epidemiology resources of 795 rectal and colon cancers in two U.S.-nationwide prospective cohort studies, we measured tumor-associated let-7a and let-7b expression levels by quantitative reverse-transcription PCR, and CD3+, CD8+, CD45RO (PTPRC)+, and FOXP3+ cell densities by tumor tissue microarray immunohistochemistry and computer-assisted image analysis. Logistic regression analysis and Cox proportional hazards regression were used to assess associations of let-7a (and let-7b) expression (quartile predictor variables) with T-cell densities (binary outcome variables) and mortality, respectively, controlling for tumor molecular features, including microsatellite instability, CpG island methylator phenotype, LINE-1 methylation, and KRAS, BRAF, and PIK3CA mutations. Compared with cases in the lowest quartile of let-7a expression, those in the highest quartile were associated with lower densities of CD3+ [multivariate odds ratio (OR), 0.40; 95% confidence interval (CI), 0.23–0.67; Ptrend = 0.003] and CD45RO+ cells (multivariate OR, 0.31; 95% CI, 0.17–0.58; Ptrend = 0.0004), and higher colorectal cancer-specific mortality (multivariate hazard ratio, 1.82; 95% CI, 1.42–3.13; Ptrend = 0.001). In contrast, let-7b expression was not significantly associated with T-cell density or colorectal cancer prognosis. Our data support the role of let-7a in suppressing antitumor immunity in colorectal cancer and suggest let-7a as a potential target of immunotherapy. Cancer Immunol Res; 4(11); 927–35. ©2016 AACR.

Supplementary Tables S1, S3 through S15, S20, and S23

PDF file 146K, This file contains the legends to Supplementary Figures 1-7 and Supplementary methods

S1 XLSX file 63K, This Supplementary Table contains the results of ORF screen to identify genes that can rescue the suppression of TBK1 in a KRAS-dependent cell line

To understand the genetic drivers of immune recognition and evasion in colorectal cancer, we analyzed 1,211 colorectal cancer primary tumor samples, including 179 classified as microsatellite instability–high (MSI-high). This set includes The Cancer Genome Atlas colorectal cancer cohort of 592 samples, completed and analyzed here. MSI-high, a hypermutated, immunogenic subtype of colorectal cancer, had a high rate of significantly mutated genes in important immune-modulating pathways and in the antigen presentation machinery, including biallelic losses of B2M and HLA genes due to copy-number alterations and copy-neutral loss of heterozygosity. WNT/β-catenin signaling genes were significantly mutated in all colorectal cancer subtypes, and activated WNT/β-catenin signaling was correlated with the absence of T-cell infiltration. This large-scale genomic analysis of colorectal cancer demonstrates that MSI-high cases frequently undergo an immunoediting process that provides them with genetic events allowing immune escape despite high mutational load and frequent lymphocytic infiltration and, furthermore, that colorectal cancer tumors have genetic and methylation events associated with activated WNT signaling and T-cell exclusion.Significance: This multi-omic analysis of 1,211 colorectal cancer primary tumors reveals that it should be possible to better monitor resistance in the 15% of cases that respond to immune blockade therapy and also to use WNT signaling inhibitors to reverse immune exclusion in the 85% of cases that currently do not. Cancer Discov; 8(6); 730–49. ©2018 AACR.This article is highlighted in the In This Issue feature, p. 663

Figure S1. Quantitative reverse-transcription PCR assays for let-7a, let-7b, and RNU6-2 using 10-fold dilution series from the same specimen. Figure S2. let-7a and let-7b expression in colorectal cancer. Table S1. Inter-assay coefficients of variation in quantitative reverse-transcription PCR assays for let-7a, let-7b and RNU6-2. Table S2. Clinical, pathological, and molecular characteristics according to tumor let-7b expression in 792 colorectal cancer cases. Table S3. Association of tumor let-7a and let-7b expression with disease stage. Table S4. Spearman's rank-correlation coefficients between the densities of T-cells. Table S5. Tumor let-7a expression and patient mortality in the 518 cases with CD45RO+ T-cell density data.

Expression of the immune checkpoint ligand CD274 (programmed cell death 1 ligand 1, PD-L1, from gene CD274) contributes to suppression of antitumor T cell–mediated immune response in various tumor types. However, the role of PDCD1LG2 (PD-L2, CD273, from gene PDCD1LG2) in the tumor microenvironment remains unclear. We hypothesized that tumor PDCD1LG2 expression might be inversely associated with lymphocytic reactions to colorectal cancer. We examined tumor PDCD1LG2 expression by IHC in 823 colon and rectal carcinoma cases within two U.S.-nationwide cohort studies and categorized tumors into quartiles according to the percentage of PDCD1LG2–expressing carcinoma cells. We conducted multivariable ordinal logistic regression analysis to assess the associations of tumor PDCD1LG2 expression with Crohn-like lymphoid reaction, peritumoral lymphocytic reaction, intratumoral periglandular reaction, or tumor-infiltrating lymphocytes, controlling for potential confounders, including microsatellite instability, CpG island methylator phenotype, long-interspersed nucleotide element-1 methylation, and KRAS, BRAF, and PIK3CA mutations. Tumor PDCD1LG2 expression was inversely associated with Crohn-like lymphoid reaction (Ptrend = 0.0003). For a unit increase in the three-tiered ordinal categories of Crohn-like lymphoid reaction, a multivariable OR in the highest (vs. lowest) quartile of the percentage of PDCD1LG2–expressing tumor cells was 0.38 (95% confidence interval, 0.22–0.67). Tumor PDCD1LG2 expression was not associated with peritumoral lymphocytic reaction, intratumoral periglandular reaction, tumor-infiltrating lymphocytes, or patient survival (Ptrend > 0.13). Thus, tumor PDCD1LG2 expression is inversely associated with Crohn-like lymphoid reaction to colorectal cancer, suggesting a possible role of PDCD1LG2-expressing tumor cells in inhibiting the development of tertiary lymphoid tissues during colorectal carcinogenesis. Cancer Immunol Res; 5(11); 1046–55. ©2017 AACR.

PDF FIle - 95K, Characteristics of colorectal cancer patients according to PTGS2 (COX-2) status.

Comparison of RAS or PIK3CA mutations between left- and right- colon in tumor tissue and ctDNA

Purpose: To assess prognostic roles of various KRAS oncogene mutations in colorectal cancer, BRAF mutation status must be controlled for because BRAF mutation is associated with poor prognosis, and almost all BRAF mutants are present among KRAS wild-type tumors. Taking into account experimental data supporting a greater oncogenic effect of codon 12 mutations compared with codon 13 mutations, we hypothesized that KRAS codon 12–mutated colorectal cancers might behave more aggressively than KRAS wild-type tumors and codon 13 mutants.Experimental design: Using molecular pathological epidemiology database of 1,261 rectal and colon cancers, we examined clinical outcome and tumor biomarkers of KRAS codon 12 and 13 mutations in 1,075 BRAF wild-type cancers (i.e., controlling for BRAF status). Cox proportional hazards model was used to compute mortality HR, adjusting for potential confounders, including stage, PIK3CA mutations, microsatellite instability, CpG island methylator phenotype, and LINE-1 methylation.Results: Compared with patients with KRAS wild-type/BRAF wild-type cancers (N = 635), those with KRAS codon 12 mutations (N = 332) experienced significantly higher colorectal cancer–specific mortality [log-rank P = 0.0001; multivariate HR, 1.30; 95% confidence interval (CI), 1.02–1.67; P = 0.037], whereas KRAS codon 13–mutated cases (N = 108) were not significantly associated with prognosis. Among the seven most common KRAS mutations, c.35G>T (p.G12V; N = 93) was associated with significantly higher colorectal cancer–specific mortality (log-rank P = 0.0007; multivariate HR, 2.00; 95% CI, 1.38–2.90, P = 0.0003) compared with KRAS wild-type/BRAF wild-type cases.Conclusions:KRAS codon 12 mutations (in particular, c.35G>T), but not codon 13 mutations, are associated with inferior survival in BRAF wild-type colorectal cancer. Our data highlight the importance of accurate molecular characterization in colorectal cancer. Clin Cancer Res; 18(17); 4753–63. ©2012 AACR.

PDF file - 98K

Assessment of aspirin use and statistical analysis to assess an interaction of tumor MIR21 expression level and regular aspirin use after diagnosis in survival analysis

Supplementary materials, methods and results

PDF file, 75K, Supplementary Table 1 shows characteristics of 1075 BRAF-wild-type cases. Supplementary Table 2 shows survival analysis limited to known stage I-IV cases. Supplementary Table 3 shows survival analysis including BRAF-mutated cases (total N=1255).

Background: Higher levels of physical activity are associated with lower colorectal carcinoma incidence and mortality, perhaps through influencing energy balance, cellular prosta7 systemic inflammation. Although evidence suggests interactive effects of energetics, sedentary lifestyle, and tumor CTNNB1 (β-catenin) or CDKN1B (p27) status on colon cancer prognosis, interactive effects of physical activity and tumor PTGS2 (the official symbol for COX-2) status on clinical outcome remain unknown.Methods: Using molecular pathological epidemiology database of 605 stage I–III colon and rectal cancers in two prospective cohort studies (the Nurse's Health Study and the Health Professionals Follow-up Study), we examined patient survival according to postdiagnosis physical activity and tumor PTGS2 status (with 382 PTGS2-positive and 223 PTGS2-negative tumors by immunohistochemistry). Cox proportional hazards models were used to calculate colorectal cancer-specific mortality HR, adjusting for clinical and other tumor variables including microsatellite instability status.Results: Among PTGS2-positive cases, compared with the least active first quartile, the multivariate HRs (95% confidence interval) were 0.30 (0.14–0.62) for the second, 0.38 (0.20–0.71) for the third, and 0.18 (0.08–0.41) for the fourth quartile of physical activity level (Ptrend = 0.0002). In contrast, among PTGS2-negative cases, physical activity level was not significantly associated with survival (Ptrend = 0.84; Pinteraction = 0.024, between physical activity and tumor PTGS2 status).Conclusions: Postdiagnosis physical activity is associated with better survival among patients with PTGS2-positive tumors but not among patients with PTGS2-negative tumors.Impact: Immunohistochemical PTGS2 expression in colorectal carcinoma may serve as a predictive biomarker in pathology practice, which may predict stronger benefit from exercise. Cancer Epidemiol Biomarkers Prev; 22(6); 1142–52. ©2013 AACR.

Supplementary Table 1. Number of pancreatic cancer cases by cohort with available plasma leptin levels and genotyped SNPs at LEPR Supplementary Table 2. Characteristics of controls from five prospective cohorts Supplementary Table 3. Partial Spearman correlation coefficients between plasma leptin and covariates among controls Supplementary Table 4. Odds ratios (ORs) and 95% confidence intervals (CIs) for pancreatic cancer according to quintiles of plasma leptin in sensitivity analyses excluding diabetics and short time intervals from blood collection to cancer diagnosis Supplementary Table 5. Multivariate-adjusted ORs (95% CI) for pancreatic cancer by plasma leptin, among subgroups* Supplementary Table 6. Association between SNPs at the LEPR gene and risk of pancreatic cancer among men Supplementary Table 7. Genomic and functional support for rs10493380 and correlated (r2 {greater than or equal to} 0.6) surrogate variants in HaploReg and Regulome DB Supplementary Table 8. Association between single nucleotide polymorphisms at the LEPR gene and plasma leptin among controls Supplementary Figure 1. Association results, recombination hotspots and linkage disequilibrium for single nucleotide polymorphisms at the LEPR gene region among women.

Potential effects on protein function of candidate mutations identified in cfDNA, as predicted using PolyPhen-2 and SIFT

Leptin is an adipokine involved in regulating energy balance, which has been identified as a potential biologic link in the development of obesity-associated cancers, such as pancreatic cancer. In this prospective, nested case–control study of 470 cases and 1,094 controls from five U.S. cohorts, we used conditional logistic regression to evaluate pancreatic cancer risk by prediagnostic plasma leptin, adjusting for race/ethnicity, diabetes, body mass index, physical activity, plasma C-peptide, adiponectin, and 25-hydroxyvitamin D. Because of known differences in leptin levels by gender, analyses were conducted separately for men and women. We also evaluated associations between 32 tagging SNPs in the leptin receptor (LEPR) gene and pancreatic cancer risk. Leptin levels were higher in female versus male control participants (median, 20.8 vs. 6.7 ng/mL; P < 0.0001). Among men, plasma leptin was positively associated with pancreatic cancer risk and those in the top quintile had a multivariable-adjusted OR of 3.02 [95% confidence interval (CI), 1.27–7.16; Ptrend = 0.02] compared with men in the bottom quintile. Among women, circulating leptin was not associated with pancreatic cancer risk (Ptrend = 0.21). Results were similar across cohorts (Pheterogeneity = 0.88 for two male cohorts and 0.35 for three female cohorts). In genetic analyses, rs10493380 in LEPR was associated with increased pancreatic cancer risk among women, with an OR per minor allele of 1.54 (95% CI, 1.18–2.02; multiple hypothesis-corrected P = 0.03). No SNPs were significantly associated with risk in men. In conclusion, higher prediagnostic levels of plasma leptin were associated with an elevated risk of pancreatic cancer among men, but not among women. Cancer Res; 76(24); 7160–7. ©2016 AACR.

Dynamics of mutations related to acquired resistance in validation cohort

Purpose: Mutations in KRAS are considered to be the main drivers of acquired resistance to epidermal growth factor receptor (EGFR) blockade in patients with metastatic colorectal cancer (mCRC). However, the potential role of other genes downstream of the EGFR signaling pathway in conferring acquired resistance has not been extensively investigated.Experimental Design: Using circulating tumor DNA (ctDNA) from patients with mCRC and with acquired cetuximab resistance, we developed a targeted amplicon ultra-deep sequencing method to screen for low-abundance somatic mutations in a panel of genes that encode components of the EGFR signaling pathway. Mutations with significantly increased variant frequencies upon disease progression were selected by using quartile analysis. The functional consequences of the identified mutations were validated in cultured cells.Results: We analyzed 32 patients with acquired cetuximab resistance in a development cohort. Of them, seven (22%) carried five novel PIK3CA mutations, whereas eight (25%) carried previously reported KRAS mutations. Functional studies showed that novel PIK3CA mutations (all in exon 19; p.K944N, p.F930S, p.V955G, p.V955I, and p.K966E) promote cell viability in the presence of cetuximab. Only one novel PIK3CA mutation (p.K944N) was verified in one of the 27 patients with acquired resistance in a validation cohort, simultaneous KRAS and PIK3CA hotspot mutations were detected in two patients. Among the above 59 acquired resistance patients, those with PIK3CA or RAS mutations detected in ctDNA showed a pronounced decrease in progression-free survival than patients with no mutation.Conclusions: The PIK3CA mutations may potentially contribute to acquired cetuximab resistance in patients with mCRC. Clin Cancer Res; 23(16); 4602–16. ©2017 AACR.

Supplemental Figure 1. Restricted cubic spline plot of CRC-specific and overall survival at 5 years by CTNNB1 status

Purpose: Mutations in PIK3CA [the gene encoding the p110α catalytic subunit of phosphatidylinositide-3-kinase (PI3K)] play an important role in colorectal carcinogenesis. Experimental evidence suggests that PIK3CA exon 9 and exon 20 mutations trigger different biologic effects, and that concomitant mutations in both exons 9 and 20 synergistically enhance tumorigenic effects. Thus, we hypothesized that PIK3CA exon 9 and exon 20 mutations might have differential effects on clinical outcome in colorectal cancer, and that concomitant PIK3CA exon 9 and 20 mutations might confer aggressive tumor behavior.Experimental Design: We sequenced PIK3CA by pyrosequencing in 1,170 rectal and colon cancers in two prospective cohort studies, and found 189 (16%) PIK3CA mutated tumors. Mortality HR according to PIK3CA status was computed using Cox proportional hazards model, adjusting for clinical and molecular features, including microsatellite instability, CpG island methylator phenotype, LINE-1 methylation, and BRAF and KRAS mutations.Results: Compared with PIK3CA wild-type cases, patients with concomitant PIK3CA mutations in exons 9 and 20 experienced significantly worse cancer-specific survival [log-rank P = 0.031; multivariate HR = 3.51; 95% confidence interval (CI): 1.28–9.62] and overall survival (log-rank P = 0.0008; multivariate HR = 2.68; 95% CI: 1.24–5.77). PIK3CA mutation in either exon 9 or 20 alone was not significantly associated with patient survival. No significant interaction of PIK3CA mutation with BRAF or KRAS mutation was observed in survival analysis.Conclusion: Coexistence of PIK3CA (the PI3K p110α subunit) exon 9 and 20 mutations, but not PIK3CA mutation in either exon 9 or 20 alone, is associated with poor prognosis of colorectal cancer patients. Clin Cancer Res; 18(8); 2257–68. ©2012 AACR.

Purpose: Prostaglandin-endoperoxide synthase 2 (PTGS2, cyclooxygenase-2; a target of aspirin) produces inflammatory mediator prostaglandin E2 (PGE2), and contributes to colorectal neoplasia development. PTGS2-driven inflammatory responses can induce tumor expression of microRNA MIR21 (miR-21) that can increase local PGE2 level by downregulating PGE2-metabolizing enzymes. We hypothesized that the prognostic association of tumor MIR21 expression level in colorectal carcinoma might depend on inflammatory tumor microenvironment and be stronger in tumors expressing high-level PTGS2.Experimental Design: Utilizing 765 rectal and colon cancer specimens in the Nurses' Health Study and the Health Professionals Follow-up Study, we measured MIR21 expression by quantitative reverse transcription PCR, and PTGS2 expression by immunohistochemistry. Cox proportional hazards regression model was used to assess statistical interaction between MIR21 and PTGS2 in colorectal cancer–specific survival analysis, controlling for potential confounders including microsatellite instability, CpG island methylator phenotype, LINE-1 methylation level, and KRAS, BRAF, and PIK3CA mutations.Results: Tumor MIR21 expression level was associated with higher colorectal cancer–specific mortality (Ptrend = 0.029), and there was a statistically significant interaction between MIR21 and PTGS2 (Pinteraction = 0.0004). The association between MIR21 expression and colorectal cancer–specific mortality was statistically significant in PTGS2-high cancers (multivariable hazard ratio of the highest vs. lowest quartile of MIR21, 2.28; 95% confidence interval, 1.42–3.67; Ptrend = 0.0004) but not in PTGS2-absent/low cancers (Ptrend = 0.22).Conclusions:MIR21 expression level in colorectal carcinoma is associated with worse clinical outcome, and this association is stronger in carcinomas expressing high-level PTGS2, suggesting complex roles of immunity and inflammation in tumor progression. Clin Cancer Res; 22(15); 3841–8. ©2016 AACR.

Regular aspirin use after diagnosis and colorectal cancer mortality according to tumor MIR21 expression level among 579 patients with stage I to III colorectal cancer

Dynamics of mutations related to acquired resistance in plasma samples of development cohort

Clinical information of validation cohort

Transcriptional addiction plays a pivotal role in maintaining the hallmarks of cancer cells. Thus, targeting super-enhancers (SEs), which modulate the transcriptional activity of oncogenes, has become an attractive strategy for cancer therapy. As yet, however, the molecular mechanisms of this process in bladder cancer (BC) remain to be elucidated. Here, we aimed to provide detailed information regarding the SE landscape in BC and to investigate new potential pharmaceutical targets for BC therapy. We employed THZ1 as a potent and specific CDK7 inhibitor. In vitro and in vivo studies were carried out to investigate the anticancer and apoptosis-inducing effects of THZ1 on BC cells. Whole-transcriptome sequencing (RNA-seq) and chromatin immunoprecipitation sequencing (ChIP-seq) were performed to investigate the mechanism and function of SE-linked oncogenic transcription in BC cells. We found that THZ1 serves as an effective and potent inhibitor with suppressive activity against BC cells. An integrative analysis of THZ1-sensitive and SE-associated oncogenes yielded potential new pharmaceutical targets, including DDIT4, B4GALT5, PSRC1 and MED22. Combination treatment with THZ1 and the DDIT4 inhibitor rapamycin effectively suppressed BC cell growth. In addition, we found that THZ1 and rapamycin sensitized BC cells to conventional chemotherapy. Our data indicate that exploring BC gene regulatory mechanisms associated with SEs through integrating RNA-seq and ChIP-seq data improves our understanding of BC biology and provides a basis for innovative therapies.

Background The high expression across multiple tumor types and restricted expression in normal tissues make B7-H3 an attractive target for immunotherapy. So far, little is known about the clinical significance of B7-H3 expression in meningiomas. We conducted this study to address this issue in a cohort of 242 patients from a single institution. Methods Expression profiles of immune checkpoint proteins (programmed death-ligand 1, B7-H3, lymphocyte activation gene-3, programmed death 1, and V-domain Ig suppressor of T cell activation) were explored by immunohistochemistry in a meningioma test cohort (n = 8). Role of B7-H3 expression was further assessed in an expanded patient cohort (n = 234) using immunohistochemical tissue microarray analysis. Results B7-H3 expression was significantly greater than all immune checkpoint proteins studied in the tested cohort. B7-H3 was detected with different degrees in all meningioma specimens, predominantly on tumor cell membranes and in cytoplasm. Tumors were classified as B7-H3 high or low group depending on immunohistochemistry histoscore (median histoscore 111.06; range, 7.313–212.008). B7-H3 expression was statistically correlated with patient sex (P = 0.0297), tumor histopathologic subtypes (P = 0.0262), and radiotherapy after surgery (P = 0.0028). However, no significant differences were observed in patient age, tumor location, and grade and extent of resection between groups. Similarly, there was no significant difference in progression-free survival and overall survival between B7-H3 high and low group. Conclusions Our study indicates variable expression and clinical role of B7-H3 in meningiomas, suggesting its potential as an immunotherapeutic target in the future.

No specific effective therapeutic drugs have been identified for COVID-19. Critically ill COVID-19 36 patients in the ICU experience high mortality. This project aims to study the effects of traditional 37 Chinese medicine (TCM) treatment on deadly outcomes caused by COVID-19. A total of 123 critically 38 ill COVID-19 patients who received close monitoring at the ICU of Vulcan Hill Hospital between 39 ICU patients received supportive management. Eighty-one patients were given additional TCM 41 treatment. Clinical characteristics during the treatment period (up to 39 days) and the clinical outcome 42 of each patient were closely monitored and analysed. We observed that patients treated with TCM had 43 lower mortality than the non-TCM treatment group (16 of 81 vs. 31 of 42; 0.3 person/month vs. 2.9 44 person/month). In the adjusted Cox proportional hazards models, TCM treatment was associated with 45 improved survival [multivariate HR, 0.13; 95% confidence interval (CI), 0.06–0.24; P < 0.001]. 46 Furthermore, we found that TCM treatment could partially improve the inflammation status by 47 reducing the levels of proinflammatory cytokines and recovering multiple organic functions. TCM 48 treatment may decrease inflammation status by reducing the level of proinflammatory cytokines and 49 allowing the recovery of multiple organic functions, which could improve the survival rate of critically 50 ill COVID-19 patients.