CD44 is an adhesion molecule expressed in cancer stem-like cells. Here, we show that a CD44 variant (CD44v) interacts with xCT, a glutamate-cystine transporter, and controls the intracellular level of reduced glutathione (GSH). Human gastrointestinal cancer cells with a high level of CD44 expression showed an enhanced capacity for GSH synthesis and defense against reactive oxygen species (ROS). Ablation of CD44 induced loss of xCT from the cell surface and suppressed tumor growth in a transgenic mouse model of gastric cancer. It also induced activation of p38(MAPK), a downstream target of ROS, and expression of the gene for the cell cycle inhibitor p21(CIP1/WAF1). These findings establish a function for CD44v in regulation of ROS defense and tumor growth.

BACKGROUND Regular use of aspirin after a diagnosis of colon cancer has been associated with a superior clinical outcome. Experimental evidence suggests that inhibition of prostaglandin-endoperoxide synthase 2 (PTGS2) (also known as cyclooxygenase-2) by aspirin down-regulates phosphatidylinositol 3-kinase (PI3K) signaling activity. We hypothesized that the effect of aspirin on survival and prognosis in patients with cancers characterized by mutated PIK3CA (the phosphatidylinositol-4,5-bisphosphonate 3-kinase, catalytic subunit alpha polypeptide gene) might differ from the effect among those with wild-type PIK3CA cancers. METHODS We obtained data on 964 patients with rectal or colon cancer from the Nurses’ Health Study and the Health Professionals Follow-up Study, including data on aspirin use after diagnosis and the presence or absence of PIK3CA mutation. We used a Cox proportional-hazards model to compute the multivariate hazard ratio for death. We examined tumor markers, including PTGS2, phosphorylated AKT, KRAS, BRAF, microsatellite instability, CpG island methylator phenotype, and methylation of long interspersed nucleotide element 1. RESULTS Among patients with mutated-PIK3CA colorectal cancers, regular use of aspirin after diagnosis was associated with superior colorectal cancer–specific survival (multivariate hazard ratio for cancer-related death, 0.18; 95% confidence interval [CI], 0.06 to 0.61; P<0.001 by the log-rank test) and overall survival (multivariate hazard ratio for death from any cause, 0.54; 95% CI, 0.31 to 0.94; P = 0.01 by the log-rank test). In contrast, among patients with wild-type PIK3CA, regular use of aspirin after diagnosis was not associated with colorectal cancer–specific survival (multivariate hazard ratio, 0.96; 95% CI, 0.69 to 1.32; P = 0.76 by the log-rank test; P = 0.009 for interaction between aspirin and PIK3CA variables) or overall survival (multivariate hazard ratio, 0.94; 95% CI, 0.75 to 1.17; P = 0.96 by the log-rank test; P = 0.07 for interaction). CONCLUSIONS Regular use of aspirin after diagnosis was associated with longer survival among patients with mutated-PIK3CA colorectal cancer, but not among patients with wild-type PIK3CA cancer. The findings from this molecular pathological epidemiology study suggest that the PIK3CA mutation in colorectal cancer may serve as a predictive molecular biomarker for adjuvant aspirin therapy. (Funded by The National Institutes of Health and others.)

The abundance of tumour-infiltrating T-cells has been associated with microsatellite instability (MSI) and a favorable prognosis in colorectal cancer. Because molecular alterations in colon cancer including MSI, the CpG island methylator phenotype (CIMP), BRAF mutation and global DNA hypomethylation have been associated with clinical outcome, potential confounding by these molecular features needs to be controlled when assessing the prognostic significance of tumour-infiltrating T-cells. We utilized a database of clinically and molecularly-annotated colon and rectal carcinoma cases (N=768; stage I-IV) in two prospective cohort studies (the Nurses' Health Study and the Health Professionals Follow-up Study). Using tissue microarray and automated Ariol image analysis system, we quantified densities of CD3 + , CD8 + , CD45RO + (PTPRC) and FOXP3 + -cells within neoplastic epithelial areas. We used Cox proportional hazard models to compute mortality hazard ratio, adjusting for clinical and molecular features including KRAS, BRAF, and PIK3CA mutations, MSI, CIMP and LINE-1 hypomethylation. The densities of CD8 + , CD45RO + and FOXP3 + -cells were significantly associated with patient survival in univariate analyses (P trend <0.007). In the multivariate model, tumour-infiltrating CD45RO + -cell density, but not that of CD3 + , CD8 + or FOXP3 + -cell, was significantly associated with survival (p=0.0032). In multivariate linear regression analysis, MSI-high (p<0.0001) and high-level tumour LINE-1 methylation (p=0.0013) were independently associated with higher CD45RO + -cell NIH-PA Author ManuscriptNIH-PA Author Manuscript NIH-PA Author Manuscript density. Nonetheless, the survival benefit associated with CD45RO + -cells was independent of MSI and LINE-1 status. In conclusion, tumour-infiltrating CD45RO + -cell density is a prognostic biomarker associated with longer survival of colorectal cancer patients, independent of clinical, pathological and molecular features. In addition, MSI-high and tumour LINE-1 methylation level are independent predictors of CD45RO + -cell density. Our data offer a possible mechanism by which MSI confers an improved clinical outcome, and support efforts to augment host immune response in the tumour microenvironment as a strategy of targeted immunotherapy.

Purpose: Host immune response to tumor may be an important prognostic factor for colon cancer patients. However, little is known on prognostic significance of histopathologic lymphoid reaction to tumor, independent of the number of lymph nodes examined and tumoral molecular alterations, including microsatellite instability (MSI) and the CpG island methylator phenotype (CIMP), both of which are associated with lymphocytic reaction and clinical outcome. Experimental Design: Using 843 colorectal cancer patients in two independent prospective cohorts, we examined patient prognosis in relation to four components of lymphocytic reaction (i.e., Crohn's-like reaction, peritumoral reaction, intratumoral periglandular reaction, and tumor-infiltrating lymphocytes) and overall lymphocytic score (0-12). CIMP was determined using eight markers including CACNA1G, CDKN2A (p16), CRABP1, IGF2, MLH1, NEUROG1, RUNX3, and SOCS1. Cox proportional hazard models computed hazard ratio for mortality, adjusted for covariates including tumor stage, body mass index, lymph node count, KRAS, BRAF, p53, cyclooxygenase-2 (PTGS2), MSI, CIMP, and LINE-1 methylation. Results: Increasing overall lymphocytic reaction score including tumor-infiltrating lymphocytes was associated with a significant improvement in colorectal cancerspecific and overall survival (log-rank P < 0.003). These findings remained significant (adjusted hazard ratio estimates, 0.49-0.71; P trend < 0.009) in multivariate models that adjusted for covariates, including body mass index, MSI, CIMP, LINE-1 hypomethylation, and cyclooxygenase-2. The beneficial effect of tumoral lymphocytic reaction was consistent across strata of clinical, pathologic, and molecular characteristics. Conclusions: Lymphocytic reactions to tumor were associated with improved prognosis among colorectal cancer patients, independent of lymph node count and other clinical, pathologic, and molecular characteristics. (Clin Cancer Res 2009;15(20): 6412-20)