Differential association of STK11 and TP53 with KRAS mutation-associated gene expression, proliferation and immune surveillance in lung adenocarcinoma.

While mutations in the KRAS oncogene are among the most prevalent in human cancer, there are few successful treatments to target these tumors. It is also likely that heterogeneity in KRAS-mutant tumor biology significantly contributes to the response to therapy. We hypothesized that the presence of commonly co-occurring mutations in STK11 and TP53 tumor suppressors may represent a significant source of heterogeneity in KRAS-mutant tumors. To address this, we utilized a large cohort of resected tumors from 442 lung adenocarcinoma patients with data including annotation of prevalent driver mutations (KRAS and EGFR) and tumor suppressor mutations (STK11 and TP53), microarray-based gene expression and clinical covariates, including overall survival (OS). Specifically, we determined impact of STK11 and TP53 mutations on a new KRAS mutation-associated gene expression signature as well as previously defined signatures of tumor cell proliferation and immune surveillance responses. Interestingly, STK11, but not TP53 mutations, were associated with highly elevated expression of KRAS mutation-associated genes. Mutations in TP53 and STK11 also impacted tumor biology regardless of KRAS status, with TP53 strongly associated with enhanced proliferation and STK11 with suppression of immune surveillance. These findings illustrate the remarkably distinct ways through which tumor suppressor mutations may contribute to heterogeneity in KRAS-mutant tumor biology. In addition, these studies point to novel associations between gene mutations and immune surveillance that could impact the response to immunotherapy.

Claudin-2 expression increases tumorigenicity of colon cancer cells: role of epidermal growth factor receptor activation.

Claudin-2 is a unique member of the claudin family of transmembrane proteins, as its expression is restricted to the leaky epithelium in vivo and correlates with epithelial leakiness in vitro. However, recent evidence suggests potential functions of claudin-2 that are relevant to neoplastic transformation and growth. In accordance, here we report, on the basis of analysis of mRNA and protein expression using a total of 309 patient samples that claudin-2 expression is significantly increased in colorectal cancer and correlates with cancer progression. We also report similar increases in claudin-2 expression in inflammatory bowel disease-associated colorectal cancer. Most importantly, we demonstrate that the increased claudin-2 expression in colorectal cancer is causally associated with tumor growth as forced claudin-2 expression in colon cancer cells that do not express claudin-2 resulted in significant increases in cell proliferation, anchorage-independent growth and tumor growth in vivo. We further show that the colonic microenvironment regulates claudin-2 expression in a manner dependent on signaling through the EGF receptor (EGFR), a key regulator of colon tumorigenesis. In addition, claudin-2 expression is specifically decreased in the colon of waved-2 mice, naturally deficient in EGFR activation. Furthermore, genetic silencing of claudin-2 expression in Caco-2, a colon cancer cell line, prevents the EGF-induced increase in cell proliferation. Taken together, these results uncover a novel role for claudin-2 in promoting colon cancer, potentially via EGFR transactivation.

HDAC inhibitors regulate claudin-1 expression in colon cancer cells through modulation of mRNA stability.

Expression and cellular distribution of claudin-1, a tight junction protein, is dysregulated in colon cancer and its overexpression in colon cancer cells induced dedifferentiation and increased invasion. However, the molecular mechanism(s) underlying dysregulated claudin-1 expression in colon cancer remains poorly understood. Histone deacetylase (HDAC)-dependent histone acetylation is an important mechanism of the regulation of cancer-related genes and inhibition of HDACs induces epithelial differentiation and decreased invasion. Therefore, in this study, we examined the role of HDAC-dependent epigenetic regulation of claudin-1 in colon cancer. In this study, we show that sodium butyrate and Trichostatin A (TSA), two structurally different and widely used HDAC inhibitors, inhibited claudin-1 expression in multiple colon cancer cell lines. Further studies revealed modulation of claudin-1 mRNA stability by its 3'-UTR as the major mechanism underlying HDAC-dependent claudin-1 expression. In addition, overexpression of claudin-1 abrogated the TSA-induced inhibition of invasion in colon cancer cells suggesting functional crosstalk. Analysis of mRNA expression in colon cancer patients, showed a similar pattern of increase in claudin-1 and HDAC-2 mRNA expression throughout all stages of colon cancer. Inhibition of claudin-1 expression by HDAC-2-specific small interfering RNA further supported the role of HDAC-2 in this regulation. Taken together, we report a novel post-transcriptional regulation of claudin-1 expression in colon cancer cells and further show a functional correlation between claudin-1 expression and TSA-mediated regulation of invasion. As HDAC inhibitors are considered to be promising anticancer drugs, these new findings will have implications in both laboratory and clinical settings.

IgG auto- and polyreactivities of normal human sera.

Using a panel of self antigens, IgM autoreactivities were clearly and constantly detected by enzyme immunoassay (EIA) in the sera of 29 normal human individuals. Similarly, IgM autoreactivities in sera were reproducibly detected by immunoblotting, using human organ extracts as the antigen sources. In contrast, IgG reactivities were low in whole sera but were considerably increased after affinity-chromatography purification on protein G-Sepharose. These increases differed from one individual IgG preparation to another and from one antigen to another (from 1-94 times) resulting in a unique IgG autoreactivity pattern for each subject. IgG reactivities diminished markedly when the IgG-depleted serum was added to the isolated autologous IgG. IgM antibodies isolated from sera on F(ab')2 IgG immunoadsorbent partially inhibited the binding of IgG to tubulin and myosin but not to actin. The individual IgG preparations examined separately exhibited, with all the autoantigens of the panel, higher autoreactivities than those of the same-but-pooled IgGs, which in turn were higher than those of a commercially available human IgG preparation obtained from approximately 8,000 healthy donors and used for intravenous injection. Depending upon the individual IgG sample, 31-65% of the IgG were bound to a DNP-Sepharose column and were eluted with DNP-glycine. The isolated anti-DNP antibodies were found to be polyreactive and possess higher autoreactivities than the original IgG preparation for all the antigens of the panel. Similarly, IgG antibodies analysed using an antibody exchange procedure were found to be essentially polyreactive but some apparently monospecific antibodies were also noted. These results suggest that the great majority of IgG present in normal humans are composed of polyreactive autoantibodies. IgG autoreactivities are only marginally expressed in these whole sera because of IgM-IgG, IgG-IgG and other, still unidentified, interactions.