Role of serine 365 in BRAF V600E sensitivity to RAF inhibition.

The serine-threonine kinase, BRAF, is an upstream regulator of the MEK-ERK1/2 pathway and is commonly mutated in cancer. 14-3-3 proteins bind to two sites in BRAF, N-terminal S365, and C-terminal S729. 14-3-3 binding modulates the activity and dimerization of both wild-type and non-V600 mutant forms of BRAF. In BRAF V600E mutants, the C-terminal S729 site affects dimerization of truncated splice variants. The N-terminal, S365, is removed in BRAF V600E splice variants but its importance in full-length BRAF V600 mutants remains uncertain. We tested the role of S365 in dimerization and RAF inhibitor resistance in full-length BRAF V600E. Mutating BRAF S365 site to an alanine (S365A) reduced 14-3-3 association and increased BRAF V600E homodimerization. BRAF V600E S365A displayed reduced sensitivity to RAF inhibitor at the level of MEK-ERK1/2 signaling, cell growth, and cell viability. These data suggest that alteration or removal of the S365 14-3-3 binding site may contribute to RAF inhibitor resistance.

Are All Periprosthetic Joint Infections the Same? Evaluating Major vs Minor Criteria.

BACKGROUND: The diagnosis of periprosthetic joint infection (PJI) can be made when 1 major criterion or 3 of 5 minor criteria are present. However, the outcomes of patients with a major vs minor criteria for diagnosis have not been studied. The objective of this study was to evaluate if a difference in outcome of surgical intervention existed between patients with PJI who were diagnosed with a major criterion or a combination of minor criteria. METHODS: A retrospective chart review identified 277 primary total hip or knee arthroplasty patients who had developed PJI based on the International Consensus Meeting definition. Patients were further stratified into "major" vs "minor" groups. Patient demographics, PJI workup, surgical treatment, microbiological growth, and clinical outcomes were recorded. Treatment success was defined by using the Delphi criteria. Standard statistical analysis was performed. RESULTS: Overall, 34 patients met minor-only criteria (12.2%), whereas 243 met major criteria. Of the minor-only patients, 16 (47%) were culture negative. When controlling for confounding variables, there was no statistically significant difference with regard to treatment success (minor 94.1% vs major 82.3%, P = .085) between groups at final follow-up (mean 110 months, range 2.3-567 months). Only higher Charlson comorbidity index (P = .001) and an initial 2-stage surgical procedure (P = .003) were associated with decreased treatment success. CONCLUSION: PJI patients were similar between both criteria groups, and there was no difference in treatment success as defined by the Delphi criteria between minor-only PJI and major criteria PJI patients.

Positive Blood Cultures in Periprosthetic Joint Infection Decrease Rate of Treatment Success.

BACKGROUND: Blood cultures are often obtained at the time of periprosthetic joint infection (PJI) diagnosis yet they are not considered part of the diagnostic criteria and the effects of a positive result on surgical outcome are unknown. The purposes of this study are to characterize the use of blood cultures when diagnosing PJI and to determine the association of positive blood cultures with PJI treatment success. METHODS: A retrospective chart review on 320 patients surgically treated for primary hip and knee PJIs was performed from 2006-2013 at 2 academic medical centers with minimum 12-month follow-up. Treatment success was defined by the Delphi criteria. Multiple logistic regression analysis was performed to identify variables associated with treatment success. RESULTS: Blood cultures were obtained from 53.1% of PJI patients (170/320) at the time of diagnosis. The same organism was identified 86.0% of the time in blood culture and operative culture. Patients with positive blood cultures at the time of PJI diagnosis had elevated synovial white blood cell count (98,979, P = .012), elevated serum C-reactive protein (24.2 mg/L, P < .001), and decreased treatment success (65.1%) compared with those with a negative blood culture (85.0%) and those without a blood culture (82.7%, P = .013). A positive blood culture remained associated with decreased PJI treatment success using multiple logistic regression analysis. CONCLUSION: The presence of positive blood cultures at the time of PJI diagnosis decreased PJI treatment success. Further prospective studies are needed to help identify the role of blood cultures in the work up of PJI and treatment optimization in these patients.

Obesity-Induced Colorectal Cancer Is Driven by Caloric Silencing of the Guanylin-GUCY2C Paracrine Signaling Axis.

Obesity is a well-known risk factor for colorectal cancer but precisely how it influences risks of malignancy remains unclear. During colon cancer development in humans or animals, attenuation of the colonic cell surface receptor guanylyl cyclase C (GUCY2C) that occurs due to loss of its paracrine hormone ligand guanylin contributes universally to malignant progression. In this study, we explored a link between obesity and GUCY2C silencing in colorectal cancer. Using genetically engineered mice on different diets, we found that diet-induced obesity caused a loss of guanylin expression in the colon with subsequent GUCY2C silencing, epithelial dysfunction, and tumorigenesis. Mechanistic investigations revealed that obesity reversibly silenced guanylin expression through calorie-dependent induction of endoplasmic reticulum stress and the unfolded protein response in intestinal epithelial cells. In transgenic mice, enforcing specific expression of guanylin in intestinal epithelial cells restored GUCY2C signaling, eliminating intestinal tumors associated with a high calorie diet. Our findings show how caloric suppression of the guanylin-GUCY2C signaling axis links obesity to negation of a universal tumor suppressor pathway in colorectal cancer, suggesting an opportunity to prevent colorectal cancer in obese patients through hormone replacement with the FDA-approved oral GUCY2C ligand linaclotide.

Translating colorectal cancer prevention through the guanylyl cyclase C signaling axis.

Colorectal cancer (CRC) is a major public health concern, ranking among the leading causes of cancer death in both men and women. Because of this continued burden there is a clear need for improved treatment, and more importantly prevention of this disease. In recent years there is significant evidence to support the hypothesis that guanylyl cyclase C (GCY2C) is a tumor suppressor in the intestine, and that the loss of hormone ligands for this receptor is an important step in the disease process. Thus, ligand replacement therapy has been proposed as a strategy to prevent CRC. Until recently this strategy was not clinically plausible; however, the recent regulatory approval of linaclotide (LINZESS™, Forest Laboratories and Ironwood Pharmaceuticals, Inc.), an oral GUCY2C ligand, has raised the possibility of utilizing this strategy clinically to prevent CRC.