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Relationships among KRAS mutation status, expression of RAS pathway signaling molecules, and clinicopathological features and prognosis of patients with colorectal cancer

Squamous cell differentiation (SCD) may occur in papillary thyroid carcinoma (PTC) only at metastatic sites. We have studied cytokeratin CK5/6 and P63 along with TTF1 (thyroid transcription factor 1) and B-Raf (V-Raf murine sarcoma viral oncogene homolog B1) immunohistochemical expression in neck lymph node metastases of thyroid PTC showing SCD. The patient (21-years) presented with a neck mass. The check-up revealed bilateral thyroid nodules. Total thyroidectomy and neck lymph node dissection were performed. The diagnosis was that of bilateral PTC with lymph node metastases (pT-1N1Mx). The metastases were peculiar by the presence of cystic change and of SCD. The thyroid PTC expressed P63 focally and, TTF1 and B-Raf diffusely. Cytokeratin 5/6 was expressed only in the lymph node metastases, in the metastatic cyst lining and in the SCD foci. The P63+ cells outnumbered those CK5/6+. TTF1 expression was faint in SCD. Metastatic, both classical PTCand SCD-epithelia expressed B-Raf. The expression patterns of CK5/6, P63, TTF1 suggest a luminal/central-to-abluminal/peripheral direction for SCD development from PTC-epithelia in lymph node metastases. Whether this metaplasia type may reflect a regression to a less aggressive morphotype or a progression-switch to squamous cell carcinoma-type differentiation in a composite tumor remains matter of debate.

Study Design: Ameloblastoma is a benign odontogenic tumor of epithelial origin, with locally aggressive behavior; and due to its invasiveness has increased recurrence rates. It is more frequent in the mandible than the maxilla. Surgical treatment is performed with approaches ranging from conservative to radical surgery, either followed by microvascular reconstruction, or not. Adjuvant treatment has shown better local control in refractory cases. Multiple relapses are associated with BRAF gene mutation at codon 600. Objective: This case series aimed to describe the clinical, imaging, histopathological, therapeutic and mutational characterization of 3 patients with refractory ameloblastoma in the maxilla. Methods: Data of 3 patients were collected, and descriptions were provided of procedures such as clinical, imaging, surgical technique, histopathological subtype and molecular analysis for detection of BRAFV600E mutation. Results It was obtained a locoregional control after RT of two cases described. After BRAF mutation molecular analysis, no patient presented it. Conclusion: Surgery remains the gold standard for the treatment of ameloblastomas, even in refractory cases; however, an expanded approach to obtain free surgical margins, and reconstruction of the maxilla itself may be challenging. Adjuvant radiotherapy is still a controversial topic, but could favor reduction of the local recurrence rate in cases where the surgical margins are compromised after surgical resection. Further studies will be necessaries for analysis of the BRAFV600E mutation, for therapeutic purposes.

Hintergrund Die Weltgesundheitsorganisation (WHO) plant eine Aktualisierung der WHO-Klassifikation maligner Lymphome.

BACKGROUND The RAS/RAF/MEK/ERK and PI3K/AKT/mTOR signaling pathways all belong to mitogen-activated protein kinase (MAPK) signaling pathways, Mutations in any one of the upstream genes (such as the RAS gene or the BRAF gene) may be transmitted to the protein through transcription or translation, resulting in abnormal activation of the signaling pathway. This study investigated the relationship between the KRAS gene mutation and the clinicopathological features and prognosis of colorectal cancer (CRC), and the effect of KRAS mutations on its associated proteins in CRC, with an aim to clarify the cause of tumor progression and drug resistance caused by mutation of the KRAS gene. AIM To investigate the KRAS gene and RAS pathway signaling molecules in CRC and to analyze their relationship with clinicopathological features and prognosis METHODS Colorectal cancer tissue specimens from 196 patients were analyzed for KRAS mutations using quantitative polymerase chain reaction and for KRAS, BRAF, MEK, and ERK protein expression levels using immunohistochemistry of tumor microarrays. To analyze differences of RAS pathway signaling molecule expression levels in different KRAS gene status, the relationships between these parameters and clinicopathological features, 4-year progression-free survival, and overall survival were analyzed by independent sample t test, Kaplan-Meier plots, and the log-rank test. Predictors of overall and disease-free survival were assessed using a Cox proportional hazards model. RESULTS Of the 196 patients, 62 (32%) carried mutations in codon 12 (53/62) or codon 13 (9/62) in exon 2 of the KRAS gene. KRAS, BRAF, ERK, and MEK protein expression was detected in 71.4%, 78.8%, 64.3%, and 50.8% of CRC tissues, respectively. There were no significant differences between KRAS mutation status and KRAS, BRAF, MEK, or ERK protein levels. Positive expression of KRAS and ERK was associated with poor tumor differentiation, and KRAS expression was also associated with age < 56 years. MEK expression was significantly associated with distant metastasis (P < 0.05). The 4-year progression-free survival rate, but not overall survival rate, was significantly higher in patients with KRAS-negative tumors than in those with KRAS-positive tumors (P < 0.05), whereas BRAF, MEK, and ERK expression was unrelated to survival. Multivariate analysis showed that only the expression of KRAS protein was a risk factor for tumor recurrence (P < 0.05). No other clinicopathological factors correlated with KRAS, BRAF, MEK, or ERK expression. CONCLUSION KRAS gene mutations do not affect downstream protein expression in CRC. KRAS protein is associated with poor tumor differentiation, older age, and a risk of tumor recurrence.

Mutations in the BRAF gene are well known for their oncogenic effects. Point mutations in V599 are particularly oncogenic and are considered important for therapeutic purposes. Along with wild type, other V599 mutated BRAF variants viz. V599E, V599D and V599R are reported and crystals of the former two with inhibitor (BAY43-9006) are further detailed. Both wild-type and mutated BRAF forms show similar interaction patterns with BAY43-9006, but the 599th residue did not show any involvement in the interactions. Upon BAY43-9006 binding, kinase domains of both forms were found adopting essentially identical conformations. However, BAY43-9006 shows a varied activity profile in the case of the wild and V599E variant of the BRAF protein. Furthermore, MMGBSA binding energy results for all four BRAF variants, further revealed the importance of the 599th residue. In-depth analysis viz. molecular dynamics, residue correlation studies and residue interaction network (RIN) analyses were conducted, providing a deep insight into the 599th residue and its impact on the overall dynamics of BRAF protein. Our findings reveal that the mutated residue at the 599th position not only changed the BAY43-9006–BRAF binding behaviour but also produced a massive impact on the overall dynamic behaviour of the protein. The insights obtained herein could be of great relevance for designing new BRAF inhibitors aimed at getting ideal activity against all BRAF forms.

Studies have shown that atypical Spitz tumors (ASTs) with homozygous deletions in 9p21 have worse prognosis than those without this finding. Conversely, numerous studies have shown that a range of other copy number aberrations including isolated 6q23 or 3p21 loss may be seen in ASTs without conferring higher risk for aggressive behavior. We studied 31 cases of ASTs with heterozygous 9p21 loss and hypothesize that heterozygous 9p21 loss in ASTs does not confer an increased risk for aggressive behavior. We compared clinical, histopathologic, and immunohistochemical features of 31 ASTs with heterozygous 9p21 deletions with 30 ASTs with homozygous 9p21 deletions. No ASTs with heterozygous 9p21 deletions resulted in distant metastasis. Severe cytologic atypia, a predominance of epithelioid cytomorphology and increased dermal mitotic activity were more frequent in ASTs with homozygous deletions versus ASTs with heterozygous deletions (P=0.0003, 0.0004, and 0.042, respectively). Expression of p16 and mutated BRAF proteins was also evaluated in 17 conventional (nonspitzoid) melanomas with homozygous 9p21 loss and the 2 groups of ASTs. Expression of p16 was retained in 67% of ASTs with heterozygous loss, whereas among ASTs with homozygous loss, 100% of cases had areas with complete loss of staining. Mutated BRAF protein expression was detected in 53% of conventional melanomas, in none of the ASTs with heterozygous loss, and in 1 AST with homozygous loss (P=0.0007 between homozygous ASTs and the conventional melanomas). Coexisting BRAF mutation and 9p21 deletion was more common in conventional melanomas than in ASTs with heterozygous or homozygous 9p21 deletion. BRAF mutation was highly uncommon among the ASTs.

Over the last few years, BRAF has emerged as a validated target in melanoma. This review summarises recent advances in the development of BRAF inhibitors, focussing on agents that have been assessed clinically.

The article summarizes the expert discussion and recommendations on the use of molecular markers and of biological targeted therapies in metastatic colorectal cancer (mCRC), as well as a proposed treatment decision strategy for mCRC treatment. The meeting was conducted during the 11th ESMO/World Gastrointestinal Cancer Congress (WGICC) in Barcelona in June 2009. The manuscript describes the outcome of an expert discussion leading to an expert recommendation. The increasing knowledge on clinical and molecular markers and the availability of biological targeted therapies have major implications in the optimal management in mCRC. 21 Suppl 6 vi1 10

Purpose This open-label, multicenter study was designed to allow access to vemurafenib for patients with metastatic melanoma, bridging the time between end of enrollment in the phase III registration trial (December 2010) and commercial availability following US Food and Drug Administration approval of vemurafenib for the treatment of unresectable or metastatic BRAF-mutated melanoma (August 2011). Patients and methods Eligible patients had metastatic melanoma with a BRAF mutation (detected by the cobas 4800 BRAF V600 Mutation Test). Unlike previous vemurafenib trials, patients with poor performance status (PS) and treated brain metastases were permitted. Enrolled patients received oral vemurafenib 960 mg twice daily. Results Of 374 patients enrolled at 29 US sites (December 2010 to October 2011), 371 patients received vemurafenib and were followed up for a median of 2.8 months (the study had a prespecified end upon vemurafenib approval and commercial availability). At baseline, most patients (75%) had stage M1c disease, and 19% had an Eastern Cooperative Oncology Group PS of 2 or 3; 72% of patients had received prior systemic therapy for metastatic melanoma, 27% received prior ipilimumab, and 29% radiotherapy for prior brain metastases. Because reassessment data to confirm response were not available for most patients, point estimates of objective response rate (ORR) are reported. Among 241 efficacy-evaluable patients, the ORR was 54% (median time to response, 1.9 months). The ORR in non-central nervous system sites in patients with previously treated brain metastases (n = 68) was 53%. The ORR in prior ipilimumab-treated patients (n = 68) was 52%. For patients with PS of 0 or 1 (n = 210) and 2 or 3 (n = 31), the ORRs were 55%, and 42%, respectively. The safety profile observed was consistent with that reported in previous studies. The number of patients with grade 3 or 4 treatment-related adverse events was higher in patients with PS 2 or 3 than in those with PS 0 or 1 (10% vs. 5%, respectively). Adverse events requiring a dose reduction (at least 1 level) occurred in 11% of patients, and 9 patients (2%) experienced events leading to vemurafenib withdrawal, including 2 with repeated QT interval prolongation. Discussion This study confirmed the established rapid and high tumor response rate achievable with vemurafenib in BRAF mutation-positive metastatic melanoma. Several groups not included in previous studies, including patients with previously treated brain metastases, Eastern Cooperative Oncology Group PS 2 to 3, or previous ipilimumab treatment had benefitted from vemurafenib similar to the overall population. No new safety signals were detected.

Personalized melanoma medicine has progressed from histopathologic features to serum markers to molecular profiles. Since the identification of activating BRAF mutations and subsequent development of drugs targeting the mutant BRAF protein, oncologists now need to incorporate prognostic and predictive biomarkers into treatment decisions for their melanoma patients. Examples include subgrouping patients by genotype profiles for targeted therapy and the development of serologic, immunohistochemical, and genotype profiles for the selection of patients for immunotherapies. In this chapter, we provide an overview of the current status of BRAF mutation testing, as well as promising serologic and molecular profiles that will impact patient care. As further research helps clarify the roles of these factors, the clinical outcomes of melanoma patients promise to be greatly improved.

Dozens of human enzymes incorporate or utilize copper, taking advantage of the metal's readiness to donate or accept electrons to catalyze key bio chemical reactions. Tumors, however, may be especially dependent on the metal. Copper, for example, promotes angiogenesis, the growth of blood vessels that can feed an expanding tumor, and depleting it may keep cancer in check. Copper also binds an enzyme that enables tumor cells to metastasize and is required for signaling by the mutant BRAF protein, which drives half of melanomas and many other cancers. "Maybe what we'll find is that particular cancer types are more susceptible to particular copper-dependent processes," says pharmacologist Donita Brady of Duke University in Durham, North Carolina.

e20120 Background: Inhibitors of the V600 mutant form of the MAPK BRAF protein induce rapid and deep responses. Unfortunately, the benefit seen with targeted therapy may be finite, with mean progre...

Objectives: Treatment decision-making in colorectal cancer is often guided by tumour tissue molecular analysis. The aim of this study was the development and validation of a high-resolution melting (HRM) method for the detection of KRAS, NRAS and BRAF mutations in Greek and Romanian patients with colorectal cancer and determination of the frequency of these mutations in the respective populations. Setting: Diagnostic molecular laboratory located in Athens, Greece. Participants: 2425 patients with colorectal cancer participated in the study. Primary and secondary outcome measures: 2071 patients with colorectal cancer (1699 of Greek and 372 of Romanian origin) were analysed for KRAS exon 2 mutations. In addition, 354 tumours from consecutive patients (196 Greek and 161 Romanian) were subjected to full KRAS (exons 2, 3 and 4), NRAS (exons 2, 3 and 4) and BRAF (exon 15) analysis. KRAS, NRAS and BRAF mutation detection was performed by a newly designed HRM analysis protocol, followed by Sanger sequencing. Results: KRAS exon 2 mutations (codons 12/13) were detected in 702 of the 1699 Greek patients with colorectal carcinoma analysed (41.3%) and in 39.2% (146/372) of the Romanian patients. Among the 354 patients who were subjected to full KRAS, NRAS and BRAF analysis, 40.96% had KRAS exon 2 mutations (codons 12/13). Among the KRAS exon 2 wild-type patients 15.31% harboured additional RAS mutations and 12.44% BRAF mutations. The newly designed HRM method used showed a higher sensitivity compared with the sequencing method. Conclusions: The HRM method developed was shown to be a reliable method for KRAS, NRAS and BRAF mutation detection. Furthermore, no difference in the mutation frequency of KRAS, NRAS and BRAF was observed between Greek and Romanian patients with colorectal cancer. 4 5