BRAF Mutation Status Research Articles

Analysis of prognostic factors and establishment of a recurrence risk prediction model for papillary thyroid carcinoma based on BRAF stratification.

Predicting the likelihood of papillary thyroid carcinoma (PTC) recurrence is crucial for improving patient outcomes. The association between the BRAF V600E (BRAF) mutation and PTC recurrence remains controversial. Our goal was to determine prognostic features of PTC patients and construct models for predicting recurrence risk according to BRAF mutation status. A total of 811 PTC patients whose clinical information and survival data were available were included in this study. Independent prognostic variables of PTC identified by screening via LASSO-Cox regression analysis were then used to construct nomograms. The performance of the predictive models was assessed according to the C-index, ROC curve, validation curve, and decision curve analyses. Kaplan-Meier curves were used to analyze differences between patients grouped according to prognostic factors and relapse risk. Multivariate Cox regression analysis demonstrated that extrathyroidal extension (ETE), vascular tumor thrombus, and lymph node yield (LNY) were correlated with recurrence-free survival (RFS) in the BRAF mutation-negative group, while extranodal extension (ENE), number of metastatic lymph node (NMLN), pathological stage, and vascular tumor thrombus were correlated with RFS in the BRAF mutation-positive group. The mutation-stratified predictive models demonstrated better performance than the model without stratification, as indicated by the greater C-index values (0.880 vs. 0.859 vs. 0.753), AUC values (1-year AUC: 0.946 vs. 0.947 vs. 0.758; 3-year AUC: 0.889 vs. 0.871 vs. 0.760; 5-year AUC: 0.845 vs. 0.793 vs. 0.758), and net clinical benefit. The calibration curves at 1 year, 3 years, and 5 years showed good consistency. The bootstrap internal validation had good AUC values exceeding 0.8 and showed a well-fitting calibration curve. Significant differences in RFS were observed between the low-risk and high-risk groups (P < 0.001). Stratifying patients based on their BRAF mutation status can facilitate the development of better and more targeted postoperative management strategies. Nomograms for BRAF mutation positive and negative patients were developed to precisely and consistently predict recurrence risk in PTC patients.

MLH1 gene promoter methylation status partially overlaps with CpG methylator phenotype (CIMP) in colorectal adenocarcinoma.

RAS/BRAF mutations, mismatch DNA repair complex deficiency (MMRd)/microsatellite instability (MSI), and CpG methylator phenotype (CIMP) are key molecular actors in colorectal carcinogenesis. To date, conflicting evidence about the correlations between these molecular features has been reported. A retrospectively selected cohort of 123 CRCs was divided into 3 groups based on the molecular characteristics: MMR proficient (MMRp)/BRAF p.V600E mutated (BRAFmut), MMRd/BRAFmut, and MMRd/BRAF wild type (BRAFwt). MLH1 promoter (pMLH1) methylation status was assessed by pyrosequencing. For 82 samples the CIMP phenotype was evaluated using the EpiTect® MethyLight kit. The MMRd/BRAFmut group showed a higher pMLH1 methylation rate compared to both the MMRd/BRAFwt and the MMRp/BRAFmut groups. Overall, the two MMRd groups had a higher methylation rate compared to the MMRp cases independently from the mutational status of BRAF (p-value <0.0001). The MMRd/BRAFmut group was characterized by a 90.0 % of CIMP high (CIMP-H) tumors of which 97.2 % were pMLH1 methylated. Instead, the MMRd/BRAFwt group presented 50.0 % of CIMP-H adenocarcinomas. Our study demonstrates that pMLH1 hypermethylation, MMRd, BRAFmut and CIMP phenotype do not completely overlap in CRC. These findings further refine the knowledge on the molecular landscape of CRC and may have critical implications also for the clinical management of the disease.

Real world characteristics and outcomes of patients with BRAFV600E-mutant metastatic colorectal cancer in Australia: The COALA project.

70 Background: BRAFV600E-mutant metastatic colorectal cancer (mCRC) represents a unique molecular subset with poor prognosis and less responsiveness to chemotherapy. The BEACON CRC study demonstrated that encorafenib plus cetuximab (EC) significantly improved overall survival (OS) compared to FOLFIRI plus cetuximab in BRAF-mutant mCRC as 2 nd or 3 rd line therapy. This combination was reimbursed in Australia from January 2022. Methods: We analysed data from the Australian TRACC mCRC clinical registry, encompassing patients (pts) with BRAF-mutant mCRC diagnosed in Australia from 2009 to 2024 at 31 hospitals. We assessed baseline demographics, uptake of EC following reimbursement and efficacy analysis of progression-free survival (PFS) and OS for pts that received EC and those who did not. Pts receiving EC on a clinical trial were excluded. Results: Among 2,800 pts with known BRAF mutation status, 365 (13%) were BRAF mutated. The BRAF mutant cohort had a median age of 63 years, with a significantly higher mutation rate in the 20-39 age group than other age groups (age 20-39, 25.5%; age 40-59, 10%; age 60+, 13.2%; P&lt;0.001); 54.5% were female, 58% right-sided tumours, 64% with synchronous metastatic disease and 21.7% deficient mismatch repair (dMMR). Overall, 314 (86%) pts with BRAF mutant tumours and 2,079 (85%) BRAF wild-type (WT) tumours received 1 st line treatment, and 156 (43%) BRAF mutant and 1,144 (47%) BRAF WT pts received 2 nd line therapy. The median OS from diagnosis was 17.3 months for BRAF mutant pts versus 34.7 months for BRAF WT pts (p &lt; 0.0001). All dMMR BRAF mutant pts received 1 st line pembrolizumab since its reimbursement in August 2021. Among 41 BRAF mutant pts progressing on 1 st line therapy since January 2022,(32 pMMR, 9 dMMR), 33 (80%) were subsequently treated with EC, and 4 (10%) did not receive further treatment. OS from the start of 2 nd line therapy was 9.6 months (95% CI: 7.2-12.5) for pts who received EC and 7.1 months (95% CI: 6.4-8.6) for those who received other systemic treatment. Median PFS for pts receiving EC was 4.6 months (95% CI: 3.4-5.8) and 3.3 months (95% CI: 2.5-5.3) for other 2 nd line treatments. Conclusions: mCRC pts younger than 40 have a high rate of BRAF mutation, where 1 in 4 tumours harbour the mutation. BRAF mutant mCRC has a poorer outcome than BRAF WT despite a similar proportion receiving 1 st and 2 nd line treatment. The majority of eligible BRAF mutant mCRC pts received EC following reimbursement in Australia, with PFS and OS outcomes consistent with the BEACON CRC trial.

Prognostic factors for thermo-ablation of liver metastases in colon cancer: A Maltese perspective.

267 Background: Malta is a small Mediterranean archipelago with an annual colon cancer incidence of 300 patients per year and a cumulative 5-year overall survival of 58%. Despite major improvements in therapeutic strategies, colon cancer remains the third leading cause of cancer deaths locally with an annual mortality rate of circa 110 patients per year. Percutaneous thermal ablation of colorectal liver metastases involves exposure of metastatic deposits to temperature extremes using radiofrequency, microwave or cryotherapy. The role of this case series was to examine the prognostic value of clinico-pathological variables to provide insight into patient selection for ablative therapy. Methods: This retrospective analysis of 66 adult patients with colorectal liver oligometastatic disease who underwent thermal ablation between January 2014 and January 2022, reviews demographic and pathological criteria including tumour grade, presence of vascular invasion, mismatch repair and k-RAS &amp; BRAF mutational status, CEA &amp; Ca19.9 assay at time of treatment. Other parameters include metachronous versus synchronous disease, hepatic burden and concurrent chemotherapy. All these co-variables were analysed using the Cox Regression Hazard model to assess their impact on progression free- (PFS) and overall survival (OS). Results: For this patient cohort, the mean PFS and OS were 13.1 and 33.8 months. Our data indicates that most individuals experience progression within the first 10 months, with a gradual decline in PFS for the remaining individuals. A Ca19.9 level within normal limits (p=0.004, HR 0.157-0.700 95% CI) was related to an improved PFS but not OS. The presence of synchronous metastases (p=0.004, HR 0.168-0.715 95% CI) and ablation of one lesion (p=0.013 HR 0.191 (0.052-0.702 95 CI) were favourable prognostic markers for improved OS. k-RAS wt status was a prognostic adverse biomarker for PFS (p=0.002, HR 1.529, 6.686 95% CI). There was a paucity of data for BRAF and MMR status so these were excluded from the final analysis. No statistical survival benefit was noted for patients treated with chemotherapy, and other variables including gender, age, primary tumour sidedness, LVSI or CEA value. Conclusions: There are two surprising trends here- the paucity of survival benefit for the use of chemotherapy in conjunction with ablation and the worse outcome for k-ras wt tumours. We postulate that the absence of BRAF sub-stratification at the time of patient accrual, impacted statistical hazard. This case series suggests that the prognosis and survival of colorectal liver oligometastatic disease is multifactorial and complex, highlighting the need for further genomic and molecular insights.

Influence of Body Mass Index on the Clinicopathological Features of Papillary Thyroid Carcinoma in a Chinese Population.

Background: Previous studies suggested a relationship between obesity and a high risk of thyroid cancer. However, the association between high body mass index (BMI) and the aggressiveness of papillary thyroid carcinoma (PTC) is controversial. In this study, we aimed to investigate the impact of excess BMI on histopathologic aggressiveness of PTC in a Chinese population. Methods: Between January 2015 and September 2020, 4369 PTC patients who were tested for BRAF mutation at Jiangyuan Hospital were enrolled. Logistic regression analyses were used to evaluate the associations between BMI and clinicopathological features of PTC as well as tumor BRAF mutational status. Results: Of 4369 PTC patients, the mean BMI was 24.06 ± 3.49 kg/m2, and BRAFV600E mutations were detected in 3528 (80.8%) patients. BMI ≥24.0 at initial surgery was associated with tumor multifocality and bilaterality, but not with advanced tumor stage, extrathyroidal extension (ETE), ratio of positive lymph nodes >0.3, distant metastasis, or BRAFV600E mutation. Conclusion: Our present study suggested that compared to patients with a normal BMI, overweight and obese patients had a greater risk of multifocality and bilaterality of PTC. No significant associations were observed between higher BMI and the more advanced tumor-node-metastasis stage or BRAFV600E mutational status.

Neoadjuvant or perioperative immunotherapy for resectable melanoma: The need for biomarkers.

Groundbreaking studies have reshaped the treatment landscape for patients with resectable stage ≥IIIB melanoma by demonstrating the benefits of neoadjuvant therapy. Data from the NADINA and SWOG S1801 trials reveal substantial improvements in event-free survival compared to adjuvant therapy alone. These studies employed distinct neoadjuvant immunotherapy approaches -ipilimumab plus nivolumab in NADINA and anti-PD-1 monotherapy in SWOG S1801-highlighting potential differences in efficacy and toxicity. The integration of biomarkers may improve the risk/benefit ratio by enabling personalized treatment selection between ipilimumab plus nivolumab versus anti-PD-1 monotherapy. In metastatic melanoma, molecular biomarkers such as BRAF mutational status and intratumoral PD-L1 expression can help guide treatment decisions; however, their, and other biomarkers', role in the neoadjuvant context has yet to be fully investigated. This commentary underscores the need to investigate whether these molecular markers, along with other biomarkers and clinical parameters, can inform neoadjuvant therapy in resectable stage ≥IIIB melanoma. We propose a coordinated effort involving retrospective analyses of current trials and prospective clinical studies, to define the role of biomarkers. A biomarker-guided approach could optimize efficacy and reduce toxicity, offering personalized treatment options for patients with resectable melanoma. As implementation of neoadjuvant therapy rapidly gains global adoption, we advocate for robust research to ensure that each patient receives the most effective and least toxic therapeutic option.

ASSOCIATION OF MELANOMA AND INCIDENCE OF PARKINSON’S DISEASE: A COHORT STUDY IN US VETERANS

Abstract Melanoma is associated with subsequent Parkinson’s disease (PD) and identifying the clinicopathologic properties of melanoma driving this association could give insight into PD pathophysiology. We undertook a retrospective cohort study of US military Veterans (n=13,737,081) to determine if malignant and premalignant phenotypes were associated with PD by using propensity score matching 4-to-1 and adjustment for competing risk of death and key confounders. Broad cancer categories were obtained from the Oncology VA registry. Data about melanoma were obtained from clinical and pathology reports using manually validated natural language processing. Risk difference and 95% confidence intervals (CI) were estimated by presence or absence of cancer. For broad cancer categories, we found decreased PD incidence associated with multiple malignancies with some novel associations (e.g. HR = 0.4 [0.2,0.5] for myeloma). In terms of melanoma, BRAF and NRAS mutational status was not associated with incident PD. Atypical/dysplastic nevi was associated with increased PD incidence (HR = 1.4 [1.2,1.7]). Melanoma subtypes in the Oncology registry were associated with PD and included superficial spreading type (HR = 1.8 [1.0, 3.3]) and lentigo maligna (HR = 1.6 [1.2, 2.1]). In people with VA pathology reports, this effect was replicated with greater effect size for superficial spreading type (HR = 10.4 [6.5,16.9]) and lentigo maligna (HR = 6.4 [3.4,11.8]) although adjustment for ascertainment bias is needed. These data show PD is associated with melanoma subtypes and precursor lesions and suggest screening for PD could be appropriate in those with melanocytic lesions once early diagnostic tests are developed.

A powerful framework for differential co-expression analysis of general risk factors.

Differential co-expression analysis (DCA) aims to identify genes in a pathway whose shared expression depends on a risk factor. While DCA provides insights into the biological activity of diseases, existing methods are limited to categorical risk factors and/or suffer from bias due to batch and variance-specific effects. We propose a new framework, Kernel-based Differential Co-expression Analysis (KDCA), that harnesses correlation patterns between genes in a pathway to detect differential co-expression arising from general (i.e., continuous, discrete, or categorical) risk factors. Using various simulated pathway architectures, we find that KDCA accounts for common sources of bias to control the type I error rate while substantially increasing the power compared to the standard eigengene approach. We then applied KDCA to The Cancer Genome Atlas thyroid data set and found several differentially co-expressed pathways by age of diagnosis and BRAF mutation status that were undetected by the eigengene method. Collectively, our results demonstrate that KDCA is a powerful testing framework that expands DCA applications in expression studies.

Electrochemical Detection of BRAF V600E Mutation in a Liquid Biopsy Format: LAMPing the Way to an Improved Diagnostics and Treatment

AbstractThe BRAF V600E substitution, predominantly found in melanoma and metastatic colorectal cancer (mCRC), is associated with aggressive traits and poor prognosis. Molecular testing of the BRAF mutational status is crucial for guiding therapeutic decisions, driving the necessity for new rapid diagnostic technologies. Here, a novel LAMP‐based assay is presented for fast and selective amplification of mutated DNA, coupled with simple electrochemical detection on electrode biochips. The assay demonstrated high selectivity across cell lines and in 51 cancer patients, detecting mutations in both tumor tissue DNA and circulating tumor DNA (ctDNA) in serum. Clear discrimination between the mutated and wild‐type sequences is achieved, in perfect agreement with either NGS or ddPCR. The bioassay offers a simple and efficient alternative to traditional PCR methods for point‐of‐care diagnostics, particularly in settings with limited equipment and resources.

The efficacy and safety of first-line metastatic melanoma treatment with ipilimumab + nivolumab versus nivolumab in a real-world setting.

The Checkmate 067 randomized controlled trial, published in 2015, demonstrated improved progression-free survival and numerically, although not statistically, superior overall survival for ipilimumab + nivolumab. The objective of this study was to compare the efficacy and safety of nivolumab to ipilimumab + nivolumab as first-line treatment for metastatic melanoma in a real-world setting. Patients were prospectively included in the French Melbase cohort from 2013 to 2022. Eligible patients were those in first-line treatment for stage-IIIc or -IV melanoma, undergoing immunotherapy with nivolumab or ipilimumab + nivolumab. The primary endpoint was overall survival at 36 months. The secondary endpoints included progression-free survival at 36 months, best radiological response, and safety analyses. We conducted a propensity score using the IPTW method to overcome the various confounding factors and also a subgroup analysis (brain metastasis, LDH levels, and BRAF mutation status). A total of 406 patients were treated with nivolumab, and 416 with ipilimumab + nivolumab. Overall survival at 36 months was higher in the ipilimumab + nivolumab group (57.1%, ([95%CI 50.7-64.2]) than in the nivolumab group (46.6% [95%CI 41.6-52.1]), HR 1.4[1.1;1.8]. Progression-free survival at 36 months was significantly improved in the ipilimumab + nivolumab group (42.3%) compared to the nivolumab group (21.9%), with a HR 1.6[1.4;1.9]. The objective response rate was similar for the two groups (44%). The overall incidence of side effects was comparable (82 vs. 84%), and severe toxicity (grade ≥ 3) was more frequent, though not significantly so, in the ipilimumab + nivolumab arm (29% vs. 41%). Our results are consistent with those from the Checkmate 067 study, except for the objective response rate and the incidence of toxicities, which proved to be lower in our analysis.

Molecular alterations influencing the prognostic outcome in small pancreatic cancer (≤ 2 cm)

Abstract Purpose Pancreatic cancer (PC) is the most lethal cancer. The prognosis of small PC (tumor smaller than 20 mm) is better than that of larger PC tumors, indicating the importance of detecting early-stage PC for patient outcome. The aim of this study was to elucidate the molecular features in small PC (≤ 20 mm). Methods This study included 79 PC tumors (≤ 20 mm in pathological examination) resected between 2004 and 2022. c-Myc, Caveolin-1, Smad4, and Thrombospondin-1 were examined by immunostaining. These molecular alterations were compared in PC patients with tumor size ≤10 mm (n = 11) (14%) and 10 mm &lt; tumor size ≤20 mm (n = 68) (86%). Mutation analyses of KRAS, PIK3CA, and BRAF were performed by pyrosequencing in 22 PCs. Results PC with 10 mm &lt; tumor size ≤20 mm showed significantly worse overall survival and disease-free survival than PC with tumor size &lt;10 mm (P = 0.024 and P = 0.028). Tumor c-Myc and stromal Caveolin-1 expressions were significantly increased in tumors larger than 10 mm (P = 0.02 and P = 0.04). c-Myc and Caveolin-1 expressions were associated with poor disease-free survival and overall survival. KRAS, PIK3CA, and BRAF mutation status did not differ between the two groups. Conclusions Tumor c-Myc and stromal Caveolin-1 overexpressions were detected in tumors larger than 10 mm. Their overexpressions were associated with worse prognosis even in small PC. These molecular alterations in small PC may be a clue for the detection of early-stage PC.

Elucidating the Prognostic Role of BRAF V600E and the Activation Status of the Downstream MAPK Pathway in PTC: A Study from a Tertiary Centre in India.

Papillary thyroid carcinoma (PTC) has an excellent prognosis, but few cases are treatment-resistant. To check the applicability of combined BRAF V600E and MEK-targeted therapy, the current study correlated BRAF V600E with the MAPK pathway activation status in a cohort of PTCs. The prognostic relevance of BRAF V600E and the usability of immunohistochemistry (IHC) for detecting the mutation were also assessed. Randomly selected 50 PTC and 15 non-PTC cases were re-classified according to the 2022 WHO classification. The BRAF mutation status was compared with the IHC of BRAFV600E, pERK1/2, pMEK1/2, and clinicopathological variables, including response to radioactive iodine and disease-free survival. BRAF V600E mutation was present in 38%. Most (87.8%) cases were immunopositive for pMEK1/2 and 40% for pMEK1/2. Although BRAF V600E mutation did not correlate with the MAPK activation status, it had an adverse impact on tumour sensitivity to radioiodine (P < 0.05). Five of the seven radioiodine-resistant tumours were BRAF V600E-mutated. An Allred cut-off score of 7 had a sensitivity of 100% and a specificity of 84% for detecting the mutation by IHC. All the non-PTC cases were BRAF-wild type, but 20% showed weak immunopositivity for mutated protein and were scored 6. BRAF V600E-mutated PTCs are more likely to be RAI-resistant. MAPK pathway activation status did not vary significantly with BRAF mutation. Immunopositivity for pMEK1/2 in most suggests a scope for MEK1-targeted therapy in recalcitrant PTC cases even in the absence of the BRAF mutation. In addition, IHC is a reliable technique for detecting BRAF V600E mutation but needs validation by correlation with molecular studies.

The Prognostic Impact of Tumor Size and BRAF Mutational Status in Middle Eastern Differentiated Thyroid Cancer.

Tumor size at diagnosis has been widely used as a major mortality risk factor in risk stratification of DTC. The current study was designed to analyze whether tumor size at diagnosis is a major prognostic factor in Middle Eastern DTC. We conducted a comparative study of the relationship between tumor size at diagnosis and event free survival (EFS) with respect to BRAF status in 1709 consecutive patients treated surgically for DTC. Patients were divided into four groups according to the size of tumor and BRAF mutation status: Group 1 (≤4 cm without BRAF mutation), Group 2 (≤4 cm with BRAF mutation), Group 3 (>4 cm without BRAF mutation) and Group 4 (>4 cm with BRAF mutation). Predictors of EFS were compared using the Log-rank test and Cox proportional hazards models. Tumor size >4cm was associated with adverse clinico-pathologic characteristics, such as older age, male gender, bilateral tumors, extrathyroidal extension, lymphovascular invasion, advanced tumor stage and persistent/recurrent disease. Tumor size was also inversely associated with BRAF mutation. Both tumor size (> 4cm) and BRAF mutation were associated with EFS on univariate analysis. On subgroup analysis, larger tumor size was an independent predictor of EFS (Group 3 vs. Group 1), irrespective of BRAF mutation status. Also, within the BRAF mutant tumors, larger tumor size was still an independent predictor of EFS (Group 4 vs. Group 2). Tumor size is an independent predictor of EFS in Middle Eastern DTC patients, regardless of BRAF mutational status.

Novel Approach for Detection of BRAF and KRAS Mutations Using ARMS (amplification refractory mutation system) Primers for Colon Cancer Therapeutic Response Prediction

Abstract Introduction/Objective KRAS and BRAF V600E mutations are confirmed predictive biomarkers for anti-EGFR monoclonal antibody therapy response and prognosis for colorectal cancer and often considered mutually exclusive with few exceptions (0.05% of mCRC cases). We have developed a multiplex PCR assay utilizing ARMS (amplification refractory mutation system) Primers and Probes for screening and further testing of BRAF mutation status in a 2 step process. Methods/Case Report The Multiplex PCR Assay or BRAF/KRAS mutation was designed as a 5 color assay. Primers and Probes consisted of Wild Type Kras and wild and mutant (V600E/V600K) type BRAF primers and probes. For specimen, Positive and negative control standards made of engineered positive control plasmids for wild type KRAS &amp; BRAF and mutant type V600E &amp; V600K BRAF as well as internal control of bet actin and 2 Patient samples with known BRAF V600E mutant status were run. Mutant type V600E &amp; V600K BRAF were mixed with wild type V600 controls in different ratios as well. Results were interpreted as BRAF wild KRAS wild/ BRAF wild KRAS mutant/ BRAF V600E KRAS wild/BRAF V600K KRAS wild type and NGS was recommended in cases of concurrent BRAF and KRAS mutations. Results (if a Case Study enter NA) Positive/negative/internal control standards for wild type KRAS &amp; BRAF and mutant type V600E &amp; V600K BRAF showed expected results in both CFX 96 and ABI 7500 runs in triplicates. Graphs were able to determine separate Ct values and sigmoidal amplification plots that readily differentiated KRAS wild/BRAF wild/ BRAF V600E types. Mutant type V600E &amp; V600K BRAF that were mixed with wild type V600 controls in different ratios all showed expected results and were able to differentiate between wild and mutant types with accuracy. Also, the 2 patient samples with known (Sanger sequencing-diagnosed) V600E mutation status showed positive Ct values for KRAs wild and V600E probes and negative CT values for V600 wild/V600K probes. No invalid runs or unexpected control results were observed. Conclusion The ARMS BRAF/KRAS multiplex test is a efficient, accurate 2 step assay. In the patients and positive and negative controls prepared, it was able to reliably diagnose KRAS and BRAF mutation status in a matter of 4 hours running time. Given the fact that this is a cost effective assay that can utilize most commonly used Real time PCR machines this test may be clinically implemented if larger clinical trials utilizing tissue samples from a larger number of colon cancer patients are conducted.

Survival of Patients with Metastatic Melanoma Treated with Ipilimumab after PD-1 Inhibitors: A Single-Center Real-World Study

Background: When monotherapy with PD-1 inhibitors in metastatic melanoma fails, there are currently no standard second-line choices. In case of the unavailability of clinical trials, ipilimumab represents a possible alternative treatment. Methods: We collected data of 44 patients who received ipilimumab after the failure of PD-1 inhibitors from July 2017 to May 2023 at our Institute. Overall survival (OS), progression-free survival (PFS), and post-progression survival (PPS) based on BRAF or NRAS mutation status, sex, and the presence of brain metastases were estimated using the Kaplan–Meier method. Cox regression was used to evaluate independence in multivariate analysis. The objective response rate (ORR) was estimated based on RECIST 1.1. Results: Among the 44 patients enrolled in this study, 28 BRAF-wildtype, 9 BRAF-mutated, and 7 NRAS-mutated patients were identified. OS analysis showed a significant difference between wildtype and BRAF- or NRAS-mutated patients: 23.2 months vs 5.3 and 4.59, respectively, p = 0.017. The presence of brain metastases and BRAF or NRAS mutation were independent factors for mortality in multivariate analysis. Conclusions: In case of failure to enroll patients in innovative clinical trials, second-line ipilimumab still represents an effective therapy in patients with metastatic wildtype melanoma and in the absence of brain metastases.

DNA methylation-array interlaboratory comparison trial demonstrates highly reproducible paediatric CNS tumour classification across 13 international centres.

DNA methylation profiling, recently endorsed by the World Health Organisation (WHO) as a pivotal diagnostic tool for brain tumours, most commonly relies on bead arrays. Despite its widespread use, limited data exist on the technical reproducibility and potential cross-institutional differences. The LOGGIC Core BioClinical Data Bank registry conducted a prospective laboratory comparison trial with 12 international laboratories to enhance diagnostic accuracy for paediatric low-grade gliomas, focusing on technical aspects of DNA methylation data generation and profile interpretation under clinical real-time conditions. Four representative low-grade gliomas of distinct histologies were centrally selected, and DNA extraction was performed. Participating laboratories received a DNA aliquot and performed the DNA methylation-based classification and result interpretation without knowledge of tumour histology. Additionally, participants were required to interpret the copy number profile derived from DNA methylation data and conduct DNA sequencing of the BRAF hotspot p.V600 due to its relevance for low-grade gliomas. Results had to be returned within 30 days. High technical reproducibility was observed, with a median pairwise correlation of 0.99 (range 0.94-0.99) between coordinating laboratory and participants. DNA methylation-based tumour classification and copy number profile interpretation were consistent across all centres, and BRAF mutation status was accurately reported for all cases. Eleven out of 12 centres successfully reported their analysis within the 30-day timeframe. Our study demonstrates remarkable concordance in DNA methylation profiling and profile interpretation across 12 international centres. These findings underscore the potential contribution of DNA methylation analysis to the harmonisation of brain tumour diagnostics.

Evaluation of Pan-RAF Inhibitor LY3009120 on Human Uveal Melanoma Cell Line 92-1.

Uveal melanoma (UM) represents a prevailing primary intraocular malignancy, with a limited median overall survival among metastatic patients, and most tumors lack RAF/RAS mutations. The pan-RAF inhibitor LY3009120 has demonstrated valuable anti-tumor effects in a wide range of RAF/RASmut and wild-type (WT) tumor models. This study aimed to evaluate the antitumor effect of LY3009120 on 92-1 UM cell line. The effect of the pan-RAF inhibitor LY3009120 on cell proliferation, metabolic activity, biomass, early and late apoptosis/necrosis, and morphology was characterized in vitro (0.1-5 μM for 48 h/72 h). Furthermore, targeted panel sequencing was used to characterize the mutational landscape of the human 92-1 UM cell line. LY3009120 showed a significant concentration-dependent anti-proliferative effect on 92-1 cells. Cell proliferation and viability were significantly reduced at the lowest effective concentration of 0.5 μM (at 48 and 72 h, p<0.001). Furthermore, LY3009120 caused significant early apoptosis and late apoptosis/necrosis in 92-1 cells at 5 μM. Except for TP53, NGS showed that all 49 additional analysed genes (Oncomine myeloid panel) of 92-1 were wild-type, including BRAF, NRAS, and KRAS. The pan-RAF inhibitor LY3009120 demonstrated a significant anti-tumor effect on human UM cell line 92-1 independent of the molecular BRAF and RAS mutational status.

An Automated Real-Time PCR Assay versus Next-Generation Sequencing in the Detection of BRAF V600 Mutations in Melanoma Tissue Samples.

Next-generation sequencing (NGS) is the most commonly used method for determining BRAF mutational status in patients with advanced melanoma. Automated PCR-based methods, such as the IdyllaTM system, are increasingly used for mutation diagnostics, but it is unclear what impact the choice of diagnostic method has on the management of melanoma. To compare the concordance rate of BRAF V600 mutational analysis using IdyllaTM and NGS and to analyze the technical and clinical turnaround time. The clinical relevance is compared by analyzing the impact on the treatment decision. In this monocentric prospective cohort study, the BRAF mutation status of 51 patients was determined using both methods in parallel. BRAF V600 mutation was detected in 23/51 cases (45%). IdyllaTM showed a 100% concordant result with a faster turnaround time (0.2 days) compared to NGS (12.2 days). In general, less tumor material was required for IdyllaTM than for NGS. Most patients received immunotherapy as a first-line therapy regardless of the BRAF V600 status. IdyllaTM testing proved to be a reliable and rapid alternative to NGS in the determination of BRAF V600 mutation. Although BRAF. status was available earlier, this had no influence on the treatment decision in most cases.

Serum Uric Acid Level May Be a Predictive Factor for BRAF V600E Mutation in Older Patients with Metastatic Colorectal Cancer: An Exploratory Analysis

Introduction: This study aimed to show the relationship between the serum uric acid level measured at diagnosis and the BRAF mutation status in the primary tumor tissue in patients with metastatic colorectal cancer. Methods: In this retrospective cross-sectional study, 264 patients (64% male) whose serum uric acid level was measured at the time of diagnosis and whose BRAF mutation status in the primary tumor was determined were included. Results: The BRAF mutation rate was 14% (n = 37). The median serum uric acid levels of all patients were 6.9 mg/dL (25%, 75% percentile range 3.7, 8.2). The serum uric acid level cut-off value was 6.6 mg/dL. Sensitivity and specificity for BRAF mutated patients were 84% and 27%, respectively. These rates were calculated as 85% and 70% in BRAF-mutated patients aged 65 and over. There was a significant correlation between BRAF mutation and high serum uric acid level, female gender, tumor located in the ascending colon, and multiple metastatic sites. The independent factors affecting BRAF mutation were age 65 and over, tumor in the ascending colon, and high serum uric acid level. Conclusion: As a result, we concluded that high serum uric acid level measured during diagnosis in metastatic colorectal cancer is an accessible and economical biomarker that can predict BRAF mutation in patients aged 65 and over.

Regorafenib synergizes with TAS102 against multiple gastrointestinal cancers and overcomes cancer stemness, trifluridine-induced angiogenesis, ERK1/2 and STAT3 signaling regardless of KRAS or BRAF mutational status.

Single-agent TAS102 (trifluridine/tipiracil) and regorafenib are FDA-approved treatments for metastatic colorectal cancer (mCRC). We previously reported that regorafenib combined with a fluoropyrimidine can delay disease progression in clinical case reports of multidrug-resistant mCRC patients. We hypothesized that the combination of TAS102 and regorafenib may be active in CRC and other gastrointestinal (GI) cancers and may in the future provide a treatment option for patients with advanced GI cancer. We investigated the therapeutic effect of TAS102 in combination with regorafenib in preclinical studies employing cell culture, colonosphere assays that enrich for cancer stem cells, and in vivo. TAS102 in combination with regorafenib has synergistic activity against multiple GI cancers in vitro including colorectal and gastric cancer, but not liver cancer cells. TAS102 inhibits colonosphere formation and this effect is potentiated by regorafenib. In vivo anti-tumor effects of TAS102 plus regorafenib appear to be due to anti-proliferative effects, necrosis and angiogenesis inhibition. Growth inhibition by TAS102 plus regorafenib occurs in xenografted tumors regardless of p53, KRAS or BRAF mutations, although more potent tumor suppression was observed with wild-type p53. Regorafenib significantly inhibits TAS102-induced angiogenesis and microvessel density in xenografted tumors, as well inhibits TAS102-induced ERK1/2 activation regardless of RAS or BRAF status in vivo. TAS102 plus regorafenib is a synergistic drug combination in preclinical models of GI cancer, with regorafenib suppressing TAS102-induced increase in microvessel density and p-ERK as contributing mechanisms. The TAS102 plus regorafenib drug combination may be further tested in gastric and other GI cancers.