Prevalence Of RAS Mutations Research Articles

Crystallographic Studies of KRAS in Complex with Small Molecules and RAS-Binding Proteins.

RAS proteins play a vital role in regulating downstream signaling and essential cellular processes, positioning them as key players in normal cellular physiology and disease development. Among the various isoforms of RAS, KRAS stands out as one of the most frequently mutated genes in human cancer. The prevalence of RAS mutations in cancer often involves single amino acid substitutions at codons 12, 13, or 61. These mutations disrupt the RAS protein's inherent ability to transition between its active and inactive states, resulting in a constant activation signal and driving uncontrolled cell growth. Crystallization and structural analysis of KRAS with inhibitors and RAS-binding proteins play a pivotal role in unraveling the structural and mechanistic details of KRAS function, aiding in drug discovery efforts, and advancing our understanding of KRAS-driven diseases. Here, we present our experimental methodology for crystallizing KRAS in the presence of covalent or non-covalent small molecules and proteins acting as effectors or regulators of RAS. We detail the techniques for successful crystallization and the subsequent optimization of crystallization conditions. The resulting crystals and their structures will provide valuable insights into the key interactions between KRAS and its partner proteins or potential inhibitors, offering a foundation for developing targeted therapies that are more potent and selective against KRAS-driven cancers.

Venetoclax (VEN) Improves Response Rates but Not Survival in Patients with Chronic Myelomonocytic Leukemia (CMML) Treated with Hypomethylating Agents (HMA): A Multicenter, Propensity Score Analysis

Introduction HMAs are the only approved therapy for CMML. In order to improve response rates and survival, the addition of VEN has been utilized in both CMML and after blast transformation (CMML-BT) with conflicting results in small single center studies. To clarify the role of upfront HMA+VEN in CMML and CMML-BT, we performed a large multicenter analysis. Methods We included patients with HMA-naïve CMML and CMML-BT treated with combination HMA and VEN from 4 academic centers. The primary outcome was overall survival (OS) with secondary outcomes including overall response rate (ORR) by 2015 MDS/MPN IWG criteria for CMML and 2022 ELN criteria for CMML-BT. To better appreciate the benefit of adding VEN to HMA treatment, we also performed 1:1 propensity score matched (PSM) analysis of patients treated with HMA alone as upfront therapy for CMML or CMML-BT. Matching was based on age, sex, ECOG performance status, CPSS-Mol, blood counts, peripheral and bone marrow blasts. Results Eighty-nine patients were treated with frontline HMA+VEN, 51 CMML and 38 CMML-BT (5 had received HMA alone for prior CMML). The median age was 71 years, 70% were male, and 83% had ECOG 0-1. Among CMML patients, 55% had MP-CMML subtype, 65% had CMML-2 and 12%, 30%, 58% were CPSS-mol intermediate-1, intermediate-2 and high risk, respectively. In the entire cohort, ASXL1 (63%), TET2 (47%), SRSF2 (39%) and RUNX1 (27%) were the most commonly mutated genes and 55% of patients harbored a RAS mutation ( KRAS [20%], NRAS [18%], CBL [13%], NF1 [11%], PTPN11 [9%]). For HMA combination partner, 46 (52%) were treated with azacitidine, 35 (39%) with decitabine, and 8 (9%) with both during their VEN course. During a median follow-up time of 15.8 months (mos) 52 patients (58%) died: 23 (45%) with CMML and 29 (76%) with CMML-BT. Among CMML patients, the ORR was 90%, including complete remission (CR) in 22%, complete cytogenic remission (CCR) in 4%, optimal marrow response (CMR) in 50%, and partial marrow response (PMR) in 6%. The OS of CMML patients was 25.1 mo (95% CI 15.7 - NR). A PSM population of HMA patients matched to HMA+VEN treated patients showing similar baseline characteristics. ORR was significantly higher with HMA+VEN (95%) as compared those treated with HMA alone (46%), p < 0.001. However, there was no significant OS difference between HMA+VEN vs HMA alone (19.1 mos vs 19.1 mos, p = 0.85, Figure 1). For CMML-BT patients, the ORR of HMA+VEN was 81% including CR in 17%, CR with partial hematologic recover (CRh) in 3%, CR with incomplete hematologic recovery (CRi) in 28%, morphologic leukemia-free state (MLFS) in 22% and partial response (PR) in 11%. The OS of CMML-BT patients was 8 mos (95% CI 4.6 - 26.2). A PSM population of CMML-BT patients treated with HMA alone showed a significantly increased ORR with HMA+VEN (82% vs 40%, p = 0.039). However, there was no difference in OS (7.49 mos vs 9.26 mos, p = 0.83, Figure 2). Among HMA-VEN treated patients, achievement of a CR was associated with an improved OS in the CMML (HR 0.31, 95% CI 0.13 - 0.75) and CMML-BT (HR 0.37, 95% CI 0.19 - 0.74). No predictors of response were identified, including the presence of a RAS mutation. In terms of safety, 7 patients (7.9%) had a grade 3/4 (G3/4) neutropenic infection and 2 patients (2.2%) had a G3/4 hemorrhagic event while being treated with HMA+VEN. A total of 29 patients went on to receive allogeneic transplant (alloSCT): 22 (43%) with CMML and 7 (18%) with CMML-BT. CMML patients who underwent alloSCT after HMA+VEN were younger as compared to non-transplanted patients (64 vs 74 years, p < 0.0001), but had otherwise similar baseline characteristics including stage, CPSS-mol category and prevalence of RAS mutations. Similarly, CMML-BT patients bridged to alloSCT were younger (66 vs 74 years, p < 0.002) and had a better ECOG performance status (p = 0.047). Of patients who received alloSCT, 10 patients (34%) died, 5 with CMML and 5 with CMML-BT. The median OS after alloSCT was 26.7 mo. Conclusions Although the use of VEN with HMA in the upfront treatment of CMML and AML secondary to CMML results in higher response rates compared to HMA alone, this combination does not improve survival based on this multi-center PSM analysis. The current sample size of CMML-BT may limit detection of OS differences between treatment groups. The role of VEN to reduce blasts and serve as a bridge to alloSCT, particularly after HMA failure, requires further investigation in larger studies in both the CMML and CMML-BT populations.

Genomic characterization of papillary thyroid carcinoma in Chinese Shandong population.

e18085 Background: With the emergence of mutation-based diagnosis and therapies for patients with papillary thyroid carcinoma (PTC), molecular profiling has become an important component of care. Although next-generation sequencing (NGS) gene panels are accessible to clinicians, the key genetic alterations of PTC in Chinese Shandong population were rarely descripted. Methods: A total of 127 papillary thyroid carcinoma patients were enrolled in this cohort, including 49 (38.6%) males and 78 (61.4%) females from January 2020 to May 2022. The median age were all 48 years. Formalin-fixed, paraffin-embedded (FFPE) tumor tissues were all collected and analyzed from these patients. Somatic and germline mutations were identified including 38 genes associated with tumor development. Sequencing data were analyzed to call tumor specific single nucleotide variants (SNV), small insertions and deletions (InDels), copy number alterations (CNA), gene fusions and rearrangements. Results: A total of 117 out of 127 papillary thyroid carcinoma were NGS-positive (92.1%), with a high prevalence of BRAF V600E mutations (52.0%, 66/127) and MLH1 mutations (23.6%, 30/127), and a low prevalence of RAS mutations (10.2%, 13/127)， ROS1 (9.4%, 12/127). Overall, 257 somatic mutations including 249 SNVs (6 splicing, 4 stopgain, 4 frameshift，and 235 nonsynonymous), 3 fusions, 3 deletions, and 1 insertions in 38 genes were identified in the PTC samples. Gene fusions, involving the RET and NTRK1 genes, were identified in three PTCs (2.4%), including NTRK1-TPM3, NTRK1-SQSTM1 and CCDC6-RET. And the three fusions patients were all BRAF V600E-negative cases. Furthermore, in our cohort, BRAF V600E mutation carriers also owned other actionable, the most frequent one was MLH1 (34.8%), followed by RET (15.2%), ROS1 (10.6%), and ATM (10.6%). We also found that three PTC patients carried germline variants of uncertain significance, including ATM (p.L857P), MET (p.L211W) and RET (p.C515R), which are heterozygous mutations. Conclusions: We described the distinct molecular profile in the Chinese Shandong population with PTC. Most patients had at least one potentially actionable mutation, predominantly driven by BRAF V600E and MLH1 alterations. These findings may help in moving toward the more comprehensive and effective personalized treatment of papillary thyroid carcinoma in Chinese.

Worldwide Prevalence and Clinical Characteristics of RAS Mutations in Head and Neck Cancer: A Systematic Review and Meta-Analysis.

In light of the development of RAS inhibitors, a reliable assessment of the prevalence of RAS mutations and their correlation with the clinical features of patients with HNC is crucially needed. This meta-analysis compiles the findings of 149 studies with over 8500 HNC patients and assesses the global prevalence of mutations in the HRAS, KRAS and NRAS genes. The available data were stratified according to geographical region, clinical features, and tumor characteristics, including human papillomavirus (HPV) infection status and tumor stage. In addition, the distribution of codon substitutions in each RAS gene was assessed. The estimated mutation rate is highest for HRAS (7%), followed by KRAS (2.89%) and NRAS (2.20%). HRAS prevalence in South Asia (15.28%) is twice as high as the global estimate. HRAS mutations are more prevalent in oral cavity and salivary gland tumors. In contrast, KRAS mutations are found more frequently in sinonasal tumors, and NRAS mutations are found chiefly in tumors of the nasopharynx. OR analyses show a significant association between HRAS mutations and a high tumor stage (OR=3.63). In addition, there is a significant association between HPV-positive status and KRAS mutations (OR=2.09). This study highlights RAS as a potential therapeutic target in certain subsets of HNC patients.

Treatment pathways and associated costs of metastatic colorectal cancer in Greece

ObjectivesColorectal cancer (CRC) is the second leading cause of cancer in Europe, with 1.931.590 people newly diagnosed in 2020. The purpose of this study is the investigation of treatment options and healthcare resource metastatic CRC (mCRC) in Greece.MethodsThis study is based on the information collected in November 2020 by an expert panel comprising of 6 medical oncologists from major public and private centers around Greece. A 3-round survey was undertaken, according to Delphi method. The treatment phases studied were: pre-progression; disease progression and terminal care. Pharmaceutical costs and resource utilization data were considered from the perspective of the Greek National Services Organization (EOPYY). RESULTS: Experts agreed that the anticipated prevalence of RAS mutation in mCRC is 47% (30% RAS/BRAF WT Left, 17% RAS/BRAF WT Right); 8% BRAF while, MSI-H/dMMR are found in 5% of mCRC tumors. Based on mutational status, 74.8% of patients receive biological targeted therapies in combination with fluoropyrimidine/based combination chemotherapy, as 1st line treatment, and 25.2% combination chemotherapy alone. At 2nd line, 58.6% of patients receive biological targeted therapies in combination with chemotherapy, 25.4% immunotherapy, 11% combination chemotherapy and 5% biological targeted therapies. At 3rd line 56% of patients receive combination chemotherapy, 28% biological targeted therapies, 10% biological targeted therapies in combination with chemotherapy and 6% immunotherapy. The weighted annual cost (pharmaceuticals and resource use cost) in 1st line per mCRC patient was calculated at €28,407, in 2nd line €33,568, in 3rd line €25,550. The annual cost beyond 3rd line per patient regardless mutation was €19,501 per mCRC patient.ConclusionsmCRC is a societal challenge for healthcare systems as the treatment is more prolonged but expand patients’ survival. Thus, reimbursement decisions should be based not just on the cost of the treatment, but on the magnitude of the benefit of its treatment on patients’ survival and quality of life.

Abstract 1999: DIRAS3 induces autophagy and enhances sensitivity to anti-autophagic therapy in KRAS-driven pancreatic and ovarian carcinomas

Abstract Mutant KRAS drives human cancers from several sites, including pancreatic ductal adenocarcinoma (PDAC) and low-grade serous ovarian cancer (LGSOC). Despite the prevalence of RAS mutations in many different cancers, effective RAS-targeted treatment remains a challenge. More than 90% of PDACs are driven by mutant KRAS. Genetic suppression of KRAS or pharmacological inhibition of its effectors, ERK/MAPK, enhances the reliance of PDAC on autophagy that can provide a target for treatment. DIRAS3 (ARHI) is an imprinted tumor suppressor gene that is downregulated in pancreatic and ovarian cancers. When re-expressed, DIRAS3 blocks cancer cell proliferation, inhibits motility, and induces autophagy by multiple mechanisms. DIRAS3 inhibits RAS signaling by heterodimerizing directly with KRAS, inhibiting homodimerization of KRAS and disrupting KRAS nanoclustering. In this study, we tested whether inhibiting RAS signaling by re-expressing DIRAS3 would sensitize RAS-driven cancers to autophagy inhibition. Re-expression of DIRAS3 decreased phosphorylation of Erk1/2 (pThr202/Tyr204), inhibiting the RAF-MEK-ERK signaling pathway. Re-expression of DIRAS3 induced autophagic flux indicated by an increased LC3-mCherry/eGFP ratio. DIRAS3 re-expression decreased LKB1 (pS428) and increased AMPK (pT172) and ULK1 (pS555), suggesting that induction of autophagic flux may be partly mediated by the activation of the energy-sensing LKB1-AMPK1-ULK1 axis following suppression of upstream RAF-MEK-ERK signaling. In addition, we confirmed that DIRAS3-induced autophagy enhanced the sensitivity of cancer cells to autophagy inhibition as evidenced by decreasing the IC50 of both chloroquine (CQ) and di-chloroquine (DC661) in human KRAS-driven PDAC and LGSOC. The greater potency of DC661, blocking autophagy with an IC50 in the low nanomolar range, should prove more effective for combinatorial therapies in vivo. Citation Format: Gamze Bildik, Joshua P. Gray, Weiqun Mao, Robert C. Bast, Zhen Lu. DIRAS3 induces autophagy and enhances sensitivity to anti-autophagic therapy in KRAS-driven pancreatic and ovarian carcinomas [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 1999.

Clinical and Molecular Characterization of Primary Plasma Cell Leukemia

Primary plasma cell leukemia (pPCL) is a rare form of multiple myeloma (MM) that is characterized by aggressive behavior and short overall survival (OS) despite significant improvement in anti-MM therapy. Due to its low incidence of 1-2% of all MM patients, most previous studies reported only on a small number of pPCL patients and the physiopathology as well as best treatment approaches remain unclear. In order to improve outcome in this patient population it will be necessary to evaluate molecular mechanisms underlying pPCL, which could potentially serve as therapeutic targets. The goal of the present study was to address clinical parameters and survival in pPCL patients. We further analyzed molecular characteristics by gene expression analysis and exome sequencing with the aim to detect distinct markers in pPCL.We retrospectively evaluated 84 pPCL patients who were treated at the Myeloma Institute in Little Rock, and had at least cytogenetic (by karyotype) and outcome data available. The cytogenetic analysis showed a complex karyotype (>3 chromosomal aberrations) in 88% of the patients and hypodiploidy in 54% patients. Deletion 13 was very common and seen in 54% of patients and translocation 11;14 and deletion 17p were evident in 22% and 19% of patients respectively. These results are consistent with previous reports and show a higher prevalence of poor prognosis alterations in pPCL patients compared to non-pPCL MM patients. Clinical and biological characteristics of pPCL patients showed very high proportion of ISS 3 stage (86%) and bone marrow (BM) plasma cell infiltration of at least 70% in 77% of patients. Median overall survival (OS) of this pPCL population was 17 months. When restricting the analysis to patients <70 years of age that underwent high dose chemotherapy induction regimen followed by ASCT (n=59) the median OS was still short at 24 months.We further evaluated gene expression profiling (GEP) from BM aspirates, which was performed on 54 patients and used the previously described 70-gene classifier (GEP70) for risk assessment. Surprisingly 40% of the patients had a low risk (LR) GEP70 profile, suggesting that pPCL might be more heterogeneous than previously thought or that some pPCL - with less aggressive biology - is caused from overcrowding of extensively involved BM space with leakage into the peripheral blood. Though median OS in LR pPCL patients was significantly higher than in HR patients (30 vs 16months, p=0.025), it still remained short suggesting that other parameters not identified by GEP70 influence outcome. In order to identify a gene signature specific to pPCL, we next studied differential gene expression of 18670 probes in BM samples from pPCL patients and compared them with healthy controls. There were 130 differentially expressed probes with at least 5 fold change at a false discovery rate of <0.01. Of these only 13 probes were upregulated with PHF19 (chr9q33.2) being the most upregulated gene in pPCL. Interestingly, when analyzing expression of PHF 19 throughout MM stages, we show that expression levels of PHF19 are similar in healthy controls and in patients with monoclonal gammopathy of undetermined significance (MGUS) as well as with smoldering MM (SMM) and increase in patients with MM and achieve highest levels in pPCL patients, suggesting that PHF19 mediates pPCL tumorigenicity and is a marker of disease aggressiveness.We further analyzed data on targeted exome sequencing which was performed on 10 pPCL patients. A mutation in the RAS oncogene was present in 9 patients (KRAS n=5, NRAS n=3, one patient had both mutation), which is a much higher prevalence of RAS mutations than the previously described ~40% in non pPCL MM. TP53 mutations were second most common and seen in 5 patients.In summary, this is the largest series of patients with pPCL reported so far. It confirms the poor prognosis of pPCL, even in young patients that were treated with an aggressive approach and in GEP70 defined LR disease. PHF19, a regulator of Polycomb Repressive Complex 2 (PRC2) that leads to transcriptional repression through H3K27 hypermethylation, was highly expressed in pCPL patients and its role in disease pathogenesis should be further evaluated. Further, targeted exome sequencing showed a RAS mutations in 90% of patients and though the patient number for this analysis was small, already available therapy targeting the RAS-pathway should be evaluated in pPCL patients. DisclosuresDavies:Seattle Genetics: Consultancy, Honoraria; Takeda: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees; Amgen: Consultancy, Honoraria; Bristol-Myers: Consultancy, Honoraria; Celgene: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees. Morgan:Bristol Myers: Consultancy, Honoraria; Takeda: Consultancy, Honoraria; Celgene: Consultancy, Honoraria, Research Funding.

Genetic Profiles of Aggressive Variants of Papillary Thyroid Carcinomas.

Simple SummaryAggressive variants of papillary thyroid carcinoma (PTC) are associated with unfavorable clinical outcomes. However, limited data exist on the genetic profile of these variants of PTC. We performed targeted next-generation sequencing in 36 tissue samples from patients with aggressive variants of PTC. Aggressive variants of PTC had a higher prevalence of the BRAF mutation and a lower prevalence of RAS mutation than other types of thyroid cancer. The prevalence of mutations in the TERT promoter, TP53, and genes encoding histone methyl transferases (HMTs), switch/sucrose non-fermenting (SWI/SNF) chromatin remodeling complex, and the phosphoinositide 3-kinase/protein kinase B (PKB/AKT)/mammalian target of the rapamycin (PI3K/AKT/mTOR) pathway was between the range of PTCs and poorly differentiated/anaplastic carcinoma from The Cancer Genome Atlas (TCGA) and the Memorial Sloan Kettering Cancer Center (MSKCC) data.Aggressive variants of papillary thyroid carcinoma (PTC) have been described with increasing frequency and are associated with unfavorable clinical outcomes. However, limited data exist on the comprehensive genetic profile of these variants. We performed targeted next-generation sequencing in 36 patients with aggressive variants of PTC and compared it to PTC from The Cancer Genome Atlas (TCGA) project and poorly differentiated thyroid cancers (PDTCs)/anaplastic thyroid cancers (ATCs) from the Memorial Sloan Kettering Cancer Center (MSKCC). BRAF mutation was the most prevalent (89%) in aggressive variants of PTC compared to that in other thyroid cancers. RAS mutation was identified in one patient (3%), which was less frequent than in others. TERT promoter mutation (17%) ranged between that of PTCs (9%) and PDTCs (40%). Tumor suppressor genes, ZFHX3, TP53, and CHEK2, were mutated in 14%, 3%, and 6% of aggressive variants of PTC, respectively. The mutation rate of TP53 (3%) was significantly higher than that of PTCs (0.7%) and lower than that of ATCs (73%). Mutations in three functional groups, histone methyl transferases, SWI/SNF chromatin remodeling complex, and the PI3K/AKT/mTOR pathway, were present in 11%, 14%, and 11% of samples, respectively. In conclusion, aggressive variants of PTC had higher BRAF and lower NRAS mutation prevalence than other thyroid cancers. The prevalence of mutations in the TERT promoter, TP53, and genes encoding three functional groups ranged between that of PTCs and PDTCs/ATCs.

Genetic differences in follicular thyroid carcinoma between Asian and Western countries: a systematic review.

Thyroid cancer is the most common endocrine malignancy, and follicular thyroid carcinoma (FTC) is the second most common thyroid cancer following papillary thyroid carcinoma (PTC). RAS mutation and PAX8/PPARγ rearrangement are the two representative genetic alterations in FTC, and there are studies from various countries on their regional frequencies. In this study, we systematically reviewed all available literature aiming to create a complete global map showing the frequencies of these common oncogenic drivers in FTC and to highlight the trends in Asian and Western countries. We performed a search in two electronic databases and identified 71 studies that fit our criteria from 1,329 studies found with our database search terms. There were 54 articles with 1,143 FTC patients and 39 articles with 764 FTC patients available for calculating the frequency of RAS mutation and PAX8/PPARγ rearrangement, respectively. NRAS mutation was the most frequent RAS mutation in all regions, followed by HRAS and KRAS mutation. The frequency of RAS mutation in Asian countries was higher than Western countries (34% vs. 27%, P=0.006) when the mutation detection method was not taken into account. In contrast, this difference in RAS mutation incidence between Asian and Western countries (28% vs. 25%, P=0.47) did not show up in our subgroup analysis incorporating only studies using direct sequencing method. The reported difference of RAS mutation frequency in the previous literature might not be due to the true prevalence of RAS mutation. They could be attributed to the difference in the detection method. As to PAX8/PPARγ rearrangement, Western countries overall had a much higher prevalence than Asian countries (23% vs. 4%, P<0.001), but some European countries had a low incidence, implying regional heterogeneity of PAX8/PPARγ rearrangement. A substantial lack of mutation data in FTC was found in several regions of the world such as Central Asia, Middle East, Africa, and Central and South America. Our results provide the most comprehensive global status of representative genetic alterations in FTC and highlight the similarities and differences between Asian and Western countries.

Targeting the Mutational Landscape of Bystander Cells: Drug-Promoted Blood Cancer From High-Prevalence Pre-neoplasias in Patients on BRAF Inhibitors.

Drug-promoted cancers are increasingly recognized as a serious clinical problem in patients receiving BRAF inhibitory treatment. Here we report on a patient with BRAF mutant hairy cell leukemia and monoclonal B-cell lymphocytosis (MBL), who responded durably to BRAF/MEK inhibitors (BRAFi/MEKi) but experienced transformation of a RAS mutant MBL to chronic lymphocytic leukemia (CLL) with accelerated nodal progression. Hypothesizing that BRAFi triggered excessive MEK-ERK signaling in the MBL/CLL clone via the CRAF/RAS complex as previously described for BRAFi-induced cancers, BRAFi was discontinued inducing a rapid remission of the CLL on MEKi alone. Liquid biopsy monitoring showed a continuous increase of the MBL/CLL clone from the start of BRAFi/MEKi treatment followed by a rapid decline upon BRAFi withdrawal. Next-generation sequencing of a cohort of patients with MBL and monoclonal gammopathy of unclear significance (MGUS) revealed that almost one third of these cases harbored RAS mutations. In view of the population frequency of lymphatic pre-malignant conditions and the prevalence of RAS mutations in such cases, vigilant surveillance remains critical in patients treated with BRAF inhibitors.

P-323 RAS mutations in metastatic colorectal cancer in central Tunisia

Colorectal cancer (CRC) is the most common cancer worldwide. RAS oncogene is one of the most important targets for drug development in metastatic colorectal cancers. Its mutation predicts cetuximab resistance and a worse prognosis. To our knowledge, the prevalence of RAS mutations in central Tunisia has not been previously reported.

Biology, pathology, and therapeutic targeting of RAS.

RAS was identified as a human oncogene in the early 1980s and subsequently found to be mutated in nearly 30% of all human cancers. More importantly, RAS plays a central role in driving tumor development and maintenance. Despite decades of effort, there remain no FDA approved drugs that directly inhibit RAS. The prevalence of RAS mutations in cancer and the lack of effective anti-RAS therapies stem from RAS' core role in growth factor signaling, unique structural features, and biochemistry. However, recent advances have brought promising new drugs to clinical trials and shone a ray of hope in the field. Here, we will exposit the details of RAS biology that illustrate its key role in cell signaling and shed light on the difficulties in therapeutically targeting RAS. Furthermore, past and current efforts to develop RAS inhibitors will be discussed in depth.

Prevalence of RAS and BRAF mutations in metastatic colorectal cancer patients by tumor sidedness: A systematic review and meta-analysis.

Studies have shown that the prevalence of RAS and BRAF mutations may differ by tumor sidedness among metastatic colorectal cancer (mCRC) patients. Both mutation status and tumor sidedness may impact survival and disease progression and RAS mutation status has been shown to predict response to anti‐epidermal growth factor receptor (EGFR) therapy. A systematic literature review and meta‐analysis were conducted to estimate the pooled prevalence of RAS and BRAF mutations by tumor sidedness in studies of mCRC patients. Forty‐four studies comprising 15 981 mCRC patients tested for RAS and/or BRAF mutations were included in the meta‐analyses. The prevalence of RAS mutations differed significantly by tumor side (32.4% among left‐sided tumors, 41.3% among right‐sided tumors; P = .017), as did the prevalence of KRAS mutations (35.8% among left‐sided tumors, 46.3% among right‐sided tumors; P < .0001) and BRAF mutations (4.3% among left‐sided tumors, 16.3% among right‐sided tumors; P < .0001). Among right‐sided tumors, the prevalence of RAS and KRAS mutations varied significantly by study design, with higher prevalence among observational studies than clinical trials, and there was significant variation by study location for the prevalence of KRAS mutations in left‐sided tumors and the prevalence of BRAF mutations in right‐sided tumors. These results help to better characterize the mCRC population to better inform clinicians and researchers. Few of the included studies reported overall or progression‐free survival (PFS) by both tumor sidedness and mutation status. As both of these factors may have prognostic impact, future studies should consider evaluating survival by these variables.

Abstract 3909: HDAC inhibition in combination with MEK or BCL-2 inhibition as novel therapeutic strategies in multiple myeloma

Abstract Multiple myeloma (MM) is an incurable malignancy of plasma cells. It is the second most common hematologic cancer, affecting nearly 30,000 people in the United States annually. Substantial progress has been made in the past fifteen years in the treatment of MM due to the approval of several new classes of drugs. However, patients inevitably relapse and become refractory to existing therapies. Hence, there is an immediate unmet need to develop novel therapies for MM based on a better understanding of the disease biology. Mutations in RAS have been found to occur in about 40% of newly diagnosed MM patients, with the frequency increasing to around 70% in relapsed/refractory patients. Such mutations are absent in patients with the premalignant conditions monoclonal gammopathy of undetermined significance (MGUS) and smoldering multiple myeloma (SMM). Clearly, RAS mutations contribute to both disease progression and relapse. However, targeting the MEK/ERK pathway has been unsuccessful in MM patients to date. Given the high frequency of RAS mutations in MM, we hypothesized that targeting this pathway could still be a promising strategy when combined with existing agents that have multifaceted mechanisms to promote tumor cell death, such as the recently approved histone deacetylase (HDAC) inhibitor LBH589 (panobinostat). Our results clearly demonstrate that low doses of LBH589 in combination with the MEK inhibitor AZD6244 induce BIM-dependent synergistic cell death in several MM cell lines and patient cells. Our studies also suggest that mutations in RAS/RAF could serve as a predictive biomarker for sensitivity to AZD6244/LBH589. RAS/RAF mutations appear to confer Mcl-1 dependence in MM cells, in part by driving up the phosphorylation of Mcl-1. The AZD6244/LBH589 combination is able to decrease the phosphorylation of Mcl-1 at several sites, which dissociates BIM-Mcl-1 complexes, ultimately leading to activation of the intrinsic apoptosis pathway. Additionally, we identified that wild-type RAS/RAF cells have relatively lower levels of phospho-Mcl-1, as well as higher levels of Bcl-2 and phospho-Bcl-2 when compared to mutated RAS/RAF cells. This seems to confer functional Bcl-2 dependence. Consequently, we found that wild-type RAS/RAF cells are sensitive to the BH3-mimetic ABT199 (venetoclax) when combined with LBH589. Through ongoing experiments, we hope to further confirm the mechanism of action of both these combinations, identify the particular HDAC that is required to be inhibited for the observed synergy, and validate RAS/RAF mutational status as a biomarker for predicting sensitivity to either combination. Our findings have broad therapeutic potential given the prevalence of RAS mutations in MM. Moreover, the ABT199/LBH589 combination could emerge as a targeted therapy for wild-type RAS patients, perhaps broadening the scope and capacity of Bcl-2 inhibition in MM. Citation Format: Kevin C. Miller, Jessica Haug, Teresa Kimlinger, Sanjay Kumar, Wilson Gonsalves, S. Vincent Rajkumar, Shaji K. Kumar, Vijay Ramakrishnan. HDAC inhibition in combination with MEK or BCL-2 inhibition as novel therapeutic strategies in multiple myeloma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 3909.

Prevalence of RAS and BRAF mutations in metastatic colorectal cancer (mCRC) patients by tumor location.

681 Background: There is evidence to suggest that tumor biology and pathology differ for right- and left-sided colon tumors which may affect treatment efficacy and clinical outcomes. Therefore, we conducted a systematic review and meta-analysis of the available scientific literature to summarize the prevalence of RAS and BRAF mutations by primary tumor location and to identify potential sources of heterogeneity in mutation prevalence. Methods: This study was conducted in accordance with PRISMA guidelines. Using comprehensive search strings, several medical research databases were queried and relevant variables abstracted including tumor mutation prevalence, study country, metastasis site, tissue source, study design, study dates, median age of the cohort, mutation assessment method, and length of follow-up. Results: Final abstraction was performed on 40 articles, 36 observational studies and 4 randomized trials. Most studies were from Europe (n = 16), followed by Asia (n = 11), USA, (n = 7), Australia (n = 3), and the remaining 3 were conducted in multiple countries. The proportion of males in each study ranged from 37% to 72%, and the mean age ranged from 55 to 76 years. The prevalence of all RAS mutations was significantly higher among right-sided colon tumors than left-sided colon tumors (44%, 95% Confidence Interval [CI]: 38 – 50% vs 34%, 95% CI: 29 – 38%; p = 0.009). BRAF mutation prevalence was also higher in right-sided tumors (16.1%, 95% CI: 13.1 – 19.6% vs 4.4%, 95% CI: 3.4 – 5.8%; p &lt; 0.0001). Conclusions: This systematic review and meta-analysis found that mutation prevalence varied by primary tumor right- or left-sided location among mCRC patients. Some of this variation may be explained by study characteristics such as mutation assessment method, country and length of follow-up. Further research will help to better understand treatment and outcome implications of tumor sidedness and mutation prevalence.

Comprehensive Analysis of the Relationship Between RAS and RAF Mutations and MSI Status of Colorectal Cancer in Northeastern China

Background/Aims: Colorectal cancer (CRC) is mainly caused by chromosomal instability (CIN) and microsatellite instability (MSI). The RAS and RAF genes are essential components of the CIN pathway, and several studies have found that RAS and RAF mutations are associated with MSI status in CRC. Here, we examined these three factors in CRC in Northeast China and aimed to reveal new details of the relationship between these mutations and MSI status. Methods: This study involved 290 patients with CRC who had RAS or RAF gene mutation detected using fluorescence-based allele-specific polymerase chain reaction or Sanger sequencing. The majority of the identified patients were found to harbor MSI (MSI status). Accurate molecular detection was carried out using formalin-fixed paraffin-embedded tissue or blood samples. Results: The rates of RAS and RAF mutations were 58.5% and 4.1%, respectively. The prevalence of RAS mutation in CRC was clearly higher and that of RAF mutation was lower in Northeast China compared with previously reported cohorts in other locations. High MSI level (MSI-H status) was more complex, at around 10%. This was consistent with previous data from China. However, compared with data reported from other continents, MSI-H was higher than that of Japan or South Korea in Asia, and lower than that of Europe or the United States. Conclusion: RAS/RAF mutations and MSI status in CRC are closely associated with tumor location and ethnicity. Further studies investigating the relationship between these three factors can help in the development of treatment strategies for patients with CRC.

Changes in the clinicopathological characteristics and genetic alterations of follicular thyroid cancer.

Changes in the clinicopathological characteristics and genetic alterations of follicular thyroid cancer (FTC) over time have not been reported. Moreover, the prognostic effects of RAS and TERT promoter mutations in FTC have not been clearly elucidated. We investigated changes in the clinicopathological characteristics of patients with FTC over four decades, as well as the clinical significance of genetic mutations of FTC. This retrospective study included 690 patients with FTC who underwent thyroidectomy between 1973 and 2015 at the Seoul National University Hospital. In 134 samples, genetic tests for N/H/KRAS and TERT promoter mutations and PAX8/PPARγ rearrangement were performed. The age at diagnosis has increased (P < 0.001) in recent decades and extrathyroidal extension of the tumor has become less common (P = 0.033). Other clinicopathological characteristics and prognosis of FTC have not significantly changed. The prevalence of RAS mutations decreased (P = 0.042) over time, whereas that of TERT promoter mutations remained stable. RAS mutations were associated with distant metastasis and persistent disease, and TERT promoter mutations were associated with distant metastasis, advanced TNM stage, recurrence and disease-specific mortality. FTC patients with coexistent RAS and TERT promoter mutations showed a higher recurrence risk than those with only one mutation. The age at diagnosis of FTC and the frequency of extrathyroidal extension have changed over four decades. Moreover, the prevalence of RAS mutations decreased. RAS and TERT promoter mutations may be associated with poor clinical outcomes in FTC, especially when the two mutations coexist.

RAS mutation prevalence among patients with metastatic colorectal cancer: a meta-analysis of real-world data.

Aim:A confirmed wild-type RAS tumor status is commonly required for prescribing anti-EGFR treatment for metastatic colorectal cancer. This noninterventional, observational research project estimated RAS mutation prevalence from real-world sources.Materials & methods:Aggregate RAS mutation data were collected from 12 sources in three regions. Each source was analyzed separately; pooled prevalence estimates were then derived from meta-analyses.Results:The pooled RAS mutation prevalence from 4431 tumor samples tested for RAS mutation status was estimated to be 43.6% (95% CI: 38.8–48.5%); ranging from 33.7% (95% CI: 28.4–39.3%) to 54.1% (95% CI: 51.7–56.5%) between sources.Conclusion:The RAS mutation prevalence estimates varied among sources. The reasons for this are not clear and highlight the need for further research.

Clinicopathological Associations of K-RAS and N-RAS Mutations in Indonesian Colorectal Cancer Cohort.

K-RAS and recently N-RAS gene mutation testing are mandatory requirements prior to anti-epidermal growth factor receptor (EGFR) monoclonal antibody treatment of metastatic CRC. Mutation prevalence and distribution in Indonesian colorectal cancer (CRC) are not known. Combined methods of PCR high-resolution melt (HRM), restriction fragment length polymorphism (RFLP), and direct DNA sequencing were used to genotype exons 2, 3, and 4 of both K-RAS and N-RAS genes for routine clinical testing of CRC patients. Descriptive analytical review of 595 consecutive CRC patients (years 2013 to 2016) was performed to find associations between gene mutations and clinicopathologic features. This retrospective study revealed overall K-RAS gene mutation in exon 2 (codon 12 and 13) rates being 34.9%. Women (42.5%), stages I and II (43.4%), and well and moderate differentiations (37.7%) had higher frequency of K-RAS exon 2 mutations than men (29%, p=0.006), stages (III and IV 31.9%, p=0.05), and poor differentiation (11.8%, p=0.002), respectively. At later period (2015-2016), 121 of 595 patients were genotyped for the remaining exons 3 and 4 of K-RAS as well as exons 2, 3, and 4 of N-RAS mutations resulting in overall RAS mutation prevalence of 41%. Mucinous histology had highest frequency of N-RAS mutation. Combination of PCR HRM with either RFLP or direct DNA sequencing was useful to detect K-RAS exon 2 and extended RAS mutations, respectively. Frequency of all RAS mutations in stage IV Indonesian (41%) was similar among Asians (41-49%), which tend to be lower than western (55%) CRC.

RAS-expanded Mutations and HER2 Expression in Metastatic Colorectal Cancer: A New Step of Precision Medicine.

Cetuximab and panitumumab monoclonal antibodies are a milestone in the history of treatment of metastatic colorectal cancer (mCRC) and point toward future directions for personalized treatment. Recent studies have shown that broader RAS testing is needed to select patients for targeted therapy. The objectives of our study were to identify the prevalence of RAS mutations and evaluate human epidermal growth factor receptor 2 (HER2) expression in KRAS exon 2 wild-type (WT) mCRC patients, correlating the findings with objective response rate, progression-free survival, and overall survival. In total, 29 mCRC patients undergoing treatment with cetuximab therapy were enrolled in this study. By pyrosequencing, mutations were found in 17% of nonresponder patients, in KRAS codon 146 and NRAS codon 12. HER2 positivity was limited to only 1 responder carcinoma specimen. There was no correlation between RAS mutation, HER2/neu expression, and clinicopathologic findings. We highlighted significantly the differences between objective response rate and RAS gene status. The overall survival and progression-free survival of RAS WT patients were higher compared with those with RAS-mutated disease. Clinical response to cetuximab therapy is impaired in the presence of RAS-expanded mutations. In fact, our finding of 5 mutations in RAS-expanded genes allowed us to understand the resistance to cetuximab in 33% of KRAS WT exon 2 nonresponder patients. HER2 does not seem to be a potential biomarker for cetuximab-targeted therapy. These analyses suggest that the assessment of other biomarkers is needed to determine the best treatment for patients with mCRC, to maximize benefit and minimize harm.