Prevalence Of KRAS Mutations Research Articles

Frequent GNAS mutations in low-grade appendiceal mucinous neoplasms

Background:The molecular basis for the development of appendiceal mucinous tumours, which can be a cause of pseudomyxoma peritonei, remains largely unknown.Methods:Thirty-five appendiceal mucinous neoplasms were analysed for GNAS and KRAS mutations. A functional analysis of mutant GNAS was performed using a colorectal cancer cell line.Results:A mutational analysis identified activating GNAS mutations in 16 of 32 low-grade appendiceal mucinous neoplasms (LAMNs) but in none of three mucinous adenocarcinomas (MACs). KRAS mutations were found in 30 LAMNs and in all MACs. We additionally analysed a total of 186 extra-appendiceal mucinous tumours and found that GNAS mutations were highly prevalent in intraductal papillary mucinous tumours of the pancreas (88%) but were rare or absent in mucinous tumours of the colorectum, ovary, lung and breast (0–9%). The prevalence of KRAS mutations was quite variable among the tumours. The introduction of the mutant GNAS into a colorectal cancer cell line markedly induced MUC2 and MUC5AC expression, but did not promote cell growth either in vitro or in vivo.Conclusion:Activating GNAS mutations are a frequent and characteristic genetic abnormality of LAMN. Mutant GNAS might play a direct role in the prominent mucin production that is a hallmark of LAMN.

KRAS and GNAS mutations and p53 overexpression in biliary intraepithelial neoplasia and intrahepatic cholangiocarcinomas

Similar to the pancreatic intraepithelial neoplasia (PanIN)-pancreatic carcinoma sequence model, intrahepatic cholangiocarcinoma (ICC) also reportedly follows a stepwise carcinogenesis process through the a precursor lesion: biliary intraepithelial neoplasia (BilIN). For this study, the authors investigated the status of v-Ki-ras2 Kirsten rat sarcoma viral oncogene homolog (KRAS) and GNAS complex locus (GNAS) mutations and tumor protein 53 (p53) overexpression in the stepwise process of cholangiocarcinogenesis. Thirty patients with hepatolithiasis were surveyed, and their lesions were categorized as follows: non-neoplastic large bile duct (LBD) (n = 12), peribiliary gland (PBG) (n = 9), BilIN-1 (low-grade dysplasia; n = 12), BilIN-2 (high-grade dysplasia; n = 16), and BilIN-3 (noninvasive or in situ carcinoma; n = 10). KRAS mutation at codons 12 and 13 and GNAS mutations at codon 201 were analyzed using genomic DNA extracted from isolated lesions by laser capture microdissection [corrected]. Immunohistochemical expression of p53 also was evaluated in BilIN lesions, ICCs, and extrahepatic cholangiocarcinomas (ExCCs). A prevalence of KRAS mutations was identified in patients with ICC (31.5%), BilIN-3 (30%), and BilIN-2 (43.8%) compared with BilIN-1 (25%). Furthermore, KRAS mutations were detected in LBD lesions (41.7%) and PBG lesions (44.4%), and these mutations were observed with greater frequency in patients who had BilIN with KRAS mutations. GNAS mutations were not identified in any of the ICCs or other lesions examined. The overexpression of p53 was not identified in BilIN lesions and was less frequent in ICCs (18.2%) compared with ExCCs (38.1%) and gallbladder carcinomas (61.5%). KRAS mutations, which were present in approximately 33% of BilIN lesions, occurred as an early molecular event during the progression of BilIN to ICC, whereas p53 overexpression was identified as a late molecular event. Furthermore, the current results indicted that BilIN also may arise from LBD and PBG lesions in patients who have hepatolithiasis with KRAS mutations.

Mutated KRAS Is an Independent Negative Prognostic Factor for Survival in NSCLC Stage III Disease Treated with High-Dose Radiotherapy.

Background. The main attention regarding prognostic and predictive markers in NSCLC directs towards the EGFR-targeted pathway, where the most studied genetic alterations include EGFR mutations, EGFR copy number, and KRAS mutations. We wanted to explore the prognostic impact of mutated KRAS in the stage III setting treated with high-dose radiochemotherapy. Methods. Samples were obtained from patients participating in two prospective studies of locally advanced NSCLC receiving combined radiochemotherapy: the RAKET study, a randomized phase II study where patients were treated with induction chemotherapy (carboplatin/paclitaxel) followed by concurrent radiochemotherapy, and the Satellite trial, a phase II study with induction chemotherapy (cisplatin/docetaxel) followed by radiotherapy concurrent cetuximab. The samples were analysed regarding KRAS mutations, EGFR mutations, and EGFR FISH positivity. Results. Patients with mutated KRAS had a significantly inferior survival, which maintained its significance in a multivariate analysis when other possible prognostic factors were taken into account. The prevalence of KRAS mutations, EGFR mutations, and EGFR FISH positivity were 28.8%, 7.5%, and 19.7%, respectively. Conclusion. Mutated KRAS is an independent negative prognostic factor for survival in NSCLC stage III disease treated with combined radiochemotherapy. The prevalence of KRAS mutations and EGFR mutations are as expected in this Scandinavian population.

Retrospective analysis of KRAS status in metastatic colorectal cancer patients: a single-center feasibility study

Background:The occurrence of KRAS mutations and their association with prognosis in metastatic colorectal cancer patients is not well documented in population-based studies.Objectives:To examine the feasibility of identifying archived colorectal cancer specimens, and through linkage with nationwide Danish population-based databases to investigate the prevalence of KRAS mutations and their association with colorectal cancer survival.Methods:We used the Danish Pathology Database to identify the physical location of primary (or in some cases secondary) tumor specimens from selected metastatic colorectal cancer patients referred to our hospital for palliative chemotherapy between November 1, 2008 and September 30, 2009. Routinely stored paraffin tissue blocks were obtained from the pathology archives of the originating hospital. KRAS mutation tumor status was assessed for each patient using the commercialized TheraScreen KRAS Mutation Kit. Using the unique identifier number, we linked the patients to the Danish National Registry of Patients and the Danish Civil Registration System to obtain data on date of first colorectal cancer diagnosis and follow-up status. We estimated prevalence of KRAS mutations and the 1-, 2-, and 5-year survival after colorectal cancer diagnosis using the Kaplan–Meier technique.Results:We identified 106 metastatic colorectal cancer patients (64% males). All were successfully linked to the registries, and archived tumor-tissue samples were obtained and analyzed in each case. The overall prevalence of KRAS mutations was 55%, and 1-, 2-, and 5-year overall survival after colorectal cancer diagnosis was 91%, 68%, and 25%, respectively.Conclusion:It is feasible to use Danish population-based registries to obtain archived tissue samples from metastatic colorectal cancer patients, and to estimate prevalence of KRAS mutation and subsequently evaluate the association with colorectal cancer survival.

Characteristics of Advanced- and Non Advanced Sporadic Polypoid Colorectal Adenomas: Correlation to KRAS Mutations

The malignant potential of colorectal adenomas highly correlates with their pathological characteristics, such as size, histology and grade of dysplasia. Currently, based on these parameters, adenomas are characterized as "non-advanced or advanced" and patient surveillance is adjusted accordingly. The aim of this study was to investigate the correlation between the KRAS mutations and characteristics of non-advanced and advanced colorectal adenomas for predicting the risk of increased malignant potential of adenomas that may influence the decision to offer follow-up endoscopic surveillance. We used a mutagenic polymerase chain reaction - restriction fragment length polymorphism method to determine KRAS mutations in 164 colorectal sporadic polypoid adenomas (51 non-advanced-, 113 advanced adenomas) and in 40 early colorectal carcinomas. The method of mutation detection was validated according to recommendation for KRAS mutation testing in colorectal carcinoma of the European Quality Assurance Program. The limit of detection of the assay was 3 % mutated DNA with a good reproducibility. Evaluation of pathological characteristics was performed according to European Guidelines for Quality Assurance in Colorectal Cancer Screening and Diagnosis. The morphological parameters of the adenoma such as size, histology, grade of dysplasia are highly correlated with one another: an increasing adenoma size raised the proportion of villous histology and degree of dysplasia (all p < 0.0001). KRAS mutations were detected in 31 % of the non-advanced adenomas, in 57.5 % of the advanced adenomas and in 62.5 % of the early carcinomas. Most mutations occurred at codon 12 rather than at codon 13 (72 %, 82 %, 76 % versus 22 %, 17 %, 24 %, respectively). There was no significant difference in association of KRAS mutation with age, gender, location among non-advanced-, and advanced adenomas and early carcinomas. KRAS mutation was found more often in tubulovillous and villous adenomas, whereas wild-type KRAS was observed more frequently in tubular adenomas (P < 0.0001) and there was an increased prevalence of KRAS mutations in larger adenomas (P < 0.0001). In this study KRAS mutation occurred with the same frequency in adenomas with low-grade (48 %) and high-grade (50 %) dysplasia. KRAS mutation is very strongly associated with a villous architecture and through villous component expansion, KRAS mutations may increase risk of tumor progression in sporadic colorectal polypoid adenomas.

Predictive biomarkers of response to cetuximab in Chinese patients with metastatic colorectal cancer.

e14085 Background: The paraffin-embedded tumor samples of 183 Chinese patients (pts) with metastatic colorectal cancer (mCRC) who had chemotherapy (chemo) over a period of 3.3 years were analyzed for the presence of KRAS, PIK3CA and BRAF mutations. Methods: PCR-direct sequencing was performed in micro-dissected tumor cells. The relationship between mutation and survival was evaluated using the Proportional Hazard Model, and relationship with drug response was evaluated with logistic regression. Multivariate analysis was used to adjust for the influence of age, sex, prior lines of chemo, use of bevacizumab (Bev) and the number of sites of metastases. Results: The median age was 58 yrs, over 50% of pts had &gt; 2 lines of chemo and 50% had cetuximab. The prevalence of KRAS, BRAF and PI3KCA mutations were: 55%, 5%, 20% respectively. The prevalence of each KRAS mutation subtype was: G12D (36%), G12V (22%) and G13D (16%). KRAS mutation and prior use of Bev were independent prognostic factors with respective hazard ratios (HR) of 1.5 (95% CI = 1.05-2.16, p = 0.03) and 0.51 (95% CI = 0.3-0.87, p = 0.01). In the subgroup of patients who received cetuximab and chemotherapy in the first-line setting, KRAS mutation was associated with a lack of response to chemo with odd ratio (OR) of 0.1 (95% CI = 0.01-0.79, p=0.03). There was no correlation between the presence of BRAF and/ or PIK3CA mutations and OS or drug response. Five pts had co-expression of KRAS and PIK3CA mutations and none of them responded to cetuximab. Of the 14 pts who had complete response or prolonged disease stabilization to cetuximab-chemo, 4 had KRAS mutation, 1 had PIK3CA mutation and none had BRAF mutations. Conclusions: The prevalence of KRAS, BRAF and PIK3CA mutations reported in this Chinese cohort is consistent with reports from other non-Asian populations. KRAS mutation is an independent negative prognostic factor in chemo-treated pts with mCRC. KRAS mutation was associated with poorer response to 1st line cetuximab-based chemo.

KRAS mutation profile in Puerto Rican patients with colorectal cancer: Trends in expression of KRAS mutation in PR between April 2009 to January 2011.

e14102 Background: The prevalence of a mutated KRAS gene in colorectal cancer shows ethnic variation with a range between 30-48% among the Caucasian, Asian, and European populations. The purpose of this study was to define the profile of KRAS mutation in colorectal cancer patients in Puerto Rico. Methods: Between April 2009 and January 2011, we collected the expression of KRAS gene among colorectal cancer patients in Puerto Rico. The assay was performed by a central laboratory which began to offer the test in the island in April 2009 (GENOPTIX). Data for the presence and type of mutation, age, sex, cancer histology, site of tumor, and cancer stage were collected. The site of the mutation in codon 12 and 13 were identified by the real time polymerase chain reaction (PCR). The study was approved by IRB. A total of 501 cases were available for study. SPPS version 14 was used for statistical analysis. Results: A mutated KRAS was present in 39% (n=195) of the cohort, more commonly in the 51-69 age group (55.4%; p=0.004), in females (55.4%; p=0.095), in the proximal area of colon (43.8%; p=0.017), in the codon 12 (91.2%), and 8.8% in the codon 13. The majority of mutated genes were in codon 12 (91.2%) with 12 ASP (39.5%) and 12 VAL (25.4%) most often seen. Mucinous histology was associated with 46.7% mutated KRAS, poorly differentiated tumor with 40.0% of KRAS mutation. The mutant expression of KRAS in cases with rectum carcinoma was 39.5%, where the most common mutation was 12 VAL. Conclusions: The prevalence of KRAS mutation in Puerto Rican population is similar to other reported studies. A higher prevalence of 12 VAL mutation in rectal carcinoma and a trend towards a higher mutation rate in females are different in this cohort as compared with published studies. This study was supported by NIH grant numbers G12RR-03035 &amp; 8U54MD007587-03.

KRAS aKtive, an Italian network for assessment of KRAS mutations in colorectal cancer patients: Results on 7,432 cases.

e14042 Background: The KRAS aKtive program was started on March 2009, promoted by the Italian Association of Medical Oncology (AIOM) and the Italian Society of Surgical Pathology and Cytopathologyy (SIAPEC) to support the activity of oncologists and pathologists involved in the management of metastatic colorectal cancer patients who need the assessment of the mutational status of the KRAS gene. Methods: The program was specifically devised to facilitate the exchange of biologic material, clinicopathological data and diagnostic reports within a network of oncologist, pathologists and pathology/molecular biology reference laboratories throughout Italy, connected through the site www.kras-aKtive.it. KRAS mutation analysis was performed by Sanger sequencing (SS), real time PCR or other techniques, including pyrosequencing and hybridization strip assays. Data were collected in a common database. Results: The KRAS aKtive program has involved 478 oncologists, 144 pathologists, and 24 reference laboratories. A total of 7,432 KRAS mutations analyses were performed. The tests were informative in 7,265 cases (98%). The vast majority of tests (5,626 cases, 77.4%) were conducted by SS. In 529 (7.3%) cases a real-time PCR assay was used, other detection techniques were used in 1,110 (15.3%) cases. KRAS mutations at codons 12-13 were detected in 2,874 cases (39,6%). The frequency of mutations detected by real-time PCR or other techniques (45%, and 43%, respectively) was significantly higher (p=0.002, and p=0.008%, respectively) than that observed by SS (38%). The percentage of cases evaluated by non-SS-based methods has increased during the first three years of the program. Conclusions: The results of this large survey allow an accurate estimation of the actual prevalence of KRAS mutations and their types in caucasian colorectal cancer patients. Our data indicate that the frequency of mutations detected by non-SS-based methods is higher than that obtained by SS.

Epidermal growth factor receptor and KRAS mutations in Brazilian lung cancer patients

OBJECTIVE:Epidermal growth factor receptor is involved in the pathogenesis of non-small cell lung cancer and has recently emerged as an important target for molecular therapeutics. The KRAS oncogene also plays an important role in the development of lung cancer. The aim of this study was to evaluate the frequency of epidermal growth factor receptor and KRAS mutations in a population of Brazilian patients with non-small cell lung cancer.METHODS:A total of 207 specimens from Brazilian patients with non-small cell lung cancer were analyzed for activating epidermal growth factor receptor and KRAS somatic mutations, and their associations with clinicopathological characteristics (including age, gender, ethnicity, smoking habits, and histological subtype) were examined.RESULTS:We identified 63 cases (30.4%) with epidermal growth factor receptor mutations and 30 cases (14.6%) with KRAS mutations. The most frequent epidermal growth factor receptor mutation we detected was a deletion in exon 19 (60.3%, 38 patients), followed by an L858R amino acid substitution in exon 21 (27%, 17 patients). The most common types of KRAS mutations were found in codon 12. There were no significant differences in epidermal growth factor receptor or KRAS mutations by gender or primary versus metastatic lung cancer. There was a higher prevalence of KRAS mutations in the non-Asian patients. Epidermal growth factor receptor mutations were more prevalent in adenocarcinomas than in non-adenocarcinoma histological types. Being a non-smoker was significantly associated with the prevalence of epidermal growth factor receptor mutations, but the prevalence of KRAS mutations was significantly associated with smoking.CONCLUSIONS:This study is the first to examine the prevalence of epidermal growth factor receptor and KRAS mutations in a Brazilian population sample with non-small cell lung cancer.

AOS17 KRAS genotypes in Thai patients with colorectal cancer

Background The reported prevalence of KRAS mutations, the predictive factor of efficacy of cetuximab in colorectal cancer, varies in 30–40% of patients. However, the prevalence and correlations between KRAS mutation and clinical outcome in Thai patients have never been investigated. We did a retrospective study to define the KRAS genotype in Thai patients with colorectal cancer. Methods Genomic DNA from patients’ paraffin-embedded tumour tissues was analysed for KRAS mutation at codons 12 and 13 by use of direct sequencing. Their clinical characteristics and outcomes were correlated with the genotype patterns. The concordance of KRAS genotype between primary tumour and available metastatic tumour was analysed Findings One hundred and seventeen patients with colorectal cancer were enroled. Eighty-three patients (70.9%) had wild-type KRAS (WT- KRAS ) whereas 34 patients (29.1%) had mutant KRAS . Gly12Asp (GGTGAT at codon 12) was the most common mutation (41.2%). The G13D mutation was detected in 14.7% of patients. Non-hepatic metastases were associated with mutant KRAS (adjusted OR = 3.699). The overall survival at 1, 2, and 5 years in patients with mutant KRAS was 85%, 75%, and 54%, respectively, compared with 96%, 83%, and 47%, respectively, in those with wild-type KRAS ( p = 0.56). Discordance of the KRAS genotype (wild-type primary tumours and mutant metastatic tumours) was detected in 2 of 9 patients (22.2%). After cetuximab-chemotherapy, the PFS of one case with discordant KRAS was shorter than the median PFS in 10 cases with primary WT- KRAS (2.7 months versus 12.8 months). Interpretation The prevalence of KRAS mutation in Thai patients with colorectal cancer was similar to that in other reports. Patients with mutant KRAS had more non-hepatic metastases than did those with WT- KRAS . Overall survival in patients with wild-type KRAS was not different from those with mutant KRAS . The mosaic pattern of tumour cells might account for the discordance in the KRAS status. Funding Ramathibodi cancer committee research budget. The authors declared no conflicts of interest. AOS18 Calreticulin expression is required for oral cancer-cell proliferation and migration, and is correlated with clinicopathologic features in oral squamous-cell carcinoma patients C. Wang, Y. Wu, C. Chiu, H. Kuo, C. Hsu, J. Chen Withdrawn.

Pharmacoeconomic analysis for K-Ras status–based decisions for first-line therapy of metastatic colorectal cancer (mCRC) in Spain.

438 Background: Personalized medicine is a challenge for current oncology practice. Nowadays there are no pharmacoeconomic analyses in Spain dealing with the clinical and financial impact secondary to K-Ras based decisions for first-line therapy in mCRC patients. So, this study was aimed to assess the cost- effectiveness of K-Ras status based decisions in first- line therapy of mCRC patients in comparison with non- K-Ras based selection of available therapies. Methods: K-Ras mutation prevalence and efficacy of available therapies (measured as response rate and progression free survival) were extracted from randomized clinical trials (RCT) that allowed on-label use of accessible drugs in Spain. Then, we have simulated all possibilities of combination therapies for first-line mCRC based on K-Ras status (wild- type vs mutated) and confronted with all therapies that could be chosen in absence of K-Ras analysis. Prices for all drugs in Spain were used to assume the best- value for each drug including all possibilities to reduce pharmacy costs. For first line, median duration of therapy reported by RCT was used to calculate the final budget. 70 kg and 1.7 m were used as reference for patients dose calculations. Results: First-line therapy that includes a biological drug in absence of K-Ras status based decisions implies an incremental cost per 1% of increased response rate of 1,237 euros for irinotecan based doublets and 3,193 euros for oxaliplatin based doublets. On the opposite, K-Ras based decisions reduce costs per objective response by 69% and 35% for irinotecan and oxaliplatin- based schedules in K-Ras wild-type population incorporating cetuximab as biological agent. These data mimic all calculi based on incremental costs secondary to improved progression free survival measured as HR when all scenarios without prior determination of K-Ras status were confronted with K-Ras based decisions. Conclusions: K-Ras based decisions reduces costs per objective response as well as per improved progression free survival. The most cost- effective scenario among all simulated was cetuximab in combination with chemotherapy for patients that harbor wt K-Ras mCRC.

K-ras Mutation is Strongly Associated with Perineural Invasion and Represents an Independent Prognostic Factor of Intrahepatic Cholangiocarcinoma after Hepatectomy

Unsatisfying long-term survival of intrahepatic cholangiocarcinoma (ICC) triggers the clinicians searching for molecular markers, such as K-ras mutation, to tailor management strategy. Additionally, emergence of tyrosine kinase inhibitors (TKIs) brings new hope to palliate advanced ICC; whether the efficacy of TKIs is influenced by k-ras mutation is largely unknown. This study was designed to determine the prevalence of k-ras mutation and its clinical significance in ICC, as well as to pave the reference for future application of TKIs. A total of 86 patients with ICC who underwent hepatectomy were retrospectively recruited. K-ras mutation was determined by using laser capture microdissection and direct sequencing method. Association among clinicopathological variables and K-ras mutation was analyzed. Prognostic factors of ICC after hepatectomy also were determined. Nineteen (22%) patients exhibited K-ras mutations. Seventeen had their K-ras mutations occurring at codon 12, and the remaining two occurring at codon 13 and codon 61 in one each. Perineural invasion was exclusively the variable associated with K-ras mutation (odds ratio, 6.9) using logistic regression analysis. Multivariate analysis demonstrated that resection margin, T-status, nodal metastasis, and K-ras mutation were independent prognostic factors. The median survival of ICC patients with K-ras mutation was 5.7 months compared with 19.0 months in those without K-ras mutation (P = 0.002). The prevalence of K-ras mutations in a considerably large cohort of ICC was 22%. K-ras mutation is strongly associated with perineural invasion phenotypically. K-ras mutation is an independent prognostic factor of ICC after hepatectomy.

Cost‐effectiveness of KRAS testing in metastatic colorectal cancer patients in the United States and Germany

The objective of this study was to determine the cost-effectiveness of testing for KRAS mutations before administering EGFR inhibitors such as cetuximab and panitumumab for patients with advanced metastatic colorectal cancer (mCRC) in the United States and Germany. We developed a lifetime Markov model of costs and survival associated with treating mCRC patients to assess the impact of KRAS testing before administering EGFR inhibitor-containing chemotherapy regimens. Overall, combination therapies involving cetuximab plus irinotecan/FOLFIRI had a better life expectancy (25.83 weeks) than cetuximab or panitumumab alone. Use of KRAS testing (assuming KRAS mutant patients receive only irinotecan) was equally effective and saved $12,428 per patient in the United States. When KRAS mutant patients received best supportive care, the life expectancy decreased slightly (24.26 weeks vs. 25.83 weeks) and the costs decreased by $13,501 in the United States and €9,560 in Germany. For patients treated with cetuximab alone, use of KRAS testing to identify mutations lowered costs by $8,040 per patient in the U.S. analysis and €3,856 per patient in the German analysis. For patients treated with panitumumab alone, use of KRAS testing to identify mutations lowered costs by $7,546 per patient in the U.S. analysis and €4,612 per patient in the German analysis. Model results were sensitive to the cost of chemotherapy regimens and the prevalence of KRAS mutations in the population. Under most scenarios, using KRAS testing to select patients for EGFR inhibitor therapy saved $7,500-$12,400 per patient in the United States and €3,900-€9,600 per patient in Germany with equivalent clinical outcomes.

STK33 Kinase Activity Is Nonessential in KRAS-Dependent Cancer Cells

Despite the prevalence of KRAS mutations in human cancers, there remain no targeted therapies for treatment. The serine-threonine kinase STK33 has been proposed to be required for the survival of mutant KRAS-dependent cell lines, suggesting that small molecule kinase inhibitors of STK33 may be useful to treat KRAS-dependent tumors. In this study, we investigated the role of STK33 in mutant KRAS human cancer cells using RNA interference, dominant mutant overexpression, and small molecule inhibitors. As expected, KRAS downregulation decreased the survival of KRAS-dependent cells. In contrast, STK33 downregulation or dominant mutant overexpression had no effect on KRAS signaling or survival of these cells. Similarly, a synthetic lethal siRNA screen conducted in a broad panel of KRAS wild-type or mutant cells identified KRAS but not STK33 as essential for survival. We also obtained similar negative results using small molecule inhibitors of the STK33 kinase identified by high-throughput screening. Taken together, our findings refute earlier proposals that STK33 inhibition may be a useful therapeutic approach to target human KRAS mutant tumors.

Differences in the frequencies of K-ras c12-13 genotypes by gender and pathologic phenotypes in colorectal tumors measured using the allele discrimination method

The frequencies of different genotypes of the K-ras oncogene in colorectal cancer (CRC) reveal complex relationships among gender, age, and tumor aggression, however, differences among these studies could also be attributed to a lack of standardization of the detection methods used. We developed the allele discrimination assay, which uses dual-color real-time polymerase chain reaction (qPCR) as a fast K-ras genotyping method, and demonstrated higher sensitivity and specificity than DNA sequencing with formalin-fixed paraffin tissues. The assay detected K-ras mutations among 83 of 204 patients with CRC (40.7%); 20.6% of these mutations were G12D (GAT) mutations, 7.4% were G13D (GAC) and G12V (GTT), and 5.3% were other types. A higher proportion of females was observed overall in tumors with K-ras mutations (60.2%, P = 0.01), codon 12 mutations (63.2%, P = 0.005), and transversions (69.6%, P = 0.02), which reflected the higher prevalence of females among the well- to moderately differentiated tumors (29% in males vs. 53% in females; interaction P = 0.03). The opposite was observed for poorly differentiated tumors (47% in males vs. 35% in females). No significant influence of age was found on the prevalence of K-ras mutation. Males with pathological changes and females with poorly differentiated tumors displayed GAT as a less common genotype compared with most other prevalence studies. In conclusion, allele discrimination, with no additional amplification step, is a fast and reliable genotyping method for detecting K-ras c12-13 mutations. Using this method, we demonstrate differences in the frequencies of K-ras genotypes by gender and pathologic phenotypes of CRC.

Unique patterns of CpG island methylation in inflammatory bowel disease-associated colorectal cancers

CpG island (CGI) hypermethylation at discrete loci is a prevalent cancer-promoting abnormality in sporadic colorectal carcinomas (S-CRCs). We investigated genome-wide CGI methylation in inflammatory bowel disease (IBD)-associated CRCs (IBD-CRCs). Methylation microarray analyses were conducted on seven IBD-CRCs, 17 S-CRCs, and eight normal control colonic tissues from patients without CRC or IBD. CGI methylator phenotype (CIMP), a surrogate marker for widespread cancer-specific CGI hypermethylation, was examined in 30 IBD-CRCs and 43 S-CRCs. The genome-wide CGI methylation pattern of IBD-CRCs was CIMP status-dependent. Based on methylation array data profiling of all autosomal loci, CIMP(+) IBD-CRCs grouped together with S-CRCs, while CIMP(-) IBD-CRCs grouped together with control tissues. CIMP(-) IBD-CRCs demonstrated less methylation than did age-matched CIMP(-) S-CRCs at autosomal CGIs (z-score -0.17 vs. 0.09, P = 3 × 10(-3)) and CRC-associated hypermethylation target CGIs (z-score -0.43 vs. 0.68, P = 1 × 10(-4)). Age-associated hypermethylation target CGIs were significantly overrepresented in CGIs that were hypermethylated in S-CRCs (P = 1 × 10(-192)), but not in CGIs that were hypermethylated in IBD-CRCs (P = 0.11). In contrast, KRAS mutation prevalence was similar between IBD-CRCs and S-CRCs. Notably, CIMP(+) prevalence was significantly higher in older than in younger IBD-CRC cases (50.0 vs. 4.2, P = 0.02), but not in S-CRC cases (9.7 vs. 16.7, P = 0.92). Cancer-specific CGI hypermethylation and age-associated CGI hypermethylation are diminished in IBD-CRCs relative to S-CRCs, while the KRAS mutation rate is comparable between these cancers. CGI hypermethylation appears to play only a minor role in IBD-associated carcinogenesis. We speculate that aging, rather than inflammation per se, promotes CIMP(+) CRCs in IBD patients.

Frequency of Kras mutation in colorectal cancer patients in a minority-based population at a community clinical oncology program (CCOP)–participating inner-city hospital.

e14051 Background: Kras gene is a downstream target of epidermal growth factor receptor (EGFR) signaling. Activating mutations of this gene confer resistance to EGFR-targeted monoclonal antibodies and may also have prognostic significance. The prevalence of this mutation has been reported in 30-40% of colorectal cancers in United States. We present the frequency and associated clinico-social characteristics of this mutation in a cohort of predominantly minority-based population. Methods: We retrospectively reviewed records of patients with metastatic colorectal cancer at our institution over an 8-year period from 2003 to 2010. The objectives were to define the frequency of Kras mutation in this predominantly minority based population, and to study its relationship with various demographic and clinical parameters. Results: Between April 2003 and August 2010, 63 patients with metastatic colorectal cancer underwent Kras mutation testing. The majority were of African American (50%) or Hispanic (21%) descent. Caucasians represented only 27% of the study population. 42 (66.7%) tested positive for Kras mutation. A difference in mutation prevalence was seen with regards to gender (males vs. females; 32/45=71% vs. 10/18=55.5%), age (>/=50 vs. < 50; 32/47=68% vs. 10/16=62%) and primary tumor location (recto-sigmoid vs. colon; 19/25=76% vs. 23/37=62%). Conclusions: Our cohort of predominantly minority ethnicities shows a significantly higher prevalence of Kras mutations than previously reported national figures. This suggests that race is an important predictor of presence of this mutation, which in turn has therapeutic and potentially prognostic implications. Further studies are needed to better understand these racial differences and adopt a more personalized approach to colorectal cancers. Kras type Wild Mutant Total (%) Number (%) 21 (33.3%) 42 (66.7%) 63 (100%) Ethnicity African American 12 19 31 (50%) Hispanic 2 10 12 (21%) Caucasian 6 11 17 (27%) Asian 1 2 3 (1%) Gender Male 13 32 45 (71.4%) Female 8 10 18 (28.6%) Age at diagnosis <50 years 7 10 17 (27%) >/=50 years 14 32 46 (73%) Primary tumor location Colon 14 23 37 (59%) Rectosigmoid 7 19 26 (41%)

KRAS mutations in colorectal cancer patients in Italy: Results from the KRAS aKtive program.

e14000 Background: The Italian Association of Medical Oncology (AIOM) and the Italian Society of Surgical Pathology and Cytopathologyy (SIAPEC) collaborated to promote a national program (KRAS aKtive) to support the activity of oncologists and pathologists involved in the management of metastatic colorectal cancer patients who need the assessment of the mutational status of the KRAS gene. Methods: The KRAS aKtive program was specifically devised to facilitate the exchange of biologic material, clinicopathological data and diagnostic reports within a network of oncologist, pathologists and pathology/molecular biology reference laboratories throughout Italy, connected through the site www.kras-aKtive.it. KRAS mutation analysis was performed by PCR-Sanger sequencing, real time PCR or other techniques. Data were collected in a common database. Results: Started on March 2009, the KRAS aKtive program has involved 407 oncologists, 125 pathologists, and 24 reference laboratories from 16 of the 21 Italian regions. A total of 2,589 KRAS mutation texts were performed. The majority of tests (1,967 cases, 78.1%) were conducted by sequencing. In 231 (9.2%) cases a real-time PCR assay was used, other detection techniques were used in 321 (12.7%) cases. The tests were informative in 97.3% of cases. KRAS mutations at codons 12-13 were detected in 940 cases (37.3%). The frequency of mutations detected by sequencing was lower than that observed by real-time PCR (35.7 and 48.1%, respectively). The frequency of the different type of mutations at codons 12 and 13 is reported in the table. Conclusions: The results of this large survey allow an accurate estimation of the actual prevalence of KRAS mutations and their types in caucasian colorectal cancer patients. Moreover, although the subset numbers are still insufficient to draw definitive conclusions, our data suggest that the frequency of mutations detected by real-time PCR amplification is higher than that obtained by sequencing. Program supported by Merck Italy. Mutation type Number % GGT->GTT G12V 226 24.04% GGT->GAT G12D 309 32.87% GGC->GAC G13D 186 19.79% GGT->TGT G12C 78 8.30% GGT->GCT G12A 49 5.21% GGT->AGT G12S 43 4.57% GGT->CGT G12R 14 1.49% Others 35 3.72% Total 940 100.00%

East meets West: ethnic differences in epidemiology and clinical behaviors of lung cancer between East Asians and Caucasians

Lung cancer is the leading cause of cancer death worldwide, with large variation of the incidence and mortality across regions. Although the mortality of lung cancer has been decreasing, or steady in the US, it has been increasing in Asia for the past two decades. Smoking is the leading cause of lung cancer, and other risk factors such as indoor coal burning, cooking fumes, and infections may play important roles in the development of lung cancer among Asian never smoking women. The median age of diagnosis in Asian patients with lung cancer is generally younger than Caucasian patients, particularly among never-smokers. Asians and Caucasians may have different genetic susceptibilities to lung cancer, as evidenced from candidate polymorphisms and genome-wide association studies. Recent epidemiologic studies and clinical trials have shown consistently that Asian ethnicity is a favorable prognostic factor for overall survival in non-small cell lung cancer (NSCLC), independent of smoking status. Compared with Caucasian patients with NSCLC, East Asian patients have a much higher prevalence of epidermal growth factor receptor (EGFR) mutation (approximately 30% vs. 7%, predominantly among patients with adenocarcinoma and never-smokers), a lower prevalence of K-Ras mutation (less than 10% vs. 18%, predominantly among patients with adenocarcinoma and smokers), and higher proportion of patients who are responsive to EGFR tyrosine kinase inhibitors. The ethnic differences in epidemiology and clinical behaviors should be taken into account when conducting global clinical trials that include different ethnic populations.

Clinicopathological and Molecular Features of Colorectal Serrated Neoplasias With Different Mucosal Crypt Patterns

Endoscopic examination shows that serrated neoplasias (SNs), such as serrated adenomas and sessile serrated adenomas, exhibit different mucosal crypt patterns. However, it remains unclear whether advanced serrated polyps with different mucosal crypt patterns have different clinicopathological or molecular features. We classified the mucosal crypt patterns of 86 SNs into three types (hyperplastic, adenomatous, and mixed pattern) and evaluated their clinicopathological and molecular features. We found significant differences in the proliferative activity status between SNs with mixed/adenomatous patterns and those with the hyperplastic patterns. SNs with the hyperplastic pattern were frequently located in the proximal colon and had a macroscopically superficial appearance, whereas SNs with the adenomatous pattern were often located in the distal colon and had a protruding appearance. Furthermore, a significant difference was observed in the frequency of the CpG island methylator phenotype (CIMP), involving the methylation of two or more CIMP-related genes (MINT1, MINT2, MINT31, p16, and MLH1), between SNs with the hyperplastic pattern and those with the mixed/adenomatous patterns (18/32 (56%) vs. 8/28 (29%) or 7/26 (27%); P=0.0309 or P=0.0249, respectively). Moreover, the prevalence of KRAS mutations was significantly higher in SNs with the adenomatous pattern than in those with the hyperplastic pattern (7/26 (27%) vs. 1/32 (3%); P=0.0173). In comparison with other patterns, the mixed pattern was detected more frequently in mixed serrated polyps (MSPs), which contain separate histological components. Some MSPs exhibited concordant molecular alterations among the different histological components. The clinicopathological and molecular features of SNs correlated strongly with their mucosal crypt patterns, which were observed using chromoendoscopy.