K-ras activation in colorectal tumors from patients with familial polyposis coli

K-ras gene mutation in colorectal tumors from patients with familial polyposis coli were investigated using oligonucleotide probes specific for a mutation at codon 12, 13, or 61 of the K-ras gene. The authors examined 22 colorectal carcinomas and 51 colorectal adenomas from 41 familial polyposis coli patients and observed mutations at codons 12 and 13 in eight of 22 colorectal carcinomas (36%) and six of 51 colorectal adenomas (12%). Thus, the frequency and sites of K-ras gene mutation in colorectal carcinoma from familial polyposis coli patients are similar to those in cases of sporadic colorectal carcinoma and may not be the first genetic event linked to the tumorigenesis.     

K-ras gene mutations in adenomas and carcinomas of the colon.

DNA extracted from 29 colorectal carcinomas and 40 sporadic adenomas was amplified by the polymerase chain reaction (PCR) and analysed for the presence of K-ras gene mutations at codon 12 using a panel of synthetic oligonucleotide probes specific for normal and mutated sequences. The presence of mutations was correlated with various histopathological and clinical data. Ten carcinomas (34.5%) and 14 sporadic adenomas (35%) showed K-ras mutations at codon 12. In the carcinoma group, no apparent correlation was found between the presence of mutant oncogenes and the degree of histological differentiation, Dukes' staging or the development of distant metastasis. In the adenoma group, the frequency of mutations increased with the size of the adenoma and the severity of the dysplastic changes. This study confirms that ras gene mutations are common and early events in colon carcinogenesis. They appear to give a selective growth advantage to those polyps with mutations which leads to their increase in size and thus possibly prepare the ground for malignant transformation.     

Mutations of the PIK3CA gene in hereditary colorectal cancers

Somatic mutations of the PIK3CA gene have recently been detected in various human cancers, including sporadic colorectal cancer. However, mutations of the PIK3CA gene in hereditary colorectal cancers have not been clarified. To elucidate the mutation status in familial adenomatous polyposis (FAP) and hereditary nonpolyposis colorectal cancer (HNPCC), which are the most common hereditary colorectal cancers, we investigated PIK3CA mutations in 163 colorectal tumors, including adenomas, intramucosal carcinomas and invasive carcinomas. For comparison, we also analyzed mutations of the same gene in 160 sporadic colorectal tumors at various histopathological stages. Analysis at exons 1, 7, 9 and 20 of the PIK3CA gene revealed somatic mutations in 21% (8 of 39) of FAP invasive carcinomas, 21% (7 of 34) of HNPCC invasive carcinomas, 15% (8 of 52) of sporadic invasive carcinomas, and 14% (7 of 50) of sporadic colorectal metastases in the liver. Mutations in FAP and HNPCC carcinomas predominantly occurred in the kinase domain (exon 20), while the majority of mutations in sporadic cases occurred in the helical domain (exon 9). Adenomas and intramucosal carcinomas from all patients exhibited no mutations (0 of 148). Our data suggest that PIK3CA mutations contribute to the invasion step from intramucosal carcinoma to invasive carcinoma in colorectal carcinogenesis in FAP and HNPCC patients at a similar extent to that seen in sporadic patients.     

K-ras mutations and HLA-DR expression in large bowel adenomas.

A total of 72 sporadic colorectal adenomas in 56 patients were studied for the presence of point mutations in codons 12 and 13 of the K-ras gene and for HLA-DR antigen expression related to clinicopathological variables. Forty K-ras mutations in 39 adenomas were found (54%): 31 (77%) in codon 12 and nine (23%) in codon 13. There was a strong relationship between the incidence of K-ras mutations and adenoma type, degree of dysplasia and sex. The highest frequency of K-ras mutations was seen in large adenomas of the villous type with high-grade dysplasia. Fourteen out of 15 adenomas obtained from 14 women above 65 years of age carried mutations. HLA-DR positivity was found in 38% of the adenomas, large tumours and those with high-grade dysplasia having the strongest staining. Coexpression of K-ras mutations and HLA-DR was found significantly more frequently in large and highly dysplastic adenomas, although two-way analysis of variance showing size and grade of dysplasia to be the most important variable. None of the adenomas with low-grade dysplasia showed both K-ras mutation and HLA-DR positivity (P = 0.004). K-ras mutation is recognised as an early event in colorectal carcinogenesis. The mutation might give rise to peptides that may be presented on the tumour cell surface by class II molecules, and thereby induce immune responses against neoplastic cells.     

Association of K-ras mutations with liver metastases from colorectal carcinoma

BACKGROUND: Colorectal carcinomas were studied regarding early proof of liver metastases through determination of K-ras mutations. PATIENTS AND METHODS: Seventy-seven colorectal carcinomas were investigated for the presence of point mutations in codon 12 and 13 of the K-ras gene, using single-strand conformation polymorphism (SSCP) analysis and direct sequencing. RESULTS: Twenty-six carcinomas were positive for K-ras mutations, of which 21 had codon 12 and 5 had codon 13 mutations. Twenty patients with K-ras-positive tumor (20 out of 26: 77%) developed liver metastases, of which 13 had simultaneous metastases and 7 had metachronous metastases. There was a significant association between K-ras mutations and liver metastases (p=0.03). A multivariate logistic regression model demonstrated that the involvement of lymph node (p<0.01) and K-ras mutations (p=0.02) were predictive factors for liver metastases from colorectal carcinoma. Sequencing in carcinomas without liver metastases showed the base change in the first position of codon 12, whereas with liver metastases it showed significantly frequent base change in the second position of codon 12 (p<0.01). CONCLUSION: It is suggested that the presence of K-ras mutation, especially base change in the second position of codon 12, may predict liver metastases from colorectal carcinoma.     

Disseminated colorectal tumor cells in organs prone to metastasis detected by new double enriched nested-PCR in comparison with recognized assays

Hypothetically, K-ras mutations can be used as a marker of disseminated tumor cells (DTCs) in patients with K-ras mutated primary carcinoma. This study focused on the development of a useful assay for detecting low numbers of DTCs in potential target tissues of metastatic K-ras codon 12 mutated colorectal cancer. Tumor, liver, lymph node and bone marrow tissues from 46 colorectal carcinoma patients were examined for K-ras codon 12 mutations with a new double enriched nested (DEN)-PCR and the incidence of mutations was compared to those obtained from three established assays. DEN-PCR followed by sequencing found one mutated cell within 107 K-ras codon 12 wild-type cells and was more sensitive than other methods (1:106-1:102). Colon carcinomas (26/46) and adenomas (1/3) harbored mutations in K-ras codon 12. Sixteen of these 27 mutated tumors were found with all assays, two with three methods and one with the two most sensitive assays. In 8 cases, only DEN-PCR identified the K-ras mutation, and thus prevailed over the other methods used (p<0.002). Eight of 26 patients with K-ras mutated colorectal carcinoma also harbored K-ras mutated DTCs in liver and lymph nodes, respectively, and 4 in bone marrow. For liver and lymph node samples, DTC-mutations were identical to those in the primary carcinoma but those in bone marrow differed from the respective mutation in the primary carcinoma. In conclusion, DEN-PCR is a highly sensitive method for detecting K-ras mutations as marker of early and late tumor cell dissemination in tissues potentially harboring colorectal carcinoma metastases.     

Involvement of APC and K-ras mutation in non-polypoid colorectal tumorigenesis

The aim of this study was to clarify the role of APC and K-ras mutations in non-polypoid colorectal tumorigenesis. DNA from 63 adenomas (31 polypoid, 17 superficial elevated, 15 superficial depressed), 66 submucosally invasive carcinomas (47 polypoid, 19 non-polypoid) and 34 advanced carcinomas were examined for K-ras codon 12 point mutations and APC mutations in the mutation cluster region. K-ras mutation: the frequency in superficial depressed adenomas was lower than that in polypoid adenomas (0% vs 31%: P= 0.018). The frequency in non-polypoid carcinomas was lower than that in polypoid carcinomas (11% vs 56%: P = 0.0008), and was relatively low compared with that in polypoid adenomas (11% vs 31%). APC mutation: the frequency in superficial depressed adenomas was lower than that in polypoid adenomas (7% vs 43%: P = 0.016), and that in polypoid carcinomas was similar to that in non-polypoid carcinomas. Polypoid adenomas, polypoid carcinomas and advanced carcinomas had almost the same frequency. There may be some pathway other than the conventional adenoma-carcinoma sequence in development of non-polypoid carcinomas. The precursors of most non-polypoid carcinomas are considered to be de novo or superficial depressed adenomas. In this non-polypoid pathway, APC mutation seems to be requisite but K-ras mutation not. It is possible that new APC mutations are acquired after the development of superficial depressed adenomas.     

Difference in the role of loss of heterozygosity at 10p15 (KLF6 locus) in colorectal carcinogenesis between sporadic and familial adenomatous polyposis and hereditary nonpolyposis colorectal cancer patients

OBJECTIVES: To clarify the role of the KLF6 (Kruppel-like factor 6) locus in multistep colorectal carcinogenesis, we analyzed loss of heterozygosity (LOH) at 10p15 (KLF6 locus) and mutations of the KLF6gene in 298 colorectal tumors at various pathological stages of sporadic and familial adenomatous polyposis (FAP) and hereditary nonpolyposis colorectal cancer (HNPCC) patients. METHODS: 10p15 LOH was analyzed using KLF6M1 and KLF6M2, and KLF6 gene mutation was analyzed using PCR-SSCP and sequencing. RESULTS: It was found that the frequencies of LOH (sum of M1 and M2) were 4% in adenomas, 0% in intramucosal carcinomas, 35% in invasive carcinomas, and 33% in liver metastases in sporadic cases. Invasive carcinomas from FAP patients showed only 6% LOH, and invasive carcinomas from HNPCC patients exhibited 0% LOH. Mutation analysis of the KLF6 gene in 298 colorectal tumors detected no somatic mutations. CONCLUSIONS: The present data suggest that LOH of the KLF6 locus at chromosome 10p15 contributes to the invasion step from an intramucosal carcinoma to an invasive carcinoma specifically in sporadic colorectal carcinogenesis, but is rarely involved in the carcinogenesis of FAP and HNPCC cases. Moreover, the absence of somatic mutations suggests the uncertainty of the KLF6 gene as a classical tumor suppressor gene at the lost 10p15 region in colorectal carcinogenesis.     

Molecular analysis of sulindac-resistant adenomas in familial adenomatous polyposis.

PURPOSE: Sulindac causes the reduction of adenomas in familial adenomatous polyposis (FAP) patients, but complete regression is unusual, and breakthrough of colorectal carcinoma during sulindac treatment has been described. The molecular features related to sulindac resistance are unknown. Therefore, we investigated molecular alterations in adenomas from FAP patients with complete adenoma regression on sulindac (responsive patients) and from FAP patients with sulindac-resistant adenomas (resistant patients). DESIGN: Fourteen baseline adenomas (removed before sulindac treatment) from six responsive patients were studied. Also, 9 baseline adenomas and 34 resistant adenomas (removed during sulindac treatment) from three resistant patients were analyzed. Using immunohistochemistry, we evaluated the expression of beta-catenin, cyclooxygenase-2 (Cox-2), p53, Bcl-2, and Bax. K-ras codon 12 mutations, loss of heterozygosity at 5q (APC locus), and microsatellite instability were studied with PCR-based techniques. RESULTS: There were no significant differences between baseline adenomas from sulindac-responsive and -resistant patients (P > 0.05). There was less loss of membranous beta-catenin staining and less nuclear beta-catenin accumulation in resistant adenomas compared with baseline adenomas from the same (sulindac-resistant) patients (P < 0.01) or baseline adenomas from responsive patients (P < 0.01). Epithelial Cox-2 expression was less, though not significant, in resistant adenomas compared with baseline adenomas from resistant patients, but was significantly less in baseline adenomas from responsive patients (P < 0.01). K-ras mutations were found in 8 of 34 resistant adenomas (24%) and in none of the baseline adenomas (P < 0.05). Stromal Cox-2 expression, staining of p53 and Bcl-2, and loss of heterozygosity at 5q were comparable in both groups. Loss of Bax staining and microsatellite instability were not found in any adenoma. CONCLUSIONS: Sulindac-resistant adenomas display less alteration in beta-catenin staining and less epithelial Cox-2 expression when compared with adenomas removed before sulindac treatment. K-ras mutations may contribute to sulindac-resistance. Continued research is needed to investigate molecular alterations related to sulindac resistance.     

The balanced induction of K-ras codon 12 and 13 mutations in mucosa differs from their ratio in neoplastic tissues

The aim of this study was to compare the ratio of K-ras codon 12 and 13 mutations in various tissues of colorectal cancer patients. Multiple samples of inconspicuous mucosa and a sample of carcinoma tissue were taken from 36 colorectal cancer patients (group I) and these results were compared with those from polyp and carcinoma tissues of another 48 colorectal cancer patients (group II). A polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) assay was used to detect the respective point mutations. The results of this assay were complemented by sequencing the K-ras mutations. In mucosa tissue, the ratio of codon 12 and 13 mutations was nearly equal (0.9:1) whereas the respective ratio in tumour tissue showed a strong preponderance of K-ras codon 12 mutations (14:1, p=0.004). In polyp tissue of patients from group II, the ratio was 2.7:1 and that in carcinomas was 19:1 (p=0.053). The prevalence of both types of mutation was 14.6% in all mucosa samples, corresponding to 30.6% of group I patients. The K-ras mutation rate in carcinoma tissue of the same patients was 38.9%. Similarly, 33.4% of all polyp and 41.7% of all carcinoma samples from group II harboured K-ras codon 12 and/or 13 mutations. Sequencing confirmed 59 of 60 K-ras codon 12 mutations, but due to the detection limit for sequencing (1:10(4)) only 10 of 20 K-ras codon 13 mutations were confirmed. It is concluded that after balanced induction K-ras codon 12 mutations increase in frequency relative to K-ras codon 13 mutations during tumour progression.     

Similarity of the phenotypic patterns associated with BRAF and KRAS mutations in colorectal neoplasia

Activation of the RAS/RAF/extracellular signal-regulated kinase-mitogen-activated protein kinase/extracellular signal-regulated kinase/mitogen-activated protein kinase pathway by RAS mutations is commonly found in human cancers. Recently, we reported that mutation of BRAF provides an alternative route for activation of this signaling pathway and can be found in melanomas, colorectal cancers, and ovarian tumors. Here we perform an extensive characterization of BRAF mutations in a large series of colorectal tumors in various stages of neoplastic transformation. BRAF mutations were found in 11 of 215 (5.1%) colorectal adenocarcinomas, 3 of 108 (2.8%) sporadic adenomas, 1 of 63 (1.6%) adenomas from familial adenomatous polyposis (FAP) patients, and 1 of 3 (33%) hyperplastic polyps. KRAS mutations were detected in 34% of carcinomas, 31% of sporadic adenomas, 9% of FAP adenomas, and no hyperplastic polyps. Eight of 16 BRAF mutations were V599E, the previously described hotspot, and none of these was associated with a KRAS mutation in the same lesion. The remaining eight mutations involve other conserved amino acids in the kinase domain, and 62.5% have a KRAS mutation in the same tumor. Our data suggest that BRAF mutations are, to some extent, biologically similar to RAS mutations in colorectal cancer because both occur at approximately the same stage of the adenoma-carcinoma sequence, both are associated with villous morphology, and both are less common in adenomas from FAP cases. By contrast, colorectal adenocarcinomas with BRAF mutations are associated with early Dukes' tumor stages (P = 0.006) and no such relationship was observed for KRAS mutations. The presence in some colorectal neoplasms of mutations in both BRAF and KRAS suggests that modulation of the RAS-RAF-extracellular signal-regulated kinase-mitogen-activated protein kinase/extracellular signal-regulated kinase/mitogen-activated protein kinase signaling pathway may occur by mutation of multiple components.     

Specific codon 13 K-ras mutations are predictive of clinical outcome in colorectal cancer patients, whereas codon 12 K-ras mutations are associated with mucinous histotype.

BACKGROUND: K-ras mutations, one of the earliest events observed in colorectal carcinogenesis, are mostly found in codons 12 and 13, and less frequently in codon 61, all three of which are estimated to be critical for the biological activity of the protein. Nevertheless the prognostic significance of such mutations remains controversial. Our purpose was to assess whether any or specific K-ras mutations in primary colorectal cancer had prognostic significance and were linked to clinico-pathological parameters. PATIENTS AND METHODS: Paired tumor and normal tissue samples from a consecutive series of 160 untreated patients (median of follow up 71 months), undergoing resective surgery for primary colorectal carcinoma, were prospectively studied for K-ras mutations by PCR/single strand conformation polymorphism sequencing. RESULTS: Seventy-four of the 160 (46%) primary colorectal carcinomas presented mutations in K-ras: 54% in codon 12, 42% in codon 13 (particularly G-->A transition) and 4% in both. Codon 12 K-ras mutations were associated with mucinous histotype (P <0.01), while codon 13 K-ras mutations were associated with advanced Dukes' stage (P <0.05), lymph-node metastasis (P <0.05) and high S-phase fraction (P <0.05). Multivariate analysis showed that codon 13 K-ras mutations, but not any mutation, were independently related to risk of relapse or death. CONCLUSIONS: Our results suggest that codon 12 K-ras mutations may have a role in the mucinous differentiation pathway, while codon 13 mutations have biological relevance in terms of colorectal cancer clinical outcome.     

A highly sensitive method for K-ras mutation detection is useful in diagnosis of gastrointestinal cancer

Detection of molecular features such as K-ras mutations has been used to evaluate potential tumour markers in a wide variety of clinical samples. Here we have applied a recently developed highly sensitive method for detection of K-ras codon 12 mutations to colorectal and pancreatic cancer diagnosis. We analysed 67 faecal samples from patients undergoing diagnostic colonoscopy under suspicion of colorectal cancer. PCR products were obtained in 62 of 67 (93%) faecal samples. Mutations were detected in exfoliated cells in 6 of 22 (27%) of the adenomas and in 6 of 11 (55%) of adenocarcinomas. No false positives were observed. Agreement between faecal samples and corresponding tissues was 100% for adenocarcinomas and 65% for adenomas. Mutations were also analysed in 61 pancreatic fine-needle aspirates. Mutations were detected in 36 of 45 (80%) of the pancreatic aspirates diagnosed as pancreatic cancer without false positives. Our findings suggest that, when colorectal cancer is suspected, detection of K-ras codon 12 mutations in faecal samples using this new method is specific for colorectal tumours. Additionally, this technique is a good alternative for evaluation of pancreatic masses.     

K-ras codon 12 and 13 mutations are correlated with differential patterns of tumor cell dissemination in colorectal cancer patients

The aim of this prospective study was to relate the incidences of cytokeratin 20 (CK20) and guanylylcyclase C (GCC) in lymph node, liver, and bone marrow specimens of 245 colorectal cancer (CRC) patients with the K-ras oncogene status of the corresponding primary tumor. Qualitative RT-PCR detection of CK20 and GCC mRNA was used as marker of circulating epithelial cells (CEC). Samples were considered positive for CEC only when both markers were detected concomitantly. For the detection of K-ras mutations, a PCR-RFLP assay was used. In the group with K-ras mutated primary carcinomas (n=92), CEC were detected in 62% of lymph node-, 43% of liver-, and 2% of bone marrow samples. No statistical significance was found when comparing these results with those from patients with K-ras wild-type carcinoma (59%, 46%, and 0%, respectively). In contrast to this combined evaluation, separate analysis of K-ras codons 12 (n=75, 82%) and 13 (n=17, 18%) revealed significantly differing CEC incidences. Lymph node specimens from corresponding K-ras codon 13 mutated carcinomas showed a significantly higher CEC incidence (82%) than the groups with codon 12 mutation (57%, p<0.05) or K-ras wild-type sequence (59%, p<0.05). Unlike these findings in lymph nodes, liver biopsies from corresponding carcinomas with K-ras codon 12 mutation or wild-type sequence were significantly more often positive for CEC (31% and 29%) than specimens from K-ras codon 13 mutated primary CRC (12%, p<0.04, respectively). In conclusion, colorectal carcinomas with K-ras codon 12 mutation showed the same pattern of tumor cell dissemination as their K-ras wild-type counterparts. Since K-ras codon 12 mutations prevailed 4-fold over codon 13 mutations, combined analysis of the two codons showed the same result. However, sub-analysis of patients with K-ras codon 13 mutation revealed that the respective CEC incidence was significantly increased in lymph nodes, but decreased in liver biopsies.     

APC gene mutations and colorectal adenomatosis in familial adenomatous polyposis

Correlations between germline APC mutation sites and colorectal pathophenotypes, as evaluated by the direct count of adenomas at colectomy, were investigated analysing colectomy specimens from 29 FAP patients carrying one mis-sense (codon 208) and 14 frame-shift or non-sense APC mutations (codons 232, 367, 437, 623, 876, 995, 1061, 1068, 1075, 1112, 1114, 1309, 1324, 1556). The mis-sense mutation at codon 208 was associated with a relatively mild colorectal pathophenotype. The mutation at codon 367, subject to alternative splicing, was associated with attenuated FAP. The mutation at codon 1309 was associated with the profuse colorectal adenomatosis. For 13 mutations, predicted to result in null alleles or truncated APC proteins, we correlated density and distribution of colorectal adenomas with the predicted functional effects of the mutation. The most severe colorectal pathophenotype was significantly associated with the truncating mutation at codon 1309, which is located downstream to the I beta-catenin binding domain but upstream II beta-catenin-binding domain. Mutations between codons 867 and 1114, which affect the I beta-catenin binding domain, as well as mutations occurring in exons 6 and 9, predicted to result in null alleles, were associated with a less severe colorectal pathophenotype. Overall, the highest number of adenomas was detected in the right colon, followed by the left colon, transverse colon sigma and rectum. However, the highest density of adenomas was observed in the left colon, followed by the right colon, sigma, transverse colon and rectum. Colorectal carcinomas, observed in only five patients, were all in the left colon.     

DNA methylation patterns in adenomas from FAP, multiple adenoma and sporadic colorectal carcinoma patients

Colorectal adenomas have traditionally been regarded as homogeneous. The aim of our study was to identify molecular features that may differentiate sporadic adenomas from familial adenomas such as Familial Adenomatous Polyposis (FAP) and Multiple Adenoma patients. DNA methylation was tested at Methylated IN Tumor (MINT) loci (1,2,12,31) and the CpG promoter region of genes MLH1, HPP1, MGMT, p14ARF and p16INK4a in FAP-associated adenomas (33) from 5 patients with a known APC mutation (Group 1, FAP), adenomas (29) from 4 Multiple Adenoma patients (Group 2 Multiple), adenomas (14) from 3 patients with sporadic colorectal cancers showing high microsatellite instability (Group 3, MSI-H) and adenomas (16) from 7 patients, with sporadic colorectal cancers showing microsatellite stable or low level instability (Group 4, MSS/MSI-L). Aberrant Crypt Foci (ACFs), Hyperplastic Polyps (HPs) and cancers were also examined for methylation status as well as K-ras mutation. Multiple Adenoma patients were examined for germline polymorphisms in the base excision repair gene, MYH. The familial syndrome, FAP -associated adenomas showed a significantly low frequency of MINT methylation (15.5%,) compared to sporadic MSS/MSI-L-associated adenomas (35.5%). Group 3 (MSI-H) adenomas were different in that many showed serration and a high level of methylation (57.1%). Group 2, Multiple Adenoma cases, resembled sporadic MSS/MSI-L-associated adenomas. However the promoter regions of key genes, MGMT, p14ARF and p16INK4a were methylated to a greater extent than MINTs in both sporadic and familial adenomas. Genetic profiling of adenomas supports the concept that adenomas belonging to familial syndromes pursue a different pathway to tumorigenesis than their sporadic counterpar/ts from their earliest formation.     

Both BRAF and KRAS mutations are rare in colorectal carcinomas from patients with hereditary nonpolyposis colorectal cancer

The BRAF mutations have been suggested to be linked with defective mismatch repair in colorectal carcinomas. To clarify the extent of BRAF mutations in HNPCC colorectal carcinomas, which are typical mismatch repair deficient carcinomas, we compared the frequency of BRAF mutations between HNPCC, familial adenomatous polyposis (FAP) and sporadic cases. The frequency of KRAS mutations was also compared between these three syndromes. No BRAF mutations were detected in 33 HNPCC colorectal carcinomas, while they were detected in 3 of 26 (12%) FAP carcinomas and 2 of 53 (4%) microsatellite stable sporadic carcinomas. KRAS mutations were detected in 2 of 33 (6%) HNPCC, 9 of 26 (35%) FAP and 18 of 53 (34%) sporadic carcinomas. Such extremely low frequencies of BRAF and KRAS mutations in HNPCC colorectal carcinomas suggest that the participation of RAS-RAF signaling is minor in HNPCC, and that the previously suggested high frequency of BRAF mutations in mismatch repair deficient colorectal carcinomas is not due to mutations of mismatch repair genes.     

K-ras mutation in colorectal carcinomas from singapore

54 sporadic colorectal cancers were analyzed for aberrations in the K-ras oncogene. DNA was extracted from frozen tissues obtained from surgical resection and analyzed for mutations in codons 12, 13 and 61 of the K-ras oncogene using single strand conformation polymorphism analysis (SSCP) and direct sequencing. Point mutations in the K-ras oncogene were found in 26/54 (48%) cases, all of which resulted in amino acid substitutions. No other types of mutations (e.g. insertions or deletions) were found. 4 of the mutations were at codon 12, 22 in codon 13 and only 1 was a codon 61 mutant. G-->A transitions were found to be predominant. A remarkable finding was the high preponderance of (13)Gly-(13)Ser mutations (54%). No correlation was observed between K-ras mutations and tumor location, Dukes' stage, differentiation levels, age or sex of the patient.     

The frequency and spectrum of K-ras mutations among Iraqi patients with sporadic colorectal carcinoma

Background: The epidemiology of colorectal cancers (CRC) is well known to differ in different geographical regions. K-ras mutations have been implicated in CRC carcinogenesis and they were extensively studied in developed countries; however, such studies are scarce from developing countries, like Iraq. Aim: To determine the frequency and spectrum of K-ras mutations among CRC Iraqi patients, and their clinico-pathological associations, if any. Materials and Methods: Fifty consecutive surgically resected sporadic CRC were evaluated. The evaluation included screening for ten K-ras mutations in codon 12 and 13 by mutant enriched polymerase chain reaction followed by reverse hybridization to oligospecific probes. Results: Out of the 50 enrolled patients, 24 (48%) had K-ras mutations. A total of 29 mutations were identified in the tumors of the latter 24 patients (20/24 tumors had single mutations, 3/24 had double mutations and 1/24 had triple mutations). The most frequently encountered mutations were the G>T transversions and G>A transitions (41.4% each). Codon 12 mutations constituted 89.7%, while codon 13 the remaining 10.3%. The most frequent mutation was GGT>GTT (Gly>Val) of codon 12 documented in 31%. No significant clinico-pathological correlations with K-ras mutational status were identified. Conclusion : The K-ras mutations are frequently encountered among Iraqi sporadic CRC patients, with relative higher frequencies of G>T transversions and Gly>Val codon 12 substitutions than encountered in their counterparts in developed countries. The latter is most likely to be related to differences in local carcinogens exposure, an aspect which requires further scrutiny.