Tatsuro Yamaguchi

Overexpression of soluble vascular endothelial growth factor receptor 1 in colorectal cancer: Association with progression and prognosis.

We examined the expression of sVEGFR1 in colorectal cancer tissue and corresponding normal colorectal mucosa to assess the clinical significance of sVEGFR1 in colorectal cancer. We also assessed the relationship between sVEGFR1 levels and various clinicopathologic factors and prognoses. sVEGFR1 and VEGF levels were measured in fresh‐frozen tumor extracts from 84 primary colorectal cancer tissues and 27 corresponding normal mucosa using ELISA. Mean of sVEGFR1 levels were 3.17 ng/mg protein. sVEGFR1 levels increased significantly in cancer tissue compared with normal mucosa. Although VEGF levels increased in cancer tissues, the ratio of sVEGFR1/VEGF in cancer tissue was significantly lower than that in normal tissue. No significant correlation between sVEGFR1 or VEGF levels and any clinicopathologic parameter was found. Overexpression of sVEGFR1 was significantly associated with a favorable prognosis. Based on sVEGFR1 levels in colorectal cancer without distant metastases, patients with higher sVEGFR1 levels (≥1.5 ng/mg protein) demonstrated significant longer recurrence‐free survival than patients with lower sVEGFR1 levels (<1.5 ng/mg protein) (P = 0.0017). Multivariate analysis showed that the sVEGFR1 levels in cancer tissue were an independent prognostic indicator of disease progression. sVEGFR1 expression was significantly elevated in colorectal cancer tissue compared with normal mucosa and the intratumoral balance between sVEGFR1 and VEGF was significantly different between tumor tissue and normal controls. Furthermore, sVEGFR1 levels showed a significant prognostic value. Further studies concerning the biologic and therapeutic significance of sVEGFR1 in colorectal cancer are warranted. (Cancer Sci 2007; 98: 405–410)

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.

Accumulation profile of frameshift mutations during development and progression of colorectal cancer from patients with hereditary nonpolyposis colorectal cancer

PurposeRole and timing of frameshift mutations during carcinogenesis in hereditary nonpolyposis colorectal cancer have not been examined. This study was designed to clarify the relationship between frameshift mutations and clinicopathologic features in colorectal cancer from patients with hereditary nonpolyposis colorectal cancer.MethodsThirty-one colorectal cancers from patients with hereditary nonpolyposis colorectal cancer at different clinicopathologic stages were analyzed for frameshift mutation in 18 genes.ResultsThe frameshift mutations of the ACVR2 and PTHLH genes were found to have an extremely high frequency (94–100 percent) in all pathologic stages, and mutation of the MARCKS gene also was high (94 percent) in Dukes B and C cancers. These frequencies were higher than the frequency of TGFβRII gene inactivation (64–88 percent). Mutations of the hMSH3, TCF4, CASP5, RIZ, RAD50, and MBD4 genes were comparatively frequent (>35 percent) in all stages. Frequencies of inactivation of the MARCKS, BAX, IGFIIR, and PTEN genes were significantly higher in Dukes B and C cancers than in Dukes A cancer (P < 0.05). The number of accumulated frameshift mutations was larger in Dukes B and C cancers (9.4) than in Dukes A cancer (6.8) (P = 0.003).ConclusionsThe present data suggest that the disruption of the transforming growth factor-β super-family signaling pathway by the alteration of the ACVR2 and/or TGFβRII genes and the disruption of antiproliferative function by the PTHLH gene alteration contribute to the development of early colorectal cancer. Moreover, the further accumulation of alterations in the MARCKS, BAX, IGFIIR, and PTEN genes seem to be associated with progression from early to advanced colorectal cancer from patients with hereditary nonpolyposis colorectal cancer.

Difference in characteristics of APC mutations between colonic and extracolonic tumors of FAP patients: Variations with phenotype

To clarify the differences in characteristics of adenomatous polyposis coli (APC) mutations between colorectal tumors from various phenotypes of familial adenomatous polyposis (FAP) and between colorectal and extracolonic tumors, we analyzed APC mutations in 86 colorectal tumors from FAP patients including profuse, sparse and attenuated types, 23 sporadic colorectal tumors and 40 FAP extracolonic tumors including duodenal, gastric and desmoid tumors. In all tumors, 1 allele of the truncated APC gene retained armadillo repeats, 15‐amino‐acid (aa) repeats and various numbers of 20‐aa repeats, but lacked SAMP repeats, as a result of germline and somatic mutations. Regarding 20‐aa repeats, the truncated APC gene retained 1 repeat due to allele loss in 96% (27/28) of colorectal tumors from profuse‐type FAP, 69% (36/52) of sparse‐type retained 2 repeats due to somatic mutation, and 100% (6/6) of attenuated‐type retained 2 or 3 repeats due to the third or second hit. In sporadic colorectal tumors 74% (17/23) retained 1 or 2 repeats. The truncated APC gene retained 3 repeats in 88% (7/8) of FAP duodenal tumors, 100% (26/26) of gastric tumors retained 2 or 3 repeats and 83% (5/6) of desmoid tumors retained 2 repeats. These data suggest that the number of β‐catenin downregulating 20‐aa repeats in truncated APC gene associated with colorectal tumorigenesis is different in profuse, sparse and attenuated types of FAP, and that the association with tumorigenesis is also different between colorectal and extracolonic tumors.

Somatic Mutations of the CDC4 (FBXW7) Gene in Hereditary Colorectal Tumors

Objectives: Cdc4 (Fbxw7) is a potential tumor suppressor that regulates ubiquitination and proteolysis of multiple targets such as cyclin E, c-Myc, c-Jun and Notch. CDC4 mutations were investigated in 194 colorectal carcinomas and adenomas for comparison between sporadic and hereditary cancers. Methods: Mutations of the CDC4 gene were analyzed by PCR-SSCP and sequencing, and loss of heterozygosity (LOH) was analyzed by microsatellite marker analysis. Results: Somatic CDC4 mutations were detected in 9% (3 of 33) of hereditary nonpolyposis colorectal cancer (HNPCC), 9% (3 of 33) of familial adenomatous polyposis (FAP) carcinomas, and 10% (7 of 73) of sporadic carcinomas. CDC4 mutations were also detected in adenomas at frequencies of 6% (2 of 31) and 4% (1 of 24) in FAP and sporadic cases, respectively. Frameshift mutations were observed in HNPCC tumors, while single-base substitutions predominantly occurred in FAP and sporadic tumors. LOH at the chromosome 4q region was rarely detected in tumors with CDC4 mutations. Conclusions: The results indicate that the frequency of CDC4 mutations in colorectal tumors is similar in patients with HNPCC and FAP compared to patients with sporadic carcinomas. Moreover, infrequent LOH suggests that the CDC4 gene does not follow the general 2-hit model.

Prognostic factors and treatment effects in patients with curatively resected brain metastasis from colorectal cancer.

BACKGROUND: Colorectal cancer infrequently causes brain metastasis, and the prognosis is poor. OBJECTIVE: The aim of this study was to identify the prognostic factors associated with survival and outcome of treatment for patients with brain metastasis from colorectal cancer. DESIGN: This is a retrospective study from a prospectively collected database. SETTINGS: The investigation took place in a high-volume multidisciplinary tertiary cancer center in Japan. PATIENTS: From 1979 to 2010, 113 consecutive patients who were treated for brain metastasis from colorectal cancer were identified. MAIN OUTCOME MEASURES: The primary outcome measure was overall survival. RESULTS: Sixty-three patients had neurosurgical resection (including curative resection for 46 patients) followed by whole brain radiotherapy, 9 had stereotactic radiosurgery, 30 had whole brain radiotherapy, and 11 had steroid and palliative care. As a whole, the overall median survival time from diagnosis of brain metastasis was 5.4 months (95% CI, 4.3–7.6 months), and the 1-year survival rate was 29% (95% CI, 22%–38%). In the group of patients who underwent curative neurosurgical resection, the overall median survival time was 15.2 months (95% CI, 9.2–17.8 months), and the 1-year survival rate was 57% (95% CI, 43%–71%). On multivariate analysis, 1 or 2 brain metastatic lesions, no extracranial metastatic lesions, and neurosurgical resection were independent favorable prognostic factors overall (p = 0.0057, 0.0197, and <0.0001), and 1 or 2 brain metastatic lesions, no extracranial metastatic lesion, and no emergence of secondary brain metastatic lesions were independent favorable prognostic factors in the group of patients who underwent curative neurosurgical resection (p = 0.0137, 0.0081, and 0.0010). LIMITATIONS: This study was limited by its single-institute, retrospective, nonrandomized design and selection bias. CONCLUSIONS: Neurosurgical resection in select patients is a reasonable option for brain metastasis from colorectal cancer, although it is not associated with long-term (5-year) survival. (see Video, Supplemental Digital Content 1, http://links.lww.com/DCR/A121 )

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.

Prognostic Impact of Primary Tumor Location on Clinical Outcomes of Metastatic Colorectal Cancer Treated With Cetuximab Plus Oxaliplatin-Based Chemotherapy: A Subgroup Analysis of the JACCRO CC-05/06 Trials

Introduction: Primary tumor location is a critical prognostic factor in metastatic colorectal cancer (mCRC); however, it remains unclear whether tumor location is a predictor of the response to cetuximab treatment. It is also uncertain if BRAF mutation contributes to the impact of tumor location on survival. We assessed the prognostic impact of tumor location on clinical outcomes in mCRC patients treated with first‐line cetuximab chemotherapy. Patients and Methods: The associations of tumor location with overall survival and progression‐free survival were evaluated in mCRC patients with KRAS exon 2 wild‐type tumors who were enrolled onto 2 clinical trials: JACCRO CC‐05 of cetuximab plus FOLFOX (n = 57, UMIN000004197) and CC‐06 of cetuximab plus SOX (n = 61, UMIN000007022). Tumors proximal or from splenic flexure to rectum were defined as right‐sided or left‐sided, respectively. In addition, exploratory RAS and BRAF mutation analyses were performed. Results: A total of 110 patients were assessable for tumor location; 90 had left‐sided tumors. Left‐sided tumors were significantly associated with longer overall survival (36.2 vs. 12.6 months, hazard ratio = 0.28, P < .0001) and progression‐free survival (11.1 vs. 5.6 months, hazard ratio = 0.47, P = .0041) than right‐sided tumors; similar results were obtained in multivariate analysis. A subanalysis showed that the association was evident in the FOLFOX group and that tumor location was an independent prognostic factor irrespective of BRAF status in RAS wild‐type patients. Conclusion: Primary tumor location might be a predictor of survival independent of BRAF status in mCRC patients who receive first‐line cetuximab combined with oxaliplatin‐based chemotherapy. &NA; Primary tumor location is a prognostic factor in metastatic colorectal cancer (mCRC). We assessed the prognostic impact of tumor location on survival and the association between BRAF mutation and tumor sidedness in mCRC patients treated with cetuximab. Tumor location is a prognostic marker for first‐line cetuximab plus oxaliplatin‐based chemotherapy, irrespective of BRAF status.

Underexpression of miR-126 and miR-20b in Hereditary and Nonhereditary Colorectal Tumors

Objectives: The aim of the study was to determine the significance of miR-126 and miR-20b in colorectal carcinogenesis. Methods: We analyzed the expressions of miR-126 and miR-20b in 136 colorectal tumors from 39 microsatellite stable (MSS) tumors, 23 high microsatellite instability (MSI-H) tumors, 16 Lynch syndrome, and 58 familial adenomatous polyposis (FAP) tumors including adenoma, intramucosal carcinoma, and invasive carcinoma. Results: All four kinds of tumors showed underexpression of both miR-126 and miR-20b. The frequency of miR-126 downregulation was 100.0% in FAP adenomas, 85.7% in FAP intramucosal carcinomas, 78.9% in invasive carcinomas, 81.3% in Lynch syndrome tumors, 68.4% in MSS tumors, and 65.4% in MSI-H tumors. The frequency of miR-20b downregulation was 64.0% in FAP adenomas, 50.0% in FAP intramucosal carcinomas, 73.3% in invasive carcinomas, 62.5% in Lynch syndrome tumors, 79.5% in MSS tumors, and 91.3% in MSI-H tumors. The current study demonstrated underexpression of miR-126 and miR-20b in various types of colorectal cancer. These findings support the hypothesis that angiogenesis results from underexpressions of miR-126 and miR-20b and occurs as an early event in colorectal carcinogenesis. Conclusions: Underexpression of miR-126 and miR-20b was observed in various types of colorectal cancer, and occurs as an early event of colorectal carcinogenesis in FAP tumors.

Assessment of SMAD4, p53, and Ki-67 alterations as a predictor of liver metastasis in human colorectal cancer

PurposeThe liver is the most common site of metastasis in patients with colorectal cancer (CRC), and this is a determinant of the prognosis. However, no reliable molecular predictors of liver metastasis have yet been identified.MethodsSixty-two surgical specimens of colorectal cancer were studied. The first group included 25 patients who achieved a disease-free survival period of at least 6 years (CRC-M0), and the second group included 37 patients with synchronous (n = 22) or metachronous (n = 15) liver metastasis (CRC-M1). SMAD4, p53, and Ki-67 expression levels were assessed immunohistochemically.ResultsThe loss of SMAD4 expression and elevated Ki-67 expression were found significantly more frequently in CRC-M1 patients than in CRC-M0 patients (P = 0.0047 and P = 0.013, respectively). Statistically significant differences were also observed between the CRC-M0 group and the metachronous metastasis group. No difference was seen in the overexpression of p53 between the groups. A combination analysis of SMAD4 and Ki-67 revealed no cases with maintained levels of SMAD4 and a low Ki-67 expression had or developed liver metastasis.ConclusionThe loss of SMAD4 expression and elevated Ki-67 expression was therefore found to significantly correlate with liver metastasis, regardless of the time of occurrence, thus indicating these factors to be predictive markers for liver metastasis in patients with CRC.