Restriction fragment length polymorphism of the L-myc gene and susceptibility to metastasis in renal cancer patients

We examined Southern blot analyses of normal and tumor DNAs from 50 patients with sporadic renal cancer, using the human L-myc oncogene fragment as a hybridization probe. Our purpose was to study the relationship between the restriction fragment length polymorphism (RFLP) of the L-myc and the frequencies of metastases. There was no individual difference in patterns of L-myc RFLP between normal and tumor-tissue DNAs digested with EcoRI. The patients were classified into 3 genetic types according to the polymorphic patterns defined by the 2 alleles [10-kilobase (kb) and 6.6-kb fragments]. The relative ratios of the 3 genotypes in the renal cancer patients were similar to those seen in healthy Japanese. However, of 16 patients who exhibited distant organ metastases at the time of surgery, only one was a 10-kb fragment homozygote. The incidence of distant metastases in 10-kb homozygotes was significantly lower than that in 6.6-kb homozygotes plus heterozygotes (p = 0.06). These results basically correspond to the previous findings in the lung cancer patients, and suggest that L-myc RFLP is a widely applicable genetic marker to predict prognosis in cancer patients.     

A new polymorphism (Ser362Thr) of the L-myc gene is not associated with lung adenocarcinoma risk and prognosis.

The non-coding variation in the second intron of the L-myc gene, generating an EcoRI polymorphism, is associated with lung cancer risk and prognosis. We carried out sequence analysis of the L-myc gene in lung adenocarcinoma (ADCA) patients to identify functional polymorphisms and identified a new single nucleotide polymorphism (SNP) in the third exon of the gene causing a Ser362Thr conservative amino acid change in the C-terminus of the encoded protein. This polymorphism showed significant linkage disequilibrium with the L-myc EcoRI polymorphism located at 1751 bp distance. Genotyping of the Ser362Thr SNP in 220 Italian ADCA patients and in 230 general population controls revealed a similar low frequency (0.10-0.11) of the Thr allele in both groups. The multivariate odds ratio was 0.68 (95% confidence interval (CI) 0.38-1.22). In the ADCA patients, no significant association between the Ser/Thr polymorphism and survival was observed. Thus, the present results do not support candidacy of the L-myc Ser362Thr polymorphism for the functional polymorphism of the L-myc genomic region.     

Restriction fragment length polymorphism analysis of the L-myc gene locus in a case-control study of lung cancer

The L-myc DNA-restriction fragment length polymorphism, revealed by EcoRI, has been studied in both a lung cancer case-control framework and a cohort of 40 non-diseased unrelated individuals. No association was found between the L-myc allelic frequencies and disease status, tumor stage or lung cancer histology. A strong association was, however, observed between the L-myc allelic frequencies and ethnic origin (black or white) of the subjects. Among American whites the allelic distribution at the L-myc proto-oncogene locus was almost identical to that previously reported for Japanese subjects. Among the American black population there was a significantly higher frequency of the presence of the polymorphic EcoRI restriction site in the second intron of the L-myc proto-oncogene. These data emphasize the importance of conducting epidemiologic studies that control for ethnic factors and indicate that L-myc EcoRI allelotypes do not appear to be predictive of lung cancer risk or disease status in American blacks and whites.     

Correlation of L-myc RFLP with metastasis, prognosis and multiple cancer in lung-cancer patients.

For further study of the correlation of L-myc restriction-fragment-length polymorphism (RFLP) and metastasis of lung cancer to lymph nodes or other organs at the time of surgery, L-myc RFLP was analyzed in 252 Japanese lung-cancer patients. A close correlation between L-myc RFLP and metastasis was confirmed in this large number of patients (p = 0.01). The correlation was particularly pronounced in cases of adenocarcinoma and squamous-cell carcinoma. Poor prognosis (additional metastases after surgery) was observed in lung-cancer patients with L-S (identified as long and short bands produced with EcoRI) and S-S type L-myc RFLP. In addition, the death rate of lung-cancer patients with the L-S and S-S types was greater than that of those with the L-L type. Lung-cancer patients of the L-S and S-S types had almost 4 times higher incidence of multiple cancer in the lung, pharynx and other organs than those with the L-L type. Our results indicate that, in patients with lung cancer, genetic disposition with respect to the L-myc gene influences the extent of metastasis, the incidence of multiple cancers and prognosis.     

Rflp analysis of the L-myc oncogene in head and neck-cancer - relationship study with susceptibility and disease progression

The L-myc DNA restriction fragment length polymorphism (RFLF), revealed by EcoRI digestion, has been evaluated in a case-control study including 161 head and neck cancer (HNSCC) patients and 160 normal healthy individuals with similar smoking and alcohol habits. No significant difference in the distribution of L-myc genotypes (LL, LS or SS) was found between the two populations implying thus no predisposition to head and neck tumour by either allele. There was no significant association between L-myc genotypes and the usual clinicopathological features such as T staging, differentiation status and lymph node involvement. Moreover, follow-up data from 154 patients was obtained and correlated with the L-myc pattern. No significant difference was observed in metastasis occurrence, multiple cancer incidence and survival data in the patients classified according to the L-myc genotypes; only a trend to preferentially develop metastasis in lung for patients with S allele was noted. In conclusion, our data shows that the L-myc typing does not contribute to HNSCC risk or prognosis assessment. A review of L-myc RFLP published studies shows contradictory results even on the same type of tumour and emphasizes the lacunae in understanding the biological role of L-myc for valid interpretation of L-myc allelic associations with cancer susceptibility or prognosis.     

Meta-analysis suggests association of L-myc EcoRI polymorphism with cancer prognosis.

The L-myc EcoRI polymorphism is a noncoding variation in the second intron of the L-myc gene, resulting in S and L alleles. Individuals carrying the S allele tend to have poor prognosis and increased risk of several tumor types, although controversial results have been reported. A meta-analysis of 36 studies on L-myc EcoRI genotyping, including 3563 patients with different types of cancer and 2953 controls, was performed. In lung cancer patients the S/S genotype was significantly associated with lymph node metastasis [odds ratio (OR), 2.8; 95% confidence interval (CI), 1.8-4.3], distant metastasis (OR, 4.7; 95% CI, 2.4-9.2), and stage (OR, 2.3; 95% CI, 1.2-4.4). No association was observed between the S/S genotype and cancer (OR, 1.1; 95% CI, 0.8-1.4). In patients with other cancers, the S/S genotype was significantly associated with tumor recurrence (OR, 2.8; 95% CI, 1.4-6.0), whereas no significant association was seen for the other prognostic parameters. When all types of cancer were examined together, the S/S genotype was associated with lymph node metastasis (OR, 2.3; 95% CI, 1.6-3.3), distant metastasis (OR, 2.9; 95% CI, 1.8-4.6), clinical stage (OR, 1.8; 95% CI, 1.2-2.9), and cancer risk (OR, 1.25; 95% CI, 1.07-1.45). The meta-analysis suggests that the L-myc EcoRI polymorphism is a marker of tumor prognosis in lung cancer and possibly in other types of cancer.     

L-myc restriction fragment length polymorphism and histological pattern of invasion in lung adenocarcinoma

It is still controversial whether restriction fragment length polymorphism (RFLP) of the L-myc proto-oncogene is correlated with the prognosis in lung cancer patients or not. On the other hand, we have shown that the histological pattern of invasion is correlated with the prognosis of surgically treated patients of lung adenocarcinoma. In this study, the L-myc RFLP was compared with the histological pattern of invasion and the prognosis in 63 Japanese lung adenocarcinoma patients to evaluate the L-myc RFLP as a prognostic marker. Although the histological pattern of invasion was correlated with the prognosis, no correlation was observed between the L-myc RFLP and the prognosis. No association was found between the L-myc RFLP and the histological pattern of invasion in this study. Our results showed that the L-myc RFLP is not a strong prognostic marker compared with histologic pattern of invasion in lung adenocarcinoma.     

L-myc Restriction Fragment Length Polymorphism in Japanese Patients with Esophageal Cancer

L-myc polymorphism is a representative genetic trait related to an individual's susceptibility to several cancers. However, there have been no reports concerning the association between esophageal cancer and L-myc polymorphism. To analyze the distribution of polymorphism in Japanese patients with esophageal cancer, a molecular genotyping method using a polymerase chain reaction-based restriction fragment length polymorphism (PCR-RFLP) was used. Based on an analysis of 65 Japanese patients with esophageal cancer and 107 healthy control subjects, a significant difference was observed in either the distribution of genotypes ( P =0.012) or of allele frequencies between the two groups ( P 0.004). The relative risk of esophageal cancer for genotypes including the shorter allele was 2.9 compared to the longer allele homozygote. Furthermore, the patients with S-allele had a tendency for poor prognosis among those with three genotypes. A significant difference between the distribution of genotypes and the incidence of lymph node metastasis was found based on the clinicopathological features of the cancers. These results suggest that L-myc polymorphism may be implicated as a genetic trait affecting an individual's susceptibility to esophageal cancer, at least among Japanese patients.     

L-myc restriction fragment length polymorphism in Japanese patients with esophageal cancer.

L-myc polymorphism is a representative genetic trait related to an individual's susceptibility to several cancers. However, there have been no reports concerning the association between esophageal cancer and L-myc polymorphism. To analyze the distribution of polymorphism in Japanese patients with esophageal cancer, a molecular genotyping method using a polymerase chain reaction-based restriction fragment length polymorphism (PCR-RFLP) was used. Based on an analysis of 65 Japanese patients with esophageal cancer and 107 healthy control subjects, a significant difference was observed in either the distribution of genotypes (P=0.012) or of allele frequencies between the two groups (P=0.004). The relative risk of esophageal cancer for genotypes including the shorter allele was 2.9 compared to the longer allele homozygote. Furthermore, the patients with S-allele had a tendency for poor prognosis among those with three genotypes. A significant difference between the distribution of genotypes and the incidence of lymph node metastasis was found based on the clinicopathological features of the cancers. These results suggest that L-myc polymorphism may be implicated as a genetic trait affecting an individual's susceptibility to esophageal cancer, at least among Japanese patients.     

Population-based mapping of pulmonary adenoma susceptibility 1 locus

Pulmonary adenoma susceptibility 1 (Pas1), the major locus affecting inherited predisposition to lung tumor development in mice, maps near the Kras2 gene. We previously reported a significant association between a KRAS2/RsaI polymorphism and the risk and prognosis of lung adenocarcinoma (ADCA) in the Italian population. In the present case-control study, we examined 269 lung ADCA patients, 121 squamous cell lung carcinoma patients, and 632 healthy individuals (general population controls) in the Japanese population with genetic markers spanning approximately 1200 kb in the KRAS2 region. Allele-specific oligonucleotide hybridization revealed the same KRAS2/RsaI polymorphism associated with risk and prognosis as in Italian lung ADCA patients; the polymorphism was significantly associated with clinical stage (P < 0.001) and survival rate (log rank = 0.0014), confirming the mapping of PAS1 and pointing to the role of this locus in human lung cancer.     

Ethnic differences in frequencies of gene polymorphisms in the MYCL1 region and modulation of lung cancer patients' survival

Linkage disequilibrium (LD) analysis to refine a region associated with lung cancer progression on chromosome 1p34 identified a 106 kb LD block that includes MYCL1, TRIT1 (tRNA isopentenyltransferase 1) and MFSD2 (major facilitator superfamily domain-containing 2). Case-only association study on SNPs mapping in TRIT1 and MFSD2 indicated that the rare Leu allele (frequency: 0.04) of the TRIT1 Phe202Leu variation predicts short survival as compared to the common Phe/Phe genotype (hazard ratio (HR)=1.7; 95% CI, 1.03-2.86; P=0.039) in 335 Italian lung adenocarcinoma samples. A replication study in an independent population of 246 Norwegian lung cancer patients confirmed the significant association of the Phe202Leu polymorphism with patients' survival, but the rare allele was associated with better survival rate (HR=0.5; 95% CI, 0.26-0.91; P=0.023). The rare allele of TRIT1 Phe202Leu SNP was approximately seven-fold more frequent in Asian than in Caucasian subjects and three additional SNPs in the TRIT1 and MFSD2 genes showed ethnic differences in allelic frequencies. These results suggest that polymorphisms in the MYCL1 LD region affect lung cancer survival but that the functional element(s) may show population-specific patterns.     

Association between restriction fragment length polymorphism of the L-myc gene and lung metastasis in human breast cancer.

EcoRI restriction fragment length polymorphism (RFLP) of the L-myc gene was examined in leukocyte DNAs isolated from 381 breast cancer patients. No differences in the patterns of L-myc RFLP were found between breast cancer patients and healthy individuals. However, among 97 patients who relapsed, a statistical correlation was found between L-myc RFLP and lung metastases (p less than 0.05). These results are in close agreement with previous findings in patients with cancer of the lung, bone or kidney, and suggest that L-myc RFLP may be a useful marker for predicting lung metastasis in some human cancers.     

L-myc genotype is associated with different susceptibility to lung cancer in smokers.

We have shown that L-myc genotype is associated with the risk of esophageal cancer from smoking and heavy drinking. In this study, we have analyzed the relationship between the L-myc genotypes and lung cancer risk from smoking in 191 Japanese lung-cancer patients and 241 non-cancer controls. The odds ratios (ORs) were markedly higher in SS and LS genotypes than in LL genotype; age-sex-adjusted ORs were 3.19, 2.30 and 0.92, respectively. This result suggests that the L-myc polymorphism may affect the induction of lung cancer by smoking. The OR for smoking in SS-genotype patients diagnosed within 2 years was higher than that in other SS patients, suggesting that smoking-related lung cancer in SS genotype might exhibit a poorer prognosis.     

L-myc Genotype is Associated with Different Susceptibility to Lung Cancer in Smokers

We have shown that L-myc genotype is associated with the risk of esophageal cancer from smoking and heavy drinking. In this study, we have analyzed the relationship between the L-myc genotypes and lung cancer risk from smoking in 191 Japanese lung-cancer patients and 241 non-cancer controls. The odds ratios (ORs) were markedly higher in SS and LS genotypes than in LL genotype; age-sex-adjusted ORs were 3.19, 2.30 and 0.92, respectively. This result suggests that the L-myc polymorphism may affect the induction of lung cancer by smoking. The OR for smoking in SS-genotype patients diagnosed within 2 years was higher than that in other SS patients, suggesting that smoking-related lung cancer in SS genotype might exhibit a poorer prognosis.     

L-myc Proto-oncogene alleles and susceptibility to hepatocellular carcinoma

A common inherited RFLP of the L-myc proto-oncogene has been reported to correlate with cancer susceptibility or prognosis for some tumors. Using a PCR-based RFLP assay on DNA extracted from peripheral blood samples, we determined the L-myc EcoRI genotype of patients with HCC, patients with other liver tumors, and healthy controls. In this polymorphism there are 2 alleles, L and S, which define 3 possible genotypes: LL, LS, and SS. While the genotype distribution of patients with other liver tumors and healthy controls do not differ from one another, both differ significantly from the genotype distribution of patients with HCC. This difference is primarily the result of a low frequency of the SS genotype among HCC patients compared to controls or other liver tumor patients. Persons with the SS genotype appear to be protected against HCC and have roughly one-ninth the risk of persons who carry one or more copies of the L allele. Within HCC cases, patients with different L-myc genotypes show slight, but not statistically significant, differences in tumor characteristics and survival.     

Studies of the L-myc DNA polymorphism and relation to metastasis in Norwegian lung cancer patients

We studied 83 lung cancer patients and 129 controls for the EcoRI polymorphism of the L-myc gene. No association was found between the L-myc RFLP and increased risk of metastasis, either to lymph nodes or metastasis to other organs. There was no difference in survival time between the three different genotypes. The S-allele of the L-myc RFLP has been correlated to increased metastasis in lung cancer. We found no tendency towards a higher frequency of this allele in the cohort of patients with positive family history compared to the patients with no known first degree relatives with cancer. A higher frequency of the S-allele in the adenocarcinomas compared to other histological groups was found, although this difference was not statistically significant. No difference in the gene frequency of the L-myc RFLP was found between the lung cancer patients and the normal controls. These results are in contrast with a previous report. Possible explanations for the discrepancies are discussed.     

Oncogene amplification in squamous cell carcinoma of the oral cavity

We have determined the prevalence of amplification of c-myc, N-myc, L-myc, H-ras, Ki-ras, and N-ras oncogenes in 23 cases of squamous cell carcinoma of the oral cavity, using Southern hybridization analysis of DNA extracted from the primary tumor tissues. Nick-translated oncogene probes and oncogene inserts labeled to high specific activities were used. We observed a 5- to 10-fold amplification of one or more of c-myc, N-myc, Ki-ras and N-ras oncogenes in 56% of the tumor tissue samples, with these oncogenes not being amplified in the peripheral blood cells of the same patients. L-myc and H-ras were not amplified in any of our samples. The oncogene amplifications seemed to be associated with advanced stages of squamous cell carcinomas, with the ras and myc family oncogenes being amplified in stages 3 and 4. Hybridization with N-myc detected an additional 2.3 kb EcoRI fragment, along with the normal 2.1 kb fragment. Our data also demonstrated amplification of multiple oncogenes in the same tumor tissue sample. About 60% of the samples with amplified oncogenes showed simultaneous amplification of 2 or more oncogenes. The results showing different oncogene amplifications in similar tumors, as well as multiple oncogene amplifications in the same tumor, suggest that these oncogenes may be alternatively or simultaneously activated in oral carcinogenesis.     

Association of L-myc polymorphism with lung cancer susceptibility and prognosis in relation to age-selected controls and stratified cases

The association of L-myc polymorphism with cancer susceptibility and prognosis has produced conflicting results. This may have been due to racial/ethnic differences and methodological variations in the studies, such as, control selection and case stratification. Therefore, we investigated the genotype distribution of the L-myc polymorphism in 169 lung cancer patients and 169 non-cancer controls, and analyzed the association of this polymorphism with cancer susceptibility and prognosis in relation to age-specific controls as well as stratified cases. The genotype frequencies in the Taiwanese non-cancer controls were 0.56 (L) and 0.44 (S). Chi-square (chi(2)) analysis indicated a significant difference in the Taiwanese genotype distribution of L-myc compared with that of African-Americans (P=0.001). Logistic regression analysis of cases/controls, adjusted for both age and sex, indicated that an increased frequency of the LL genotype was observed in early-staged patients compared with the non-cancer controls (OR=0.43, 95% CI, 0.20-0.94, P=0.03). In addition, the frequency of the LL genotype was significantly higher in stages I+II patients (47.4%) than in stages III+IV patients (28.4%) (P=0.05). Furthermore, the S allele frequency was significantly increased in stages III+IV patients (P=0.005). As both L-myc and p53 polymorphisms were analyzed for their prognostic value, the patients with an S allele of the L-myc gene and a Pro/Pro variant genotype of the p53 gene had significantly poorer prognoses compared with other patients (P=0.004, by the log rank test). These data suggest that the S allele of the L-myc polymorphism may be associated with lung cancer progression.     

Increased frequency of the S allele of the L-myc oncogene in non-Hodgkin's lymphoma.

We studied 100 patients with non-Hodgkin''s lymphoma, 44 patients with Hodgkin''s disease and 100 controls for the prevalence of the EcoRI restriction fragment polymorphism of the L-myc oncogene. No difference in the frequency of the three genotypes (LL, LS, SS) was found between the patient and control groups. However, the S allele was found to occur more frequently in the non-Hodgkin''s lymphoma patients (chi 2 = 4.57, P = 0.032). These data confirm an earlier report and suggest that the presence of the S allele is associated with susceptibility to non-Hodgkin''s lymphoma.