Chih-Hsiung Lai

Germ line MLH1 and MSH2 mutations in Taiwanese Lynch syndrome families: characterization of a founder genomic mutation in the MLH1 gene

This multicenter study evaluated the mutation spectrum and frequencies of the MLH1 and MSH2 genes and determined the occurrence of large genomic deletions in 93 unrelated Taiwanese families that fulfilled the Amsterdam criteria II by denaturing high‐performance liquid chromatography analysis, DNA sequencing for aberrant chromatograms, and multiplex ligation‐dependent probe amplification analysis. In total, 38 pathogenic mutations (10 large deletions and 28 point mutations or small deletion/insertions) in the MSH2 or MLH1 gene were identified in 61 of the 93 families (66%). Three of the 10 large deletions and 14 of the 28 point mutations or small insertions/deletions have not been reported elsewhere. Three mutations in the MLH1 gene, the MLH1c.1846_1848delAAG (5 families), deletion exons 11–15 (4 unrelated families), and MLH1c.793C>T (13 unrelated families), accounted for 35% of all cases with pathogenic mutations. Haplotype analysis indicated that mutant c.793C>T alleles were derived from two distinct common founders that might be inherited from a single ancestor of presumably Chinese origin. As a mutation detection strategy for Taiwanese Lynch syndrome patients, we recommend that diagnosis starts with screening for large genomic deletions and continues by screening for common mutations in exons 10 and 16 of the MLH1 gene prior to searching for small mutations in the remaining exons.

Preoperative SCC Antigen, CRP Serum Levels, and Lymph Node Density in Oral Squamous Cell Carcinoma.

AbstractThe prognostic significance of squamous cell carcinoma antigen (SCC-Ag) and C-reactive protein (CRP) levels and lymph node density (LND) has been individually recognized in oral squamous cell carcinoma (OSCC). We investigated the relationship between preoperative serum markers (SCC-Ag and CRP) and postoperative prognostic marker (LND) in this study. We retrospectively analyzed 277 OSCC patients who underwent primary curative resection and neck dissection with/or without adjuvant therapy between March 2008 and November 2013. Serum SCC-Ag and CRP levels were measured preoperatively. Distant metastasis, overall survival (OS), and disease-free survival (DFS) were used to evaluate the prognostic significance of preoperative SCC-Ag and CRP levels in relation to LND. LND (cutoff point ≥0.06) correlated with the pathologic tumor status, pathologic nodal metastasis, degree of differentiation, tumor stage, tumor depth (≥10 mm vs <10 mm), and perineural invasion (all P values were <0.001). LND was significantly associated with development of distant metastasis, DFS, and OS (all P values were <0.001). Preoperative elevated CRP and SCC-Ag levels were significantly associated with LND (P = 0.006), DFS (P < 0.001), and OS (P < 0.001). LND+ patients were further stratified into prognostic groups according to their SCC-Ag and CRP levels (DFS: P = 0.010; OS: P = 0.003). LND correlated with the incidence of DM, DFS, and OS in patients with OSCC. Concurrent elevated preoperative SCC-Ag and CRP levels are predictors for LND. In addition, SCC-Ag and CRP are markers for classifying high-risk LND+ patients with OSCC into subgroups.

The Mitochondrial DNA Northeast Asia CZD Haplogroup Is Associated with Good Disease-Free Survival among Male Oral Squamous Cell Carcinoma Patients

Reprogramming of energy metabolism in cancer cells has been directly/indirectly linked to mitochondria and mitochondrial functional defects and these changes seem to contribute to the development and progression of cancer. Studies have indicated that mitochondrial DNA haplogroups are associated with risk in relation to various diseases including cancer. However, few studies have examined the effect of haplogroups on cancer prognosis outcome. In order to explore the role of haplogroups on oral squamous cell carcinoma (OSCC) prognosis, the mitochondrial genomes of 300 male OSCC patients were comprehensively analyzed by direct sequencing. They were then haplotyped and grouped into four major geographic haplogroups, namely the East Asia AN, Southeast Asia RBF, East Asia MGE and Northeast Asia CZD groups. The Kaplan-Meier plot analysis indicated that individuals who were members of the CZD haplogroup showed a significant association with better disease-free survival (DFS) than the other three haplogroups and this phenomenon still existed after adjusting for tumor stage, differentiation and age at diagnosis (hazard ratio = 0.55; 95% CI = 0.36–0.84). In addition, an interaction between membership of the RBF haplogroup and radiotherapy/chemo-radiotherapy in DFS was also identified. The results strongly support the hypothesis that an individual’s haplogroup, by defining their genomic background, plays an important role in tumor behavior and mitochondrially-targeted anticancer drugs are promising future therapeutic approaches.

Polymorphisms of DNA repair genes are associated with colorectal cancer in patients with Lynch syndrome

DNA repair genes are crucial for maintaining genomic stability by preventing mutagenesis and carcinogenesis. The present retrospective cohort study aimed at investigating whether MLH1, APEX1, MUTYH, OGG1, NUDT1, XRCC5, XPA, and ERCC2 single nucleotide polymorphisms (SNPs) are associated with colorectal cancer (CRC) in Chinese population with Lynch syndrome.

EGFR, SMAD7, and TGFBR2 Polymorphisms Are Associated with Colorectal Cancer in Patients with Lynch Syndrome

Background/Aim: Epidermal growth factor receptor (EGFR), mothers against decapentaplegic homolog 7 (SMAD7) and transforming growth factor betta (TGFB) are crucial for colorectal cancer (CRC) tumorigenesis. This study investigated whether polymorphisms in EGFR, SMAD7, and TGFB are associated with CRC risk in patients with Lynch syndrome. Materials and Methods: Genotyping was performed using Sequenom iPLEX MassArray. Association between genetic polymorphisms and CRC was assessed using a weighted Cox proportional hazard model. Results: Patients carrying the AA genotype of EGFR rs2227983 had a significantly higher CRC risk than those carrying the G allele (HR=2.55, 95% CI=1.25-5.17). The dominant model of SMAD7 rs12953717 (CT + TT genotypes) significantly increased CRC risk (HR=2.17, 95% CI=1.12-4.16) when compared to the wild-type CC genotype. Similarly, the GG genotype of TGFBR2 rs6785358 significantly increased the risk of CRC (HR=21.1, 95% CI=5.06-88.1) compared to the AA genotype. Conclusion: EGFR, SMAD7, and TGFBR2 are associated with CRC risk in patients with Lynch syndrome.

Epidermal growth factor receptor intron-1 CA repeat polymorphism on protein expression and clinical outcome in Taiwanese oral squamous cell carcinoma

This study was designed to explore the relationship between epidermal growth factor receptor (EGFR) CA repeats polymorphism and protein expression in oral cavity squamous cell carcinoma (OSCC). A total of 194 OSCCs were examined for EGFR protein overexpression, gene copy number and the length of their CA repeats. The length of the EGFR CA repeats was found not to be associated with EGFR gene copy number or with protein overexpression. To exclude the effect of EGFR gene copy number on protein overexpression, only those OSCC tumors with disomy of the EGFR gene were included in further analysis. In this subgroup, EGFR protein overexpression was significantly associated with poor differentiation of the tumor cells and lymph node metastasis, especially extra-capsular spread. However, EGFR CA repeats were not related to any clinicopathological factor. Interestingly, patients genetically found to have the EGFR CA repeats SS genotype and having tumors with EGFR protein overexpression were found to have a worst prognosis in terms of disease-free survival (DFS) (HR = 2.68; 95% CI, 1.03–6.98) after multivariate adjustment. The present study demonstrates that concurrent overexpression of EGFR protein in the presence genetically of the SS form CA repeats acts as a predictor for poor DFS.

Abstract 5260: Mitochondrial DNA phylogenetic analysis for local recurrences and second primary tumors from patients with oral cavity squamous cell carcinoma

Local recurrences (LRs) and second primary tumors (SPTs) are commonly observed in patients with oral squamous cell carcinoma (OSCC). A lower survival rate has been demonstrated in patients with LRs with respect to patients with SPTs. Therefore, the criteria for differentiating LRs and SPTs are indispensable for determining appropriate treatment. Currently, criteria based on histology, the distance and relapse time between new lesions and index primary tumor are generally used in clinical classification. In the genomic era, molecular classification based on the clonal relationship between the first and the second lesions may be more reliable than traditional clinical classification. The aim of the present study tried to establish a feasible molecular classification based on mitochondrial DNA (mtDNA) phylogenetic analysis. The study population consisted of 44 OSCC patients with multiple lesions for a total of 47 (19 LRs and 28 SPTs) paired lesions. The mtDNA analysis was performed by direct sequencing. The phylogenetic and molecular evolutionary analyses were conducted by using MEGA software version 6 and three common phylogenetic evolutionary methods (Maximun Likelihood (ML), Neighbor-Joining (NJ) and Unweighted Pair Group Method with Arithmetic Mean (UPGMA) method) were applied. Preliminary analysis indicated that the prevalence of clonality for LRs and SPTs was 26.3% and 3.6%, respectively. A discrepancy between the clinical classification and mtDNA analysis was found in 15 paired tumors (31.9%) in which 14 paired lesions were classified as LRs based on clinical criteria, while mtDNA analysis suggested that they were nonclonal-related SPTs. Survival analysis indicated that there was no difference between patients with LRs and patients with SPTs based on clinical classification. On the other hand, patients with clonal second lesions had poorer overall survival than patients with nonclonal second lesions based on mtDNA analysis. In conclusion, our preliminary findings suggested that molecular clonality analysis could improve the accuracy in differentiating LRs from SPTs. Citation Format: Chih-Hsiung Lai. Mitochondrial DNA phylogenetic analysis for local recurrences and second primary tumors from patients with oral cavity squamous cell carcinoma. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 5260. doi:10.1158/1538-7445.AM2015-5260

Abstract 4146: Polymorphisms of GSTT1, APE1 and MUTYh gene and the risk of oral squamous cell carcinoma

In Taiwan, oral cancer (including sub-sites in the oral cavity, oropharynx and hypopharynx) is the fourth most common cancer in men. According to the cancer registry report from the Ministry of Health and Welfare of the Executive Yuan, the annual age-adjusted incidence for oral cancer in males has shown a 1.55-fold increase in the past decade. Cigarette smoking, areca quid (AQ) chewing and alcohol drinking have been unveiled as the major risk factors. Previous studies have implicated that oxidative stress generated from cigarette smoking, AQ chewing and alcohol drinking contributed to oral cancer development. It is well known that GSTM1 and GSTT1 are involved in the detoxification of oxidative free radicals and nearly all oxidative DNA lesions are repaired via the DNA base excision repair (BER) pathway. To investigate the role of GSTM1, GSTT1 and BER genes (including OGG1, APE1, MUTYh, WRN and XRCC1) involved in OSCC risk in Taiwan, a comprehensive study on evaluation of polymorphisms of above genes was carried out in 1139 OSCC male cases and 1470 general males. A significant effect of AQ chewing or AQ chewing combined with cigarette smoking or alcohol drinking on the risk of OSCC was observed. GSTT1 null genotype was significantly associated with an increased risk of OSCC (odds ratio (OR), 2.10; 95% confidence interval (CI), 1.64-2.68, p = 0.000000003). After adjustment for cigarette smoking, AQ chewing and alcohol drinking, GSTT1 null-type variant was still significantly associated with an increased risk of OSCC (OR= 2.00; 95% CI, 1.42-2.81, p = 0.00007), while GSTM1 was not associated with OSCC risk. Polymorphisms of OGG1 Ser326Cys, APE1 Asp148Glu, APE1 T-656G, MUTYh Gln324His, WRN Cys1367Arg, and XRCC1 Arg399Gln were not associated with OSCC risk in general. However, stratification analysis indicated that polymorphisms of APE1 Asp148Glu and MUTYh Gln324His were associated with OSCC risk in individuals with cigarette smoking combined with AQ chewing. Furthermore, those individuals with either APE1 148 Asp/Asp or MUTYh 324 His/His genotype had an increased OSCC risks when compared to those with APE1 148 Glu and MUTYh 324 Gln allele (OR = 1.48; 95% CI, 1.13-1.95, p = 0.005). In conclusion, our results demonstrated that polymorphisms of genes involved in detoxification and BER of oxidative stress could influence the OSCC risk especially for those have history of cigarette smoking and AQ chewing. Citation Format: Chih-Hsiung Lai, Li-Ling Chiu, Huei-Tzu Chien, Saou-Hsing Liou, Shiang-Fu Huang, I-How Chen, Chun-Ta Liao, Ling-Ling Hsieh. Polymorphisms of GSTT1, APE1 and MUTYh gene and the risk of oral squamous cell carcinoma. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 4146. doi:10.1158/1538-7445.AM2014-4146

Abstract 2009: Clinical value of pathogenic mitochondrial mutations in oral cavity squamous cell carcinoma.

Proceedings: AACR 104th Annual Meeting 2013; Apr 6-10, 2013; Washington, DC Mitochondria appear to have a central role in two of the basic cancer cellular hallmarks made by Hanahan and Weinberg: reprogramming of energy metabolism and glucose-shortage induced apoptosis. Somatic mutations in mitochondrial DNA (mtDNA) have been frequently observed in human cancers and proposed as important oncological biomarkers. However, the clinical significance of mtDNA mutations in cancer remains unclear. In the present study, we screened the entire mitochondrial genome of 300 oral cavity squamous cell carcinomas (OSCC) with their matched control tissues by direct sequencing. We found that 240 (80%) OSCC tumor tissues had one or more somatic mutations and 85 % (203/240) of somatic mutation occurs in the control region (D-loop region) of mitochondrial genome, 47.1% (113/240) in the coding region, 13.8% (33/240) in rRNA and 8.3% (20/240) cases with in tRNA. Overall, 645 somatic mutations at 329 unique nucleotide positions were identified and 355 (55.0%) of 645 somatic mutations were located in the D-loop. The relative mutation frequency (mutations/nucleotides) in the D-loop was 15.6-fold higher than in the other regions (355/1121 versus 290/15448). The highest mutation was in the D310 polycytidine stretch region, where 165 (25.6%, 165/645) mutations were found. Among the 10 somatic mtDNA hot mutation sites, 6 are known mono-/di-nucleotide repeat length polymorphic sites, namely nucleotide position 310-317 (D310, also known as hypervariable region 1), 514-523 (CA repeat), 568-576 (C stretch 1), 956-965 (12S RNA), 5895-5899 (C stretch 2) and 16180-16195 (hypervariable region 2. We found that the mutation frequency at these 6 mono-/di-nucleotide repeat length polymorphic sites was strongly associated with the number of the repeat length and status of heteroplasmy. In the total 645 mutations, most common mutation type was base substitution (58.3%, 376/645), and transition mutation (G:C > A:T and A:T > G:C) accounted for the most mutations (94.7%, 356/376). The total and pathogenic mutation load for mitochondrial respiratory complexes was 1.10% and 0.76%, respectively. No significant association was found between somatic mtDNA mutations in whole mitochondrial genome and the clinicopathological features. However, we found that individuals with p53 R allele had higher frequency of pathogenic somatic mtDNA mutation than those with PP genotype. The Kaplan-Meier plot analysis indicated that pathogenic somatic mtDNA mutations was associated with poor disease-free survival (p = 0.02) and this phenomenon still existed after adjusting for tumor stage, differentiation and age at diagnosis (hazard ratio = 1.557; 95% confidence interval = 1.058 - 2.292); while, there was no difference in overall survival. Taken together, these data suggest that pathogenic somatic mtDNA mutations may play important roles in the aggressiveness of OSCCs. Citation Format: Ling-Ling Hsieh, Chih-Hsiung Lai, Shiang-Fu Huang, Chun-Ta Liao, I-How Chen, Hung-Ming Wang. Clinical value of pathogenic mitochondrial mutations in oral cavity squamous cell carcinoma. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 2009. doi:10.1158/1538-7445.AM2013-2009