Atsuko Hirai

Multiple oral squamous epithelial lesions: are they genetically related?

The development of second primary tumors (SPTs) in patients with head and neck squamous cell carcinoma (HNSCC) has become an increasingly important factor in clinical treatment decison. Currently, clinical and histologic parameters are used to determine whether or not SPT is present. Recent studies suggest that many SPTs in the upper aerodigestive tract have a common clonal origin, challenging the longstanding multiclonal origin concept. To determine genetic relationships among multiple oral cancerous and precancerous lesions (MOCP), we analysed 100 lesions from 26 Japanese patients. Lesion development was synchronous and metachronous. We looked for patterns of microsatellite alterations (MA) using seven markers at chromosomes 3p14, 9p21, and 17p13, where MA occurs early in oral carcinogenesis. Loss of heterozygosity (LOH) was found in 52.6% (41/78), 62.5% (60/96), and 59.3% (32/54) of informative MOCP at 3p14, 9p21, and 17p13, respectively. Microsatellite instability (MI) was observed in 11, 26 and 13% of the samples at 3p14, 9p21, and 17p13 markers, respectively. Patterns of MA were concordant in only nine (14%) of 63 lesions from four (18%) of 22 patients who initially presented with noninvasive lesions. However, two of four patients with invasive cancer as indexed lesion showed 16 (43%) clonally related MOCP among 37 lesions (P=0.003). The results suggest that the majority of MOCP arise from clonally independent cells affected by field cancerization. However, the probability of mucosal spread of clonal malignant or premalignant cells may increase along with malignant progression.

Specific p53 mutations predict poor prognosis in oral squamous cell carcinoma

In this study, we focused on p53 mutations in specific regions, including DNA-binding surface regions, to clarify the correlation between mutations within the specific regions of p53 and clinical outcomes of patients with oral cancers. We analyzed p53 mutations in 121 fresh primary oral squamous cell carcinomas (SCCs) by polymerase chain reaction-single-strand conformation polymorphism or a yeast functional assay. p53 mutations were detected in 51/121 (42%) cases. Mutation of p53 was not associated with any clinicopathological parameters; however, tumors containing specific p53 mutations, e.g. DNA-binding surface regions (L2, L3 and the LSH motif) and conserved regions (II-V), had significantly poorer prognoses than tumors with mutations outside of those regions. Moreover, locoregional failure, lymph node metastasis and the occurrence of subsequent distant metastasis were also significantly associated with mutations within DNA-binding surface regions. These data indicate that specific mutations of p53 could be important prognostic factors in oral SCCs.

Clinical value of genetically diagnosed lymph node micrometastasis for patients with oral squamous cell carcinoma.

The purpose of this study was to investigate the incidence and clinical significance of genetically diagnosed lymph node micrometastasis for patients with oral squamous cell carcinoma (SCC).

Interaction of MCC2, a novel homologue of MCC tumor suppressor, with PDZ-domain Protein AIE-75

AIE-75 is a protein identified as an autoantigen in patients with autoimmune enteropathy and as a colon cancer-related antigen. It has recently been assigned to be a causative gene for Usher type 1C congenital syndromic hearing loss. The novel protein has three PSD-95/Dlg/ZO-1 (PDZ) protein-protein interaction domains and is therefore implicated to function as a molecular anchor or sorter. We have identified a novel protein that binds to AIE-75 by yeast two-hybrid screening. The protein has a high homology to the tumor suppressor MCC (mutated in colon cancer; or MCC1 hereafter) and was named MCC2. MCC2 protein binds the first PDZ domain of AIE-75 with its C-terminal amino acids -DTFL. Since the MCC1 does not bind to AIE-75 and the MCC2 displays different expression patterns in various organs compared to MCC1, they appear to play distinct roles in cells. The MCC2 gene is located on chromosome 19p13 in the vicinity of APCL gene, while MCC1 maps near to APC tumor suppressor gene. Because of negative expression of MCC2 in a panel of cancer cell-lines compared to the corresponding normal tissues, we suggest that further study is necessary to investigate a possible role of MCC2 as a tumor suppressor.

Radioresistance in Oral Squamous Cell Carcinoma With p 53 DNA Contact Mutation

Reliable variables to predict the radiosensitivity of each tumor have not been identified. Recent studies have demonstrated that specific regions of mutations within the core domain of p53 protein correlate with responses to chemotherapy and radiotherapy in some tumor types. In this study, we evaluated the relationship between specific p 53 mutations and radiosensitivity in 49 patients with oral squamous cell carcinomas (SCCs) who underwent preoperative radiotherapy. Exons 5 through 8 of the p 53 gene were examined by polymerase chain reaction-single-strand conformation polymorphism and direct sequencing. We detected p 53 mutations in 27 (55.1%) cases. DNA contact mutations were detected in 11 (40.7%) of these 27 cases in L3 loop, loop-sheet-helix motif, and zinc-binding residues. Tumors containing p 53 DNA contact mutations had significantly poorer responses to radiation than the other tumors, although no statistically significant difference between tumors with and without p 53 mutations was found. These data indicate that DNA contact mutation of p 53 could be a useful marker to predict the radioresistance of oral SCCs.

A functional and quantitative mutational analysis of p53 mutations in yeast indicates strand biases and different roles of mutations in DMBA- and BBN-induced tumors in rats

In order to analyze the mutational events and to understand the biological significance of the p53 gene in chemical carcinogenesis, we applied a new yeast‐based p53 functional assay to ovarian tumors induced by 7, 12‐dimethylbenz[a]anthracene (DMBA), as well as to transitional cell carcinomas of the urinary bladder induced by N‐butyl‐N‐(4‐hydroxybutyl) nitrosamine (BBN) in rats. The assay demonstrated that 15 of 19 DMBA induced tumors harbored clonal p53 mutations, which is consistent with the expectations of the “clonal expansion” hypothesis. The majority of the mutations were purine (AG) to pyrimidine (CT) transversions (12/19) on the non‐transcribed (sense) strand (NTS), which is likely to be due to depurination created by DMBA adduct formation on the NTS. In contrast, we found no purine to pyrimidine transversion on the NTS. After cessation of BBN treatment, BBN‐induced multifocal lesions in the bladder contained heterogeneous p53 mutations at an early stage. In the later stage, however, clonal p53 mutations were identified in 4 out of 7 bladders analyzed, conforming with the concept of “field cancerization”. The observed base substitutions were G→A (1/6) or C →T transitions (2/6), and mutations at T (3/6) on the NTS in clonal mutations, together with non‐clonal mutations, showing a preference of C→T to G→A (17 vs. 0). Thus, preferential repair was found in the transcribed strand of the p53 gene, whether modified by DMBA or by BBN carcinogens. Very similar mutation patterns were observed between clonal and non‐clonal mutations in the DMBA‐ and BBN‐induced tumors, indicating that the rat yeast p53 functional assay can be a potential tool for the characterization of in vivo mutation patterns of p53, when modified by chemical carcinogens. Int. J. Cancer 83:700‐705, 1999.