Expression of facilitative glucose transport proteins during development of squamous cell carcinomas of the head and neck.

Positron emission tomography studies on malignant head and neck tumors have shown that tumor growth and elevated glucose uptake are associated. On a molecular level, glucose uptake is mediated by specific glucose transport proteins, which exhibit an altered expression in head and neck malignant neoplasms. However, it is unknown when during development of squamous cell carcinomas an alteration of the expression of glucose transport proteins occurs. We have studied the expression of different facilitating glucose transport proteins (GLUT 1, 2, 3 and 4) by immunohistochemistry in a variety of preneoplastic and neoplastic mucosal lesions of the head and neck. We have observed weak expression of GLUT 1 in normal mucosa, a marked expression of GLUT 1 throughout preneoplastic lesions, which correlated well with the degree of dysplasia. In squamous cell carcinomas of the head and neck (HNSCC) and metastases, GLUT 1 was always expressed strongly. In contrast, GLUT 2, 3 and 4 were not detected in any of the epithelial tissues examined. The increased expression of GLUT 1 in dysplastic lesions and its sustained expression in SCC indicate that changes of GLUT 1 expression are early events during development of HNSCC. Therefore, the detection of GLUT 1 might be a reliable marker in the diagnosis of premalignant lesions of the oropharyngeal mucosa.

Aberrant p21(CIP1/WAF1) protein accumulation in head-and-neck cancer.

p21(CIP1/WAF1) is an inhibitor of cyclin-dependent kinases and, in normal tissues including squamous epithelia, has been associated with cell-cycle exit and differentiation. As shown in this pilot study, however, the majority of head-and-neck squamous-cell carcinomas (HNSCC) display aberrant p21(CIP1/WAF1) expression: of 42 tumors analyzed by immunohistochemical staining, 28 (67%) over-expressed the p21(CIP1/WAF1) protein. Accumulation of p21(CIP1/WAF1) was independent of the histological grade of the tumors as well as the genetic status of the p53 gene. In many cases, most notably in poorly differentiated or undifferentiated HNSCC, p21(CIP1/WAF1)-positive cells were actively proliferating tumor cells, since they also expressed proliferating-cell nuclear antigen (PCNA) and Ki-67. Accumulation of p21(CIP1/WAF1) occurred through a post-transcriptional mechanism since, in contrast to immunohistochemical analysis of the p21(CIP1/WAF1) protein, in situ hybridization showed no increase of mRNA levels as compared with cells in normal mucosa (n = 25). Clinically, among the patients with p21(CIP1/WAF1)-over-expressing tumors, there was increased recurring disease (p = 0.03; chi2-test), shortened disease-free survival (p = 0.0019; log-rank test) and shortened overall survival (p = 0.0071; log-rank test). These in vivo data indicate that in many HNSCC, accumulated p21(CIP1/WAF1) is compatible with increased tumor-cell proliferation, and they provide preliminary evidence that p21(CIP1/WAF1) may be of prognostic and predictive significance.

Expression of CD44 splice variants in squamous epithelia and squamous cell carcinomas of the head and neck.

Splice variants of the adhesion molecule CD44 have been described as essential for the lymphatic spread of rat tumour cells and are claimed to be involved in the metastatic spread of several human tumours. Immunohistochemistry has been used to analyse the expression pattern of CD44 standard (CD44s) and variant (CD44v) isoforms in normal and dysplastic squamous epithelia, as well as in primary and metastatic squamous cell carcinomas (SCCs), which spread predominantly by way of the lymphatic system. Frozen sections of squamous epithelia and of squamous cell carcinomas were stained with a panel of monoclonal antibodies recognizing epitopes of CD44s as well as of the variant exons v5, v6, v7, v7-v8, and v10. The stratum basale and stratum suprabasale of squamous epithelia stained with all antibodies; the stratum spinosum stained with anti-CD44v5, anti-CD44v6, anti-CD44v7-8 and anti-CD44v10; the lower layers of the stratum corneum stained with anti-CD44v5. This expression profile was seen in epithelia of the lip, the tongue, the gingiva, the hard palate, the floor of the mouth, the buccal mucosa, and the pharynx. The same pattern of expression was also noted in dysplastic epithelia, but expression of the variant exons v7, v8, and v10 was significantly downregulated in primary squamous cell carcinomas and was not detected at all in the majority of metastasis-derived specimens. Expression of CD44v5 and CD44v6, on the other hand, was mainly unaltered. Thus, epithelial cell layers representing different stages of differentiation express distinct sets of CD44 variant isoforms, where especially exons v8-v10 might be required for the maintenance of the structural integrity of squamous epithelium. Downregulation of these exons on tumour cells could indicate that they are irrelevant for tumour progression or may even hamper infiltration of surrounding tissue or of lymphatics.

Expression of the histone H3 gene in benign, semi-malignant and malignant lesions of the head and neck: a reliable proliferation marker.

To search for a reliable proliferation marker in epithelial head and neck lesions, we have analysed the expression of the histone H3 gene by in situ hybridisation and compared this with the immunoreactivity of the widely used monoclonal antibody Ki-67. In many lesions, the Ki-67 staining failed to delineate proliferation. In contrast, the H3 hybridisation signals were in accordance with the histopathology of the biopsies: in hyperplastic epithelia, significant H3 mRNA levels were only seen in areas with inflammation. Dysplastic cells showed distinctly elevated H3 expression. Benign and semi-malignant tumours, i.e. basal cell carcinomas, showed moderate H3 signals at the periphery. In squamous cell carcinomas, H3 expression was always high at the expanding zone of the tumour and was most extensive in undifferentiated carcinomas. Thus, the expression of the histone H3 gene closely reflected the dynamics of neoplastic growth within and around head and neck tumours.