Patients affected by squamous cell carcinoma of the head and neck (HNSCC) show frequent occurrence of multiple cancers and widespread precancerous lesions in the mucosa of the upper respiratory tract, a phenomenon known as field cancerization. In this study, we investigated the role of genetic instability in the development of HNSCC and in particular in tumour multiplicity phenomena of the upper respiratory tract. For this purpose, we analysed microsatellite instability (MI) and loss of heterozygosity (LOH) at 20 loci mapping on five chromosomal arms in 67 HNSCC patients, 45 of whom had a single cancer and 22 had multiple primary tumours. The possible involvement of the hMLH1 gene in genetic instability and as a potential target of 3p21 deletion phenomena in head and neck cancers was also investigated. Our data indicate that mismatch repair-related genetic instability plays a minor role in the carcinogenesis of HNSCC and in tumour multiplicity of the head and neck region. Moreover, our results exclude a role for the hMLH1 gene as a determinant of MI and as a specific gene target of deletion at 3p21 in HNSCC. We conclude that presumably other genetic mechanisms, such as those hypothesized for MI-negative hereditary non-polyposis colorectal cancer patients, may play a major role in the carcinogenesis of the mucosa of the upper respiratory tract.