Jesper Reibel

Prognosis of Oral Pre-malignant Lesions: Significance of Clinical, Histopathological, and Molecular Biological Characteristics

The concept of a two-step process of cancer development in the oral mucosa, i.e., the initial presence of a precursor subsequently developing into cancer, is well-established. Oral leukoplakia is the best-known precursor lesion. The evidence that oral leukoplakias are pre-malignant is mainly derived from follow-up studies showing that between < 1 and 18% of oral pre-malignant lesions will develop into oral cancer; it has been shown that certain clinical sub-types of leukoplakia are at a higher risk for malignant transformation than others. The presence of epithelial dysplasia may be even more important in predicting malignant development than the clinical characteristics. Three major problems, however, are attached to the importance of epithelial dysplasia in predicting malignant development: (1) The diagnosis is essentially subjective, (2) it seems that not all lesions exhibiting dysplasia will eventually become malignant and some may even regress, and (3) carcinoma can develop from lesions in which epithelial dysplasia was not diagnosed in previous biopsies. There is, therefore, a substantial need to improve the histologic assessment of epithelial dysplasia or, since epithelial dysplasia does not seem to be invariably associated with or even a necessary prerequisite for malignant development, it may be necessary to develop other methods for predicting the malignant potential of pre-malignant lesions. As a consequence of these problems, numerous attempts have been made to relate biological characteristics to the malignant potential of leukoplakias. Molecular biological markers have been suggested to be of value in the diagnosis and prognostic evaluation of leukoplakias. Markers of epithelial differentiation and, more recently, genomic markers could potentially be good candidates for improving the prognostic evaluation of precursors of oral cancer. As yet, one or a panel of molecular markers has not been determined that allows for a prognostic prediction of oral pre-cancer which is any more reliable than dysplasia recording. However, these new markers could be considered complementary to conventional prognostic evaluation.

Oral epithelial dysplasia classification systems: predictive value, utility, weaknesses and scope for improvement

At a workshop coordinated by the WHO Collaborating Centre for Oral Cancer and Precancer in the United Kingdom issues related to potentially malignant disorders of the oral cavity were discussed by an expert group. The consensus views of the Working Group are presented in a series of papers. In this report, we review the oral epithelial dysplasia classification systems. The three classification schemes [oral epithelial dysplasia scoring system, squamous intraepithelial neoplasia and Ljubljana classification] were presented and the Working Group recommended epithelial dysplasia grading for routine use. Although most oral pathologists possibly recognize and accept the criteria for grading epithelial dysplasia, firstly based on architectural features and then of cytology, there is great variability in their interpretation of the presence, degree and significance of the individual criteria. Several studies have shown great interexaminer and intraexaminer variability in the assessment of the presence or absence and the grade of oral epithelial dysplasia. The Working Group considered the two class classification (no/questionable/ mild - low risk; moderate or severe - implying high risk) and was of the view that reducing the number of choices from 3 to 2 may increase the likelihood of agreement between pathologists. The utility of this need to be tested in future studies. The variables that are likely to affect oral epithelial dysplasia scoring were discussed and are outlined here; these need to be researched in longitudinal studies to explore the biological significance of a low-risk or high-risk dysplasia.

Xerostomia and hypofunction of the salivary glands in cancer therapy.

Abstract. This review presents data from the literature on oral adverse reactions from the perspectives of subjective feelings of dry mouth (xerostomia) and objective measures of salivary gland hypofunction during and after cancer therapy. Special emphasis is paid to the mechanisms behind xerostomia, impaired saliva secretion and changes in the composition of saliva and to how these relate to radiation therapy involving the salivary glands and to systemic chemotherapy. The oral complications that relate to such iatrogenic changes in salivary gland function are also discussed.

Tobacco and Oral Diseases

It is well known that smoking contributes to the development of lung cancer and cardiovascular disease, and there is weighty evidence that it has a considerable influence on oral health. Smoking has many negative effects on the mouth, including staining of teeth and dental restorations, reduction of the ability to smell and taste, and the development of oral diseases such as smoker’s palate, smoker’s melanosis, coated tongue, and, possibly, oral candidosis and dental caries, periodontal disease, implant failure, oral precancer and cancer. From a qualitative point of view the latter is obviously the most serious tobacco-related effect in the mouth. Quantitatively, however, importance has been attached to periodontitis, which affects a large proportion of the population, and during recent years more attention has been given to implant survival rates. Dentists have an important role to play in preventing the harmful effects of smoking in the mouth, and consequently smoking counselling should be as much a part of the dentist’s job as plaque control and dietary advice.

Human papillomavirus in oral premalignant lesions

The aetiology of oral premalignant lesions is generally accepted to be multifactorial. Tobacco and alcohol are established as important cofactors in malignant development in the oral cavity, but in addition microorganisms, such as human papillomavirus (HPV), have gained much interest over the past decade. For many years, HPV has been accepted as an important cofactor in the development of cervical cancer, originating from a mucous membrane with similarities to the oral mucosa. 49 patients with oral premalignant lesions and 20 control patients with normal oral mucosa and no history of HPV infection were examined for the presence of HPV by immune histochemical staining using the peroxidase anti-peroxidase technique (PAP), DNA-DNA in situ hybridisation (ISH), and polymerase chain reaction (PCR) analysed by Southern blot hybridisation with an HPV 16 specific probe. The investigations revealed that HPV was found in 62.5% of the verrucous leucoplakias, 50.0% of the erythroplakias, 45.5% of the homogeneous leucoplakias, 33.3% of erythroleucoplakias and in 12.5% of the nodular leucoplakias. An overall HPV detection rate in the examined premalignant lesions was 40.8% and no patients in the control sample were positive. Concerning oral cancer development, it seems likely that HPV may be a cofactor, as 100% of patients who developed oral cancers within 4-12 years were all positive for HPV, one being positive for HPV 16.

Genetic and epigenetic alterations of the blood group ABO gene in oral squamous cell carcinoma

Loss of histo‐blood group A and B antigen expression is a frequent event in oral carcinomas and is associated with decreased activity of glycosyltransferases encoded by the ABO gene. We examined 30 oral squamous cell carcinomas for expression of A and B antigens and glycosyltransferases. We also examined DNA from these tumors for loss of heterozygosity (LOH) at markers surrounding the ABO locus at chromosome 9q34, for loss of specific ABO alleles, and for hypermethylation of the ABO promoters. Loss of A or B antigen expression was found in 21 of 25 tumors (84%) and was a consistent feature of tumors lacking expression of A/B glycosyltransferases. LOH at 9q34 was found in 7 of 27 cases (26%), and one case showed microsatellite instability. Among 20 AO/BO cases, 3 showed loss of the A/B allele and 3 showed loss of the O allele. Analysis of the proximal ABO promoter by methylation‐specific PCR and melting curve analysis showed hypermethylation in 10 of 30 tumors (33.3%), which was associated with loss of A/B antigen expression. ABO promoter hypermethylation was also found in hyperplastic or dysplastic tissues adjacent to the tumors, suggesting that it is an early event in tumorigenesis. Collectively, we have identified molecular events that may account for loss of A/B antigen expression in 67% of oral squamous cell carcinomas.

Oral mucosal lesions, microbial changes, and taste disturbances induced by adjuvant chemotherapy in breast cancer patients

OBJECTIVE The aim of the study was to examine oral mucosal lesions, microbial changes, and taste disturbances induced by adjuvant chemotherapy (CT) in breast cancer patients during and 1 year after treatment. STUDY DESIGN Forty-five consecutive breast cancer patients, eligible for adjuvant CT with cyclophosphamide, epirubicin or methotrexate, and 5-fluorouracil were followed before, during, 6 months and 1 year after CT and were compared to a control group of 31 breast cancer patients not receiving adjuvant CT. RESULTS During CT, oral mucosal lesions developed including erythema (n = 10, 22%) and ulceration (n = 7, 16%). Five patients (11%) were diagnosed with oral candidosis. Scores of dental bacterial plaque and gingival inflammation increased during CT and the oral microbial composition changed towards a more acidophilic flora. Taste disturbances were experienced by 84% (n = 38) of the patients in the CT group. CONCLUSION In breast cancer patients, moderate-intensive adjuvant CT caused oral mucosal lesions, oral candidosis, taste disturbances and a more acidophilic oral microflora. These adverse effects were temporary and the majority of the patients were mildly affected.

Comparison of nonexposed and exposed bisphosphonate-induced osteonecrosis of the jaws: a retrospective analysis from the Copenhagen cohort and a proposal for an updated classification system.

OBJECTIVE Nonexposed osteonecrosis of the jaws (NE-ONJ) does not fit into the current definition of osteonecrosis, which requires exposed bone. A modification of the classification of bisphosphonate-induced osteonecrosis of the jaws (ONJ) is proposed. This study aimed to test proposed criteria for NE-ONJ and compare NE-ONJ with exposed ONJ (E-ONJ) in a retrospective analysis. STUDY DESIGN In 102 patients with E-ONJ diagnosed according to Ruggiero et al. (2006, 2009), criteria for NE-ONJ were developed. Subgroups of NE-ONJ and E-ONJ were tested against each other using nonparametric and parametric statistics. RESULTS Among 102 patients with ONJ, 14 had NE-ONJ and 88 had E-ONJ. NE-ONJ and E-ONJ were similar in all important data (P > .05) except bone exposure. CONCLUSIONS NE-ONJ belongs to the same disease condition as E-ONJ. NE-ONJ may be otherwise classified as ONJ stage 1, 2, or 3 and is different from ONJ stage 0. We propose to include the criteria for NE-ONJ into the classification.

Differential expression of integrins and laminin-5 in normal oral epithelia

β1 and β4 integrins are receptors on epithelial cells mediating cell‐extracellular matrix adhesion. Furthermore, α2β1 and α3β1 contribute to cell‐cell adhesion. Laminin‐5 in epithelial basement membranes (BMs) is a ligand for α6β4 and α3β1. Expression of different integrins and laminin‐5 was studied in oral epithelium to characterize regional variations in these adhesion molecules. Monoclonal antibodies directed against α2‐α6β1/α6β4 and laminin‐5 were examined in cryopreserved biopsies of normal mucosa by immunohistochemistry. Laminin‐5 was expressed as a line along the BMs. The junctional epithelium showed a unique phenotype: Laminin‐5 was detected in the internal BM at the tooth surface and in the external BM, where excessive laminin‐5 was seen in the stroma. α6β4 was expressed in all cells of the junctional epithelium. Integrins α4β1 and α5β1 were not detected in the epithelia, whereas α2β1 and α3β1 showed differential expression. Epithelia with well‐developed rete pegs and connective tissue papillae showed polarized α3β1 expression along the BM in the rete pegs, in contrast to negative expression at the tips of the connective tissue papillae. A variation in the suprabasal distribution of α2β1 and α3β1 was observed between epithelia from different regions. α2β1 and α3β1 were detected in basal/parabasal cells in keratinized epithelia, whereas there was increased suprabasal expression in nonkeratinized mucosa. These results indicate inhomogeneity in the basal cell population of oral squamous epithelia and differential expression of integrins, which may reflect differences in the underlying stroma. Laminin‐5 deposits in the stroma underneath the junctional epithelium may indicate subclinical gingival inflammation.

Cancer‐associated changes in glycosylation of fibronectin

The extracellular matrix adhesion molecule fibronectin exhibits different isoforms derived by alternative splicing as well as recently demonstrated variation in O‐glycosylation. Although fibronectin is widely distributed in normal tissues, the individual isoforms have been found to show restricted tissue distribution and association with malignancies. The monoclonal antibody FDC‐6 defines a cancer‐associated de novo glycosylation of a specific threonine residue in the C‐terminal region of the fibronectin molecule termed oncofetal fibronectin. Here we report an immunohistological study of oral squamous cell carcinomas (n = 33), premalignant lesions (n = 15), and normal oral mucosa (n = 10) using the FDC‐6 antibody. A selective expression of the oncofetal fibronectin epitope was demonstrated in close relation to the invading carcinoma, whereas no staining was observed in premalignant lesions without epithelial dysplasia, or in normal epithelium. Furthermore, we attempted to identify additional carbohydrate‐related epitopes distinguishing fibronectin of human hepatoma cell line HUH‐7 from plasma fibronectin. No novel epitopes were identified, as all generated monoclonal antibodies lacking reactivity with plasma fibronectin showed the same specificity as FDC‐6. Previous studies have indicated that the de novo glycosylation is induced by a novel transferase activity only found in fetal and carcinoma cell lines, placenta and hepatoma tissues. Here we provide further evidence that a purified UDP‐GalNAc:peptide N‐acetylgalactosaminyltransferase from normal bovine thymus and human placentae is incapable of utilizing the hexapeptide VTHPGY as a substrate. The results demonstrate that oncofetal fibronectin is highly associated with malignancy, and appears to be induced by expression of a unique glycosyltransferase or modification of the specificity of the normally expressed transferase.