Davide Bartolomeo Gissi

Multiple squamous cell carcinomas of the oral cavity in a young patient with graft-versus-host disease following allogenic bone marrow transplantation

The development of secondary malignancies is a potential long-term complication after haematopoietic stem cell transplantation (HSCT). In particular, a higher incidence of oral squamous cell carcinoma (OSCC) has been reported in patients experiencing chronic graft versus host disease (cGvHD) secondary to HSCT. This report describes the development of two synchronous SCC of the buccal mucosa in a young female patient treated with HSCT for beta thalassemia major. She had undergone HSCT at the age of 9 years and developed oral GvHD 6 months after transplant. 17 years after HSCT she developed two synchronous carcinomatous lesions on the tongue and floor of the mouth. The current case highlights the association between oral cGvHD and OSCC, and the possible development of OSCC in young patients even many years after HSCT. This evidence suggests closer follow-up for all patients treated with HSCT who developed cGvHD, and more effective strategies to prevent and treat cGvHD.

Cancerization of cutaneous flap reconstruction for oral squamous cell carcinoma: report of three cases studied with the mtDNA D-loop sequence analysis

Foschini M P, Morandi L, Marchetti C, Cocchi R, Eusebi L H, Farnedi A, Badiali G, Gissi D B, Pennesi M G & Montebugnoli L
(2011) Histopathology58, 361–367
Cancerization of cutaneous flap reconstruction for oral squamous cell carcinoma: report of three cases studied with the mtDNA D‐loop sequence analysis

Predictive Role of p53 Protein as a Single Marker or Associated to Ki67 Antigen in Oral Carcinogenesis

p53 over-expression has been proposed as a reliable marker associated to oral carcinogenesis, although only about 50% of oral carcinomas (OSCC) are associated with p53 over-expression and even p53-negative lesions can progress to OSCC. The aim of the study was to determine whether the combination of p53 over-expression and p53 low-expression associated with Ki67 over-expression (high Ki67/p53 ratio) could lead to a more sensitive parameter. Immunohistochemical expression of Ki67 and p53 was measured in 54 specimens from OSCC; 27 specimens from moderate/severe epithelial dysplasia; 32 specimens from oral leukoplakias without epithelial dysplasia, and 13 specimens with normal epithelium. p53 over-expression was found in 31 (53%) samples from OSCC, in 10 (37%) samples from severe dysplasias, and in 5 (15%) samples from non-dysplastic lesions, while the combination of high p53 values with high Ki67/p53 ratio was observed in 93% of OSCC, in 81% of dysplastic lesions, and in 50% of non-dysplastic lesions. This parameter may have a clinical implication to detect early lesions with an impairment of p53 pathway, and probably at risk of progress to OSCC.

Immunohistochemical expression of p16INK4A protein in oral lichen planus

The expression of p16(INK4A) has been investigated in oral leukoplakias (OLK), but no data are available about oral lichen planus (OLP). In this study, p16(INK4A) immunohistochemical expression was evaluated in 56 OLP and 36 OLK (12 without inflammation [NI-OLK] and 24 with chronic inflammation [I-OLK]) and compared with 23 reactive nonspecific inflammations (INF) and 14 normal control samples. The p16(INK4A) immunostaining was considered to be positive when >5% of keratinocytes were stained. All normal control samples were negative. Positive p16(INK4A) was detected in OLP, IOLK, and INF. Significant differences in p16(INK4A) positivity were found between OLP (64%) and OLK (28%) (χ(2) = 17.7; P < .01), and between I-OLK and NI-OLK (χ(2) = 4.5; P < .05). No significant difference was found between OLP and INF (43%). In conclusion, positive p16(INK4A) in OLP patients seems to be related to reactive inflammatory processes rather than to a risk of progression to oral squamous cell carcinoma.

DNA methylation analysis by bisulfite next-generation sequencing for early detection of oral squamous cell carcinoma and high-grade squamous intraepithelial lesion from oral brushing.

PURPOSE Oral squamous cell carcinoma (OSCC) is commonly preceded by oral potentially malignant lesions (OPML). The aim of the present study was to assess, by bisulfite next-generation sequencing (NGS), the methylation status of a list of candidate genes obtained from oral brushings to early detect OPML and OSCC. MATERIAL AND METHODS Oral brushing specimens from 11 OSCC, 11 high-grade squamous intraepithelial lesions (HG-SIL), 9 low-grade SIL (LG-SIL), 9 oral lichen planus (OLP), and 8 healthy donors were included in this study. We investigated, by means of bisulfite NGS, the promoter of GP1BB, ZAP70, KIF1A, p16[CDKN2A], CDH1, miR137, and miR375. Statistical significance between lesions and a pool of healthy donors were evaluated with the Mann-Whitney U test. RESULTS ZAP70 was found to be hypermethylated in 100% of OSCC and HG-SIL and in 28.6% of LG-SIL. GP1BB hypomethylation was detected in 90.9% OSCC and HG-SIL and in 37.5% of LG-SIL. MiR137 was hypermethylated in 100% of OLP, 44.4% of OSCC, 40% HG-SIL, and 25% LG-SIL. KIF1A hypermethylation was found to be associated with TP53 mutations (p < 0.0001). CONCLUSION In the present preliminary cohort of patients, DNA methylation analysis of GP1BB and ZAP70 seems to be a promising noninvasive tool for early detection of OSCC and HG SIL from oral brushing specimens.

Predictive Role of p53 Protein as a Single Marker or Associated with ki67 Antigen in Oral Leukoplakia: A Retrospective Longitudinal Study

Oral leukoplakia (OL) is the most common potentially malignant lesion of the oral cavity. Immunohistochemical analysis of p53 and Ki67 proteins is a simple and inexpensive method widely used in non-dysplastic OLs to reveal lesions predicted to develop oral cancer. The present longitudinal study evaluated the predictive role of p53 and Ki67 proteins alone or in combination in a group of OLs without dysplasia followed for many years. Seventy-seven OL patients referred to our Department between January 2006 and October 2013 underwent histochemical analysis of p53 and Ki67 expression. OLs were considered at high risk in the presence of either high p53 expression (>20%), or low/normal p53 expression associated with high Ki67 expression (Ki67/p53 ratio >3). Seven OLs evolved to OSCC during the follow-up period. Three cases had p53 overexpression, while four had a high Ki67/p53 ratio. Statistical significance was reached when samples with p53 overexpression were combined with samples with high Ki67/p53 ratio (Chi square 5.3; p<0.02). The combined immunohistochemical expression of p53 and Ki67 proteins could be a useful and simple molecular marker for early detection of non-dysplastic OLs at risk of developing oral cancer.

Clonality analysis in primary oral squamous cell carcinoma and related lymph-node metastasis revealed by TP53 and mitochondrial DNA next generation sequencing analysis.

The chance of developing a neck nodal metastasis after initial treatment of oral squamous cell carcinoma varies from 12.4% to 62%. Despite being the main reason for cancer-related mortality, nodal metastases are still rarely subjected to molecular analyses, and our knowledge of the clonal heterogeneity of multiple lesions within the same patient is limited. The aim of the present study was to evaluate the relationship between primary oral cancer and lymph node metastasis in a series of patients with synchronous and metachronous metastases by 2 clonality tests: mt-DNA and TP53 sequence analysis. The study population consisted of 10 consecutive patients. Data identified in this study demonstrate that our assay based on next-generation analysis of TP53 and mt-DNA is simple, is reliable, allows high throughput, and may be applied to retrospective cases. The combination of mt-DNA and TP53 data analysis helped us to evaluate more precisely and consistently the genetic relationship among different tumor clones.

Genetic relationship between multiple squamous cell carcinomas arising in the oral cavity.

Histological and clinical criteria are generally used to differentiate second primary tumors (SPTs) from local recurrences. The purpose of the present study was to apply mitochondrial DNA (mtDNA) D‐loop analysis to differentiate SPTs from local recurrences and to validate the clinical classification.

Clinical and histologic healing of lichenoid oral lesions following amalgam removal: a prospective study

OBJECTIVE This study aimed to see whether clinical healing after amalgam removal corresponds to histologic healing, i.e., a complete disappearance of any histologic sign of lichenoid lesion. STUDY DESIGN The study evaluated 64 patients with lichenoid lesions and at least one amalgam filling. RESULTS After amalgam removal, complete clinical healing was obtained in 14 patients (22%) and was significantly related to lesion topography (χ(2) 4.7; P < .05) and positive patch test (χ(2) 6.3; P < .01). Complete histologic healing was obtained in only 7 cases (50% of clinically healed patients), and was significantly related to the combination of positive patch test and strict contact with amalgams (Fishers exact test P < .01). CONCLUSIONS Contact with amalgams and positive patch testing are good but not absolute indicators of the beneficial effect of amalgam replacement. In addition, complete clinical healing does not necessarily mean a disappearance of the histologic characteristics of OLL/OLP lesions.

Histological and immunohistochemical evaluation of new epithelium after removal of oral leukoplakia with Nd:YAG laser treatment

Laser excision has been used with increasing success to treat oral leukoplakia. The aim of the present study was to evaluate whether clinical healing of a leukoplakia after laser surgery is associated with a normal functional status of the new epithelium and whether pathological alterations in these parameters are related to the risk of local recurrence. The study population consisted of 13 consecutive patients with oral leukoplakia in which clinical healing was achieved after laser therapy using an Nd:YAG laser (Model 6000, Laser Sonics, Cooper Laser Sonics). At the end of the therapy, all patients underwent a second biopsy of the clinically healthy tissue. Epithelial cell turnover was evaluated before and after laser surgery by Ki67 protein expression, and positive staining of more than 20% was considered abnormal. All patients were followed on a monthly basis. Eight patients had abnormally high Ki67 values before laser therapy (mean 27.4 ± 6.2%), but the levels decreased significantly after treatment (17.6 ± 8.5%; t = 2.6, p < 0.05). High Ki67 values persisted in three patients, and local recurrences in the new epithelium was observed in two of these patients. Kaplan–Meier statistics showed that the between-group difference was statistically significant (Chi square 7.3; p < 0.01). In conclusion, this is the first prospective study to show that clinical healing of leukoplakia treated by laser surgery may be accompanied by altered cell turnover in 20% of the cases. Ki67, as a marker of proliferative status, may be a prognostic indicator in the mucosa replacing the lesion.