Oral epithelial dysplasia detection and grading in oral leukoplakia using deep learning.

BACKGROUND: The grading of oral epithelial dysplasia is often time-consuming for oral pathologists and the results are poorly reproducible between observers. In this study, we aimed to establish an objective, accurate and useful detection and grading system for oral epithelial dysplasia in the whole-slides of oral leukoplakia. METHODS: Four convolutional neural networks were compared using the image patches from 56 whole-slide of oral leukoplakia labeled by pathologists as the gold standard. Sequentially, feature detection models were trained, validated and tested with 1,000 image patches using the optimal network. Lastly, a comprehensive system named E-MOD-plus was established by combining feature detection models and a multiclass logistic model. RESULTS: EfficientNet-B0 was selected as the optimal network to build feature detection models. In the internal dataset of whole-slide images, the prediction accuracy of E-MOD-plus was 81.3% (95% confidence interval: 71.4-90.5%) and the area under the receiver operating characteristic curve was 0.793 (95% confidence interval: 0.650 to 0.925); in the external dataset of 229 tissue microarray images, the prediction accuracy was 86.5% (95% confidence interval: 82.4-90.0%) and the area under the receiver operating characteristic curve was 0.669 (95% confidence interval: 0.496 to 0.843). CONCLUSIONS: E-MOD-plus was objective and accurate in the detection of pathological features as well as the grading of oral epithelial dysplasia, and had potential to assist pathologists in clinical practice.

Proliferative verrucous leukoplakia: a general dental practitioner-focused clinical review.

Proliferative verrucous leukoplakia (PVL) is a distinct type of oral leukoplakia which has the potential to enlarge or develop into new areas of leukoplakia coupled with areas of a warty surface texture. PVL is usually diagnosed from the fifth decade onwards and is more common in female patients. The most frequent sites involved tend to be gingivae, followed by buccal mucosa and lateral border of tongue. It is one of the oral potentially malignant conditions with a high risk of malignant transformation. It is important for general dental practitioners (GDPs) to identify such lesions to facilitate referral for further investigation and diagnosis. Management is challenging with long-term monitoring and surgical excision when appropriate; however, PVL tends to recur following surgical excision. This article provides an up-to-date review tailored for GDPs on the present knowledge of PVL and illustrates the management challenges with clinical cases.

Global gene expression profile of proliferative verrucous leukoplakia and its underlying biological disease mechanisms.

BACKGROUND: Proliferative verrucous leukoplakia (PVL) is a rare and enigmatic oral potentially malignant disorder which almost invariably results in oral squamous cell carcinoma (OSCC). The aims of this project were to use transcriptome profiling to characterise PVL gene expression patterns for biomarker identification and gain insight into the molecular aetiopathogenesis of PVL. METHODS: Forty-three oral cavity mucosal biopsies from 32 patients with oral lesions clinically compatible with either PVL or non-PVL conventional oral leukoplakia (OLK) underwent transcriptome profiling by RNA sequencing. Data was analysed by hierarchical clustering, differential gene expression, functional enrichment and network analysis, sparse partial least squares discriminant analysis sPLS-DA, and immune cell phenotypic estimation. RESULTS: We found 464 genes significantly differentially expressed at least 2-fold between PVL and non-PVL OLK (193 up and 271 down). HOX genes, including HOXA1 and HOXB7, keratin-associated proteins (KRTAPs) and olfactory receptor G proteins (OR) were significantly upregulated in PVL. Other upregulated genes in PVL included FOS, WNT16 and IFNA1. Pathway analysis showed that there was a significant downregulation of connective tissue signalling in PVL. Classifying multivariate models based upon 22 genes discriminated PVL from non-PVL OLK. Bioinformatic profiling showed that immune cell profiles in PVL and OLK were similar except that fibroblast markers were reduced in PVL. CONCLUSION: These results demonstrate that PVL and conventional OLK are molecularly distinct with upregulation of many cancer-associated genes. They provide insight into the pathogenesis of PVL and show that biomarker based molecular diagnostics is feasible to discriminate and inform diagnosis and management.

Clinical evaluation of photodynamic therapy for oral leukoplakia: a retrospective study of 50 patients.

BACKGROUND: Topical photodynamic therapy (PDT) has demonstrated encouraging results in the treatment of oral leukoplakia (OLK). However, data on the clinical efficacy of PDT in Chinese patients with OLK are still limited. METHODS: Fifty patients diagnosed with OLK were enrolled, including patients with various dysplastic tissues. All patients received topical PDT with 5-aminolevulinic acid (5-ALA) as a photosensitizer. Clinical efficacy was evaluated 4 weeks after treatment. Follow-up was performed every 3 months during the first year and every 6 months during the second year. RESULTS: The overall response rate was 68% (34/50): 12% (n = 6) complete and 56% (n = 28) partial responses. Aneuploidy was reduced in the patients with dysplastic lesions. Oral pain and local ulcers developed in 52% of the patients (n = 26). Patients with a long history of OLK including hyperplasia and dysplastic lesions, as well as those with non-homogenous lesions, were more likely to develop pain and ulcer. During follow-up, the recurrence rate of hyperplasia and dysplastic lesions was 32% (n = 16) and the malignant transformation rate of dysplastic lesions was 4% (n = 2). Lesions on the buccal mucosa were associated with recurrence (P = 0.044; OR: 0.108, 95% CI: 0.013-0.915). CONCLUSION: Topical 5-ALA-mediated PDT is an effective treatment for OLK, particularly for homogenous leukoplakia, with few side effects. The buccal mucosa may be a protective factor that can reduce recurrence.

Oral bacteriome and oral potentially malignant disorders: A systematic review of the associations.

INTRODUCTION: Periodontal bacteria can infiltrate the epithelium, activate signaling pathways, induce inflammation, and block natural killer and cytotoxic cells, all of which contribute to the vicious circle of carcinogenesis. It is unknown whether oral dysbiosis has an impact on the etiology or prognosis of OPMD. AIMS: Within this paradigm, this work systemically investigated and reported on the composition of oral microbiota in patients with oral potentially malignant disorders (OPMD) versus healthy controls. METHODS: Observational studies that reported next generation sequencing analysis of oral tissue or salivary samples and found at least three bacterial species were included. Identification, screening, citation analysis, and graphical synthesis were carried out. RESULTS: For oral lichen planus (OLP), the bacteria with the highest abundance were Fusobacterium, Capnocytophaga, Gemella, Granulicatella, Porphyromonas, and Rothia; for oral leukoplakia (OLK), Prevotella. Streptococci levels in OLK and OLP were lower. The usage of alcohol or smoke had no effect on the outcomes. CONCLUSIONS: An increase in periodontal pathogenic bacteria could promote the development and exacerbation of lichen. Effective bacteriome-based biomarkers are worthy of further investigation and application, as are bacteriome-based treatments.

Molecular landscape of proliferative verrucous leukoplakia: a systematic review.

Proliferative verrucous leukoplakia (PVL) is a rare oral potentially malignant disorder characterised by multifocal origin and unpredictable long-term evolution to oral squamous cell carcinoma (OSCC) or oral verrucous carcinoma (OVC). Currently no predictive biomarkers are in clinical use. We aimed to explore the genomic profile of PVL. A total of 685 cases in 26 studies were included in this review. Genomic data were presented in 15% of studies and biomarker analysis was reported in 85% of studies. At first clinical presentation, PVL is characterised by a high loss of heterozygosity (LOH), similar to OSCC, and low copy number alterations (CNA). As these progress, more CNAs and mutations in CDKN2A and alterations to ELAVL1 expression are noted, but no TP53 mutations are identified. There is significantly lower LOH at 17p in early PVL compared with OSCC (p = 0.037). Deletions in chromosomal loci 17q12, 5q31.1 and amplifications in 7q11.2, 7q22 are shared between early lesions and OVC. PVL shows CNAs at 11q31. WNT signalling pathway genes (SUZ12, CTTN and FOLR3) are enriched in CN-altered regions. PVL stroma shows significantly lower α-SMA and higher CD34 expression than OVC and OSCC. The exact genomic landscape is currently unclear, and further studies are necessary to unravel this mystery.

Copy Number Alterations Predict Development of OSCC from Oral Leukoplakia.

Oral leukoplakia (OLK) is a common type of potentially malignant disorder. Early identification of the malignancy potential leads to a better management of OLK and prediction of development of oral squamous cell carcinoma (OSCC). However, there has been no effective biomarker to assess the risk of malignancy in OLK. Genomic copy number alteration (CNA) is a complex chromosomal structural variation in the genome and has been identified as a potential biomarker in multiple cancers. This study aimed to develop a predictive model for the malignant transformation risk of OLK by copy number analysis. A total of 431 OLK samples with long-term follow-up (median follow-up of 67 mo) from multiple academic centers were analyzed for CNAs. CNA events increased with the severity of hyperplasia, mild dysplasia, moderate dysplasia, and severe dysplasia. More CNA events were present in patients with OLK who later developed OSCC than in those with OLK who did not. By multivariate Cox regression analysis, the OLK of the CNA scorehigh group showed an increased risk of malignant transformation than the CNA scorelow group (P < 0.001). A CNA score model was developed to accurately predict the prognosis (area under the receiver operating characteristic curve [AUC] = 0.879; 95% confidence interval [CI], 0.799-0.959) and was validated using data from 2 external centers (AUC = 0.836, 95% CI, 0.683-0.989; AUC = 0.876, 95% CI, 0.682-1.000), and all of them showed better prediction performances than histopathological grade in assessing the transformation risk of OLK. Furthermore, we performed CNA models among 4 subgroups of OLK with hyperplasia, mild dysplasia, moderate dysplasia, and severe dysplasia and found that CNA score can accurately predict malignant transformation of different subgroups. CNA score may be a useful biomarker to predict malignant transformation of OLK. Subtyping of OLK by the CNA score could contribute to better management of OLK and predicting development of OSCC.

Immunoexpression of SIRT1, 6, and 7 in oral leukoplakia and oral squamous cell carcinoma.

Sirtuins (SIRTs) are a family of proteins involved in the metabolic process responsible for extending the lifespan. The role of SIRT1, 6, and 7 in oral squamous cell carcinoma (OSCC) and oral leukoplakia (OLP), one of its precursors, is still elusive. In this study, 82 OLP and 77 OSCC were immunohistochemically examined for SIRT1, 6, and 7. Stained sections were thoroughly scanned and evaluated using a digital image analysis program. The SIRT1, 6, and 7 expressions were detected in the nuclei of epithelial and carcinoma cells in various degrees. Afterward, any correlations among SIRTs, including associations with clinicopathological features and the Kaplan-Meier curves were analyzed. OSCC demonstrated significantly higher SIRT1 expression than OLP, while non-dysplastic lesions showed significantly higher SIRT6 expression than other lesions. A strong correlation was observed between SIRT6 and 7 in OLP, SIRT1 and 6 in in OSCC and in SIRT6 and 7 when all lesion types were considered. There were no significant differences between SIRTs reactivity and the clinical features in OLP. For OSCC, SIRT1 and 6 was found to be directly associated with site of the lesion, while SIRT7 showed a direct relationship between gender, stromal lymphocytic infiltration, and depth of the invasion. OSCC with high SIRT7 expression revealed a slightly lower survival probability, although not statistically significant (p = 0.1019). Our findings suggest that SIRT1, 6, and 7 may play correlated and diverse roles in the development and advancement of OSCC.

Nivolumab for Patients With High-Risk Oral Leukoplakia: A Nonrandomized Controlled Trial.

Importance: Proliferative verrucous leukoplakia (PVL) is an aggressive oral precancerous disease characterized by a high risk of transformation to invasive oral squamous cell carcinoma (OSCC), and no therapies have been shown to affect its natural history. A recent study of the PVL immune landscape revealed a cytotoxic T-cell-rich microenvironment, providing strong rationale to investigate immune checkpoint therapy. Objective: To determine the safety and clinical activity of anti-programmed cell death 1 protein (PD-1) therapy to treat high-risk PVL. Design, Setting, and Participants: This nonrandomized, open-label, phase 2 clinical trial was conducted from January 2019 to December 2021 at a single academic medical center; median (range) follow-up was 21.1 (5.4-43.6) months. Participants were a population-based sample of patients with PVL (multifocal, contiguous, or a single lesion ≥4 cm with any degree of dysplasia). Intervention: Patients underwent pretreatment biopsy (1-3 sites) and then received 4 doses of nivolumab (480 mg intravenously) every 28 days, followed by rebiopsy and intraoral photographs at each visit. Main Outcomes and Measures: The primary end point was the change in composite score (size and degree of dysplasia) from before to after treatment (major response [MR]: >80% decrease in score; partial response: 40%-80% decrease). Secondary analyses included immune-related adverse events, cancer-free survival (CFS), PD-1 ligand 1 (PD-L1) expression, 9p21.3 deletion, and other exploratory immunologic and genomic associations of response. Results: A total of 33 patients were enrolled (median [range] age, 63 [32-80] years; 18 [55%] were female), including 8 (24%) with previously resected early-stage OSCC. Twelve patients (36%) (95% CI, 20.4%-54.8%) had a response by composite score (3 MRs [9%]), 4 had progressive disease (>10% composite score increase, or cancer). Nine patients (27%) developed OSCC during the trial, with a 2-year CFS of 73% (95% CI, 53%-86%). Two patients (6%) discontinued because of toxic effects; 7 (21%) experienced grade 3 to 4 immune-related adverse events. PD-L1 combined positive scores were not associated with response or CFS. Of 20 whole-exome sequenced patients, all 6 patients who had progression to OSCC after nivolumab treatment exhibited 9p21.3 somatic copy-number loss on pretreatment biopsy, while only 4 of the 14 patients (29%) who did not develop OSCC had 9p21.3 loss. Conclusions and Relevance: This immune checkpoint therapy precancer nonrandomized clinical trial met its prespecified response end point, suggesting potential clinical activity for nivolumab in high-risk PVL. Findings identified immunogenomic associations to inform future trials in this precancerous disease with unmet medical need that has been difficult to study. Trial Registration: ClinicalTrials.gov Identifier: NCT03692325.

Proliferative verrucous leukoplakia/multifocal leukoplakia in patients with and without oral submucous fibrosis.

BACKGROUND: Oral submucous fibrosis (OSF) and proliferative verrucous leukoplakia (PVL) are established as oral potentially malignant disorders. Dual pathology of the two conditions is not commonly encountered in clinical practice. This study aims to present a case series of multifocal leukoplakia in patients with and without OSF to outline the clinical behavior and challenges in the management of this high-risk group in clinical practice. MATERIAL AND METHODS: We retrospectively analyzed cases of six Indian patients (four with OSF) managed over a period of 5.5 to 13 years at the Government Dental College, Nagpur. Patient data consisting of age, gender, medical history, habits, clinical findings, and biopsy reports were recorded at the initial visit. During follow-up visits, the clinicopathological data were reassessed. When surgical intervention failed to arrest the disease or when surgery was contraindicated metronomic therapy with Folitrax 15 mg once a week and Celecoxib 100mg twice daily was initiated. RESULTS: All patients developed PVL after the initial pathology diagnosis of OSF or oral leukoplakia. Initial lesions were either homogenous or non-homogenous leukoplakia. All patients developed multiple recurrences, regional or systemic metastasis. Despite thorough interventions, the patients died of, or with the disease. CONCLUSIONS: The occurrence of two or more oral potentially malignant disorders poses challenges in patient management and possibly presents a higher risk of malignant transformation. More clinical trials are necessary to assess the benefits of metronomic therapy for patients diagnosed with aggressive PVL concurrently found with OSF.

Nano-drug delivery system targeting FAP for the combined treatment of oral leukoplakia.

Oral leukoplakia (OLK) has received much attention due to its potential risk of malignant transformation. Studies have shown that when drug therapy is combined with photothermal therapy (PTT), not only can the cytotoxicity of the drug be enhanced, but also the heat energy can be used to kill the lesion cells, so we can combine drug therapy with PTT to enhance the therapeutic effect on OLK. However, with certain drawbacks due to its lack of targeting, fibroblast activating protein (FAP) has become an attractive target for OLK combination therapy. In this study, we used NGO-PEG loaded with FAP-targeting peptide (F-TP) and celecoxib (CXB) to construct a nano-drug delivery system CGPF for targeting OLK with high FAP expression and confirmed the biocompatibility and therapeutic efficacy of CGPF by in vitro and in vivo experiments. Overall, the novel nano-drug delivery system CGPF proposed in this study showed a very significant potential for the combination therapy of OLK.

Correlation of PD-1 and PD-L1 expression in oral leukoplakia and oral squamous cell carcinoma: an immunohistochemical study.

The programmed cell death protein (PD-1)/programmed cell death protein ligand (PD-L1) pathway and cytotoxic T lymphocyte antigen are the most important co-stimulatory molecules that play a key role in the negative regulation of T cells during carcinogenesis. We aimed to evaluate the immunohistochemical expression of PD-1 and PD-L1 in oral leukoplakia and squamous cell carcinoma compared with normal oral mucosa. Twenty-five cases of oral squamous cell carcinoma, oral leukoplakia and normal oral mucosa tissue specimens were immunohistochemically stained to assess PD-1 and PD-L1 expression. The PD-L1 positivity of subepithelial TAFs (p < 0.001) increased with increasing grades of oral leukoplakia. Pearson's correlation indicated a high positive correlation between the PD-L1 labelling index of epithelial tumour cells and the PD-1 labelling index of tumour infiltrating lymphocytes (p value: 0.005) in OSCC. A high positive correlation was noted between the H-score of PD-L1 positive tumour epithelial cells and the H-score of PD-1 positive tumour infiltrating lymphocytes in OSCC (p value: 0.001). PD-L1 positivity increased in dysplastic epithelial cells from premalignant lesions to malignancy. The sub-epithelial PD-L1 positive TAFs were higher in oral leukoplakia compared to OSCC inferring that PD-L1 positivity in TAFs decreased with malignant transformation. The PD-1 positivity in TILs was higher in oral leukoplakia than in OSCC.

Compositional and functional changes in the salivary microbiota related to oral leukoplakia and oral squamous cell carcinoma: a case control study.

BACKGROUND: Oral squamous cell carcinoma (OSCC) is one of the most common malignant tumours with increasing incidence, and oral leukoplakia (OLK) has a strong tendency to undergo malignant transformation. The oral microbiota may influence oral cancer progression, but the salivary bacterial composition and functional changes in OSCC and OLK have not been comprehensively elucidated. Therefore, we compared salivary bacteria in OLK and OSCC patients with healthy controls (HC). METHODS: Metagenomic sequencing was used to compare the bacterial composition and functional changes of 18 OSCC patients, 21 OLK patients and 21 HC. Spearman correlation was used to identify possible associations between functions and bacteria. RESULTS: Gemella was the most differentially enriched genus in OSCC. At the species level, Streptococcus sp. NPS 308, Streptococcus agalactiae, Gemella haemolysans and Gemella morbillorum were slightly increased in OLK and OSCC. Kyoto Encyclopedia of Genes and Genomes (KEGG) results showed that OSCC was mainly associated with metabolism functions, including lipid metabolism, carbohydrate metabolism and glycan biosynthesis and metabolism. The synthesis and degradation of ketone bodies, cysteine and methionine metabolism and glycerolipid metabolism differed significantly among the three groups, and were highest in OSCC and lowest in HC. And G. haemolysans was significantly associated with these selected metabolic pathways. CONCLUSIONS: Metagenomic analysis revealed significant differences in the salivary microbiota among OSCC, OLK and HC. Thus, salivary microbiota composition and functional changes may be associated with OSCC progression. Metabolism of nonessential amino acids such as cysteine and methionine in bacteria may play an important role in oral oncogenesis, and more studies of the mechanism between metabolisms of bacteria and oral oncogenesis are needed in the future.

Implication of TIPE2 Expression on the Malignant Transformation of Oral Leukoplakia.

BACKGROUND/AIM: Identification of biomarkers involved in the malignant transformation of oral leukoplakia (OL) is required for the early diagnosis and management of patients with OL. This study aimed to evaluate the functions of tumor necrosis factor-alpha-induced protein 8-like 2 (TIPE2) expression in the malignant transformation of OL. MATERIALS AND METHODS: The expression levels of TIPE2 and dormant cell markers phospho-ERK and phospho-p38 in a cohort containing 103 surgical specimens from patients with OL were evaluated using immunohistochemistry. The influence of TIPE2 expression on the biological behavior of the immortalized human oral keratinocyte (IHOK) line was investigated in vitro. RESULTS: Increased TIPE2 expression was detected in 40 (38.8%) patients with OL. In a multivariate analysis using clinicopathological variables and TIPE2 expression as cofactors, the presence of dysplasia (p=0.003) and TIPE2 abundance (p=0.019) were identified as independent risk factors for the malignant transformation of OL. Moreover, the in vitro analysis revealed that TIPE2 knockdown can promote the proliferating ability of IHOK; however, the number of apoptotic cells also increased after TIPE2 knockdown in IHOK. Furthermore, TIPE2 expression was significantly associated with phospho-p38 expression, a dormant cell marker, in our cohort (p=0.047). CONCLUSION: TIPE2 expression may contribute to the malignant transformation of OL, and its function may be related to cellular dormancy in OL pathogenesis.

Proliferative verrucous leukoplakia: a case report of multiple metachronous oral verrucous carcinoma.

Proliferative verrucous leukoplakia (PVL) is a distinct progressive and multi-focal form of oral leukoplakia, not associated with the traditional risk factors (ie, tobacco and alcohol consumption). The incidence of oral squamous cell carcinoma in PVL patients is high. Here, we describe the case of a patient affected by PVL, who developed two metachronous oral verrucous carcinomas at different sites of the oral mucosa. Owing to the high risk of multiple oral squamous cell carcinoma, periodical clinical and histopathological follow-up is mandatory and should continue lifelong.

Etiology of Oral Potentially Malignant Disorders and Squamous Cell Carcinoma Based on Cellular Stress Regulation and Matrix Stiffness.

The oral cavity serves as the initial segment of the digestive system and is responsible for both nutritional supplementation and the mechanical breakdown of food. It comprises distinct hard and soft tissues; the oral mucosa is subject to mechanical stress and interaction with microbiota. In oral cancer, tumors exhibit abnormal cellular networks and aberrant cell-cell interactions arising from complex interplays between environmental and genetic factors. This presents a challenge for clinicians and researchers, impeding the understanding of mechanisms driving oral cancer development and treatment strategies. Lesions with dysplastic features are categorized under oral potentially malignant disorders, including oral leukoplakia, erythroplakia, oral submucous fibrosis, and proliferative verrucous leukoplakia, carrying a high malignancy risk. In this review, we discuss oral cancer cell characteristics and the stiffness of the surrounding matrix. We also discuss the significance of stiffness equilibrium in oral potentially malignant disorders, particularly oral submucous fibrosis, possibly triggered by mechanical stress such as betel quid chewing.

Oral erythroplakia and oral erythroplakia-like oral squamous cell carcinoma - what's the difference?

BACKGROUND: Oral erythroplakia (OE) is a rare oral potentially malignant disorder, that has a high rate of malignant transformation. The definition of OE still lacks uniformity. In particular, lesions that look clinically like erythroplakias, but are histopathologically diagnosed as squamous cell carcinomas are still sometimes called erythroplakias. The purpose of this study is to present demographic and clinicopathologic features of a series of OEs and clinically oral erythroplakia -like squamous cell carcinomas (OELSCC), to study their differences and to discuss the definition of OE. METHODS: A multicenter retrospective case series of OEs and OELSCCs. Descriptive statistics were used to analyze the data. RESULTS: 11 cases of OEs and 9 cases of OELSCCs were identified. The mean age of the OE patients was 71 years and 72.7% were female, while the mean age of the OELSCC patients was 69 years, and all were female. 9% of the OE and 22% of the OELSCC patients had smoked or were current smokers. 72.7% of the OEs and 55.5% of OELSCCs were uniformly red lesions. 63.6% of the OE and 22% of the OELSCC patients had a previous diagnosis of oral lichenoid disease (OLD). The malignant transformation rate of OE was 9% in a mean of 73 months. CONCLUSIONS: OE and OELSCC may arise de novo or in association with OLD. Tobacco and alcohol use were not prevalent in the present cases. The clinical features of OEs and OELSCC are similar, but symptoms, uneven surface and ulceration may be more common in OELSCCs than in OEs. Clinical recognition of OE is important since it may mimic other, more innocuous red lesions of the oral mucosa. The diagnosis of OE requires biopsy and preferably an excision. Clarification of the definition of OE would aid in clinical diagnostics.

Assessment of brush biopsy findings and salivary LDH levels in oral mucosal lesions of tobacco users.

Introduction: The oral brush cytology is an alternative method developed to improve the efficacy of conventional cytology in oral potentially malignant disorder (OPMD), and salivary lactate dehydrogenase (LDH) which is a cytoplasmic enzyme has been widely used as a marker for diagnosing various diseases. The purpose of the study is to evaluate the brush biopsy findings and salivary LDH levels for the early diagnosis of premalignant and malignant lesions of the oral cavity. Materials and Methods: Patients with deleterious habits including tobacco-related lesions such as leukoplakia, tobacco pouch keratosis, and oral cancer were included in the study. For each patient, saliva sample was collected, brush biopsy was done and smears were prepared. Collected saliva samples were analysed for salivary LDH levels and prepared smears were analysed for dysplastic changes and statistical analysis was performed. Results: Out of 80 samples, 30 were leukoplakia, 45 were tobacco pouch keratosis and 5 were oral cancer, and 13 samples showed positive dysplastic changes, 26 samples showed atypical dysplastic changes and 41 samples showed no signs of dysplastic changes and concluded as negative. On comparing the results of brush biopsy findings and salivary LDH levels, the mean salivary LDH value for positive dysplasia was elevated and the P value was statistically significant (P value: 0.00). Conclusion: Brush biopsy showed good potential in detecting premalignant lesions and salivary LDH levels showed a marked increase which can be used as a diagnostic biomarker and serve as a potent diagnostic aid for early detection of malignancy.