Background Microtubule-associated protein light chains (LC) 3A and 3B are the structural proteins of the autophagosomal membrane widely used as endogenous autophagy markers. LC3A and LC3B autophagosomes reportedly have a distinct subcellular localization yet their role in the transition from premalignant to malignant phase remains unclear. This exploratory study aimed to investigate the expression of autophagy-related proteins LC3A and LC3B in oral premalignant lesions (OPL) and oral squamous cell carcinoma (OSCC) cases. Methodology A cross-sectional study was conducted on 100 OPL and 39 OSCC samples. OPL samples comprised both dysplastic and non-dysplastic lesions. The expression of LC3A and LC3B markers was evaluated in the study samples using immunohistochemistry and associated with dysplasia in OPL and with invasive OSCC versus OPL. Fisher's exact test was used for statistical analysis. Results There was a higher ratio of LC3A positivity in non-dysplastic OPL (31/38) compared to dysplastic premalignant lesions (36/62, p=0.017). There was a higher ratio of LC3B positivity in dysplastic OPL (16/62) compared to non-dysplastic lesions (4/38) with a trend towards statistical significance (p=0.075). There was no statistical difference in the ratio ofLC3A positivity between OSCC (23/39) and premalignant (67/100) lesions, while the ratio of LC3B marker positivity was higher in OSCC cases (18/39) relative to premalignant lesions (20/100, p=0.003). Conclusion Autophagy-related proteins LC3A and LC3B may have different roles to play in a disease context manner. LC3A is likely to be negatively associated with dysplasia in OPL while LC3B expression is positively associated with carcinogenesis of OSCC, possibly including dysplasia.