Objective: To investigate the expression of phosphatidylinositol proteosan-3 (Glypican-3) and Heat shock protein 70 (HSP70) in Hepatitis B virus-associated liver cancer and their relationship with clinicopathology. Methods: Sixty-two patients with HBV-related liver cancer who were admitted to our hospital from October 2022 to October 2024 were selected as the study objects. Immunohistochemistry was used to detect the expression levels of Glypican-3 and HSP70 in liver cancer and adjacent tissues, and the relationship between the expression of Glypican-3 and HSP70 and clinicopathological features of patients was analyzed. Pearson correlation was used to analyze the relationship between Glypican-3, HSP70 and clinicopathological parameters. Results: The positive expression rates of Glypican-3 and HSP70 in liver cancer tissues were 75.81% and 80.65%, respectively, which were significantly higher than the positive expression rates of Glypican-3 and HSP70 in paracancer tissues (24.19% and 29.03%, P<0.05). The positive expression rate of Glypican-3 in liver cancer tissues was significantly different among patients with different degrees of differentiation (P<0.05). There were significant differences in the positive expression rate of HSP70 among patients with different tumor length, TNM stage and differentiation degree (P<0.05). The expression of Glypican-3 was positively correlated with TNM stage and differentiation degree, and the expression of HSP70 was positively correlated with tumor length, TNM stage and differentiation degree (P<0.05). Conclusion: Both Glypican-3 and HSP70 are highly expressed in HBV-related liver cancer tissues, and their positive expression is correlated with tumor length, TNM stage and differentiation degree.