81P - Evaluating the effect of lenvatinib-resistance in hepatocellular carcinoma cells and in lenvatinib-resistant patient-derived PBMCs

Background

Lenvatinib (LNV) represents a first-line therapeutic agent for the treatment of patients with advanced hepatocellular carcinoma (HCC). The efficacy of LNV in the targeted therapy of numerous cancer types has been demonstrated; however, a significant proportion of patients do not receive long-term benefit due to primary and acquired drug resistance. In this work we aimed to identify molecular pathways involved in acquired resistance to lenvatinib in different HCC models.

Methods

For this study, different models of HCC were used: (1) resistant cell lines, (2) resistant patients PBMCs and (3) co-culture model. Huh-7, SNU449 and HepG2 cell lines were characterised using western blot (WB), colony formation and migration assay. LNV-resistant cell lines (Huh-7/LR and SNU449/LR) were established by increasing doses of LNV (1 to 40 μM). Mechanisms of resistance were explored, and MTT assays were conducted to determine IC50 and cross-resistance acquisition in LR cells to other TKIs. Validation study in co-culture of the HCC-LR cell line and LR-patient-derived PBMC were performed.

Results

WB, migration and colony formation assay of the HCC cell lines enabled the characterisation of the cell lines together with the evaluation of EMT markers. Huh-7 and SNU449, showing a high mesenchymal phenotype and a higher IC50 value than HepG2, were exposed to LNV to induce resistance. SNU449/LR showed the highest levels of p-AKT compared to parental cell lines, together with an increase of the EMT phenotype, i.e. a decrease in E-cadherin and an increase in vimentin and snail, as compared to Huh7-LR. Similarly, an increase in p-EGFR and EGFR was observed in SNU449/LR, while a decrease was seen in Huh-7/LR. Interestingly, LR-cell lines showed sensitivity to Cabozantinib treatment compared to Sorafenib. Further experiments with PBMCs from LR-patients are ongoing to validate findings and explore cross-sensitivity to TKIs alone or in combination with immunotherapy (IO).

Conclusions

Our findings indicate that Cabozantinib is the most effective TKI in LR-cells. Co-culture models of LR-HCC cells and LR-patient-derived PBMCs represent a useful tool to test the cross-resistance ex vivo for TKI combined with IO.

Editorial acknowledgement

Clinical trial identification

Legal entity responsible for the study

The authors.

Funding

Has not received any funding.

Disclosure

All authors have declared no conflicts of interest.