IN VITRO EVALUATION OF STRATEGIES TO OVERCOME PHARMACOLOGICAL RESISTANCE IN RETINOBLASTOMA

Abstract

Purpose : During the last decades, retinoblastoma (Rb) treatments have changed to local delivery of chemotherapy greatly improving ocular preservation rates. Still, some eyes relapse or present recurrence probably due to acquired pharmacological resistance. Our aim was to establish a topotecan- or melphalan-resistant cell subtype (HSJD-RBT-7-TP, HSJD-RBT-7-ML) as tools for studying the pharmacological sensitivity and alternative treatments to overcome resistance.

Methods : A primary cell culture was established from the tumor biopsy of an upfront enucleated patient with intraocular Rb. The parental cell line (HSJD-RBT-7) was exposed to three weekly doses of topotecan or melphalan equivalent to the IC50 to generate the resistant cell subtypes (HSJD-RBT-7-TP and HSJD-RBT-7-ML, respectively). Once established, we determined the sensitivity to topotecan, melphalan and carboplatin to evaluate the generation of resistant and cross-resistance. Alternative treatments included digoxin exposure and metronomic topotecan or melphalan. Cell viability was determined using the MTT assay.

Results : Mean (range) topotecan IC50 in HSJD-RBT-7-TP cells was 8.1 nM (6.8-9.3), 3-fold higher than that in HSJD-RBT-7 (p<0.05). Mean (range) melphalan IC50 in HSJD-RBT-7-ML was 1.3 µM (0.8-1.4), 4.3-fold higher than that in HSJD-RBT-7 (p<0.05). Cross-resistance to melphalan and carboplatin was observed in HSJD-RBT-7-TP cells resulting in a 2.5-fold and 1.8-fold increase in IC50 (p<0.05). Also, cross-resistance to topotecan and carboplatin was observed in HSJD-RBT-7-ML cells showing a 2.4-fold and 3.4-fold increase in IC50 (p<0.05). Generation of pharmacological resistance did not alter the sensitivity to digoxin as the IC50 remained similar between the parental and resistant cells (0.05 µM and 0.06 µM, respectively). HSJD-RBT-7-TP and HSJD-RBT-7-ML showed a significant decrease in topotecan (4 fold) and melphalan (2.5 fold) IC50 after treating cells with metronomic schemes (p<0.05).

Conclusions : Topotecan- and melphalan-resistant cell subtypes showed a significant decrease in drug sensitivity. Cross-resistance to frequently used drugs was observed demonstrating the importance of using new drug. Digoxin and metronomic treatment are two alternatives that successfully overcome melphalan and topotecan resistance in Rb tumors.

This abstract was presented at the 2023 ARVO Annual Meeting, held in New Orleans, LA, April 23-27, 2023.