In vitro triple culture model of retinoblastoma for pre‐clinical investigations

Abstract

Background: Retinoblastoma (Rb) is a rare cancer of the retina that occurs during early childhood. The disease is relatively rare but aggressive, accounting for ∼3% of childhood cancers. Treatment modalities encompass the administration of large doses of chemotherapeutic drugs, which result in multiple side-effects. Therefore, it is essential to have safe and effective newer therapies and suitable physiologically relevant, alternative-to-animal, in vitro cell culture-based models to enable rapid and efficient evaluation of potential therapies. Methodology:This investigation was focused on the development of a triple co-culture model comprising Rb, retinal epithelium, and choroid endothelial cells, using a protein coating cocktail, to recapitulate this ocular cancer under in vitro conditions. This resulting model was used for screening drug toxicity, based on the growth profile of Rb cells, using carboplatin as the model drug. Further, a combination of bevacizumab and carboplatin was evaluated using the developed model, to lower the concentration of carboplatin and thereby reduce its physiological side-effects. Major Results:The effect of drug treatment on the triple co-culture was assessed by increase in the apoptotic profile of Rb cells. Further, the barrier properties were found to be lower with a decrease in the angiogenetic signals that included expression of vimentin. Measurement of cytokine levels signified reduced inflammatory signals due to the combinatorial drug treatment. Conclusions:These findings validated that the triple co-culture Rb model was suitable for evaluating anti-Rb therapeutics and could thereby decrease the immense load on animal trials, which are the primary screens employed for evaluating retinal therapies. K E Y W O R D Sdisease model, pre-clinical drug testing, retinal cells, retinoblastoma, triple culture INTRODUCTIONRetinoblastoma (Rb) is a type of ocular cancer that is particularly prevalent in children up to 5 years of age. This form of cancer accounts for ∼3% of the pediatric cancer population. [1] Its progression is activated due to the mutation in the Rb gene, responsible for the transcription tumor suppressor Rb protein. [2] Rb tumors occur either as endophytic or exophytic tumors. Endophytic tumors arise in the inner retinal layer, whereas exophytic tumors develop in the outer retinal layer further extending the sub-retinal space beneath the retina. [3] The currently available treatment options for Rb can be classified as (1) focal therapy including cryotherapy, conventional laser photocoagulation, transpupillary thermotherapy, photodynamic therapy (in vitro study), and plaque radiotherapy; (2) external beam radiotherapy;(3) systemic chemotherapy; and (4) enucleation. Proper management of Rb, by tumor reduction, excision, and curbing the side effects of

Histone Deacetylases in Retinoblastoma

“…This model, established using a protein coating cocktail, aimed to replicate the complex interactions within the ocular environment under in vitro conditions. By incorporating multiple cell types relevant to retinoblastoma, this model provided a comprehensive platform for studying the disease and evaluating potential therapeutic interventions [32].…”