Investigating the effects of cisplatin on long non-coding RNAs ANRIL and PANDA in oesophageal squamous cell carcinoma (OSCC) cell lines

Abstract

Oesophageal cancer is the 8th most common cancer type and the 6th most death causing cancer type worldwide. One of its subtypes, oesophageal squamous cell carcinoma (OSCC) is prevalent in Eastern to Central Asia and along the Rift Valley in East Africa and into South Africa. The prognosis of OSCC remains poor, detected at the advanced stage with low survival rates. There are various treatment options for OSCC and cisplatin is the first line chemotherapeutic drug used in South Africa and other developing countries as it is readily available and affordable. However, it is associated with poor efficacy, chemoresistance and chemotoxicity.

This study investigated the expression of ANRIL and PANDA long non-coding RNAs (lncRNAs) in response to cisplatin and immediately after cisplatin withdrawal, aiming to provide potential lncRNA-targeted strategies for overcoming cisplatin resistance in cancer therapy, especially OSCC. Cisplatin cytotoxicity and EC50 concentrations in OSCC (WHCO1 and WHCO5) and HEK293 cell lines were determined by an MTT assay. Cisplatin cytotoxic effects were exerted in all cell lines, with the HEK293 control being more sensitive to cisplatin compared to the cancerous OSCC cell lines. Western blot was used to observe DNA double stranded break (DSB) formation in the presence of cisplatin and in the first hour after cisplatin withdrawal in HEK293, WHCO1 and WHCO5 cell lines by measuring gammaH2AX (γH2AX), a DSB marker. The western blot results showed an increase in γH2AX levels post cisplatin treatment in all three cell lines. This showed cisplatin treatment induced DNA DSBs and activated the DNA damage response (DDR) in HEK293 control as well as WHCO1 and WHCO5. RT-PCR measured ANRIL and PANDA expression during cisplatin treatment and DDR, where PANDA expression appeared upregulated and ANRIL fluctuated during DDR in OSCC cell lines.

In conclusion, cisplatin activates DDR, upregulates PANDA but causes fluctuating ANRIL expression in OSCC cell lines. This study indicates roles for PANDA and ANRIL lncRNAs during cisplatin chemotherapy that warrant further investigation in OSCC.