Higher isoform of hnRNPA1 confer Temozolomide resistance in U87MG & LN229 glioma cells.

BACKGROUND

Gliblastoma is a malignant brain tumor; despite available treatment modalities, the tumor reoccurrence rate persist in the currently prescribed Temozolomide chemotherapy. Study aimed to study the inquisitive role of RNA binding splice factor protein hnRNPA1 in promoting glioma resistance against Temozolomide drug and therapeutic insights.

METHODS

In this study two non-expressing O-methylguanine-DNA methyltransferase (MGMT) glioma cell lines U87MG & LN229. U87MG cells were grown in Temozolomide from 50μM upto 400μM & LN229 cells grown upto 200μM, till then both these cells acquired Temozolomide resistance. Both of these cells were grown & maintained continously in its highest dose of Temozolomide (TMZ). Splice factor protein SF2/ASF1 was functionally correlated with abundance of hnRNPA1 protein in Temozolomide (TMZ) resistant cells using its specific siRNA transfection approach, in detrmining SF2/ASF1 mediated hnRNPA1 splicing and Temozolomide resistant reversal.

RESULTS

U87MG TMZ resistance, results an increase in the expression of pre mRNA-splicing factor SF2/ASF1, Heterogeneous Ribonucleoprotein A1 (hnRNPA1) and O-methylguanine-DNA methyltransferase (MGMT) protein. MGMT expression was not observed in LN229 TMZ resistant cells. Further, mRNA sequencing of hnRNPA1 confirmed the exclusive abundance of its higher isoform in TMZ- resistant cells along with increase in SF2/ASF1 expression. Knocking down of SF2/ASF1 using its specific siRNA reverted the higher isoform of hnRNPA1 isoform Var2 to its lower isoform hnRNPA1 Var1 in U87 TMZ resistant cells, reveals hnRNPA1 alternative higher isoform abundance is SF2/ASF1 splice factor dependent. Additionally, selective knock down of hnRNPA1 higher isoform Var2 in TMZ resistant U87MG & LN229 promotes apoptosis, was further specfically enhanced on Wortmannin (PI3Kinase inhibitor) treatment.

CONCLUSION

Targeting higher isoform Var2 of hnRNPA1 specifically induces chemosensitization in MGMT expressed Temozolomide resistant U87MG as well as in MGMT non-expressed LN229 TMZ resistant cells.