Temozolomide Resistance is Associated with Upregulated MICB mRNA Expression and Protein Secretion in Human Glioblastoma Cells

Abstract

BACKGROUND: Standard treatment for glioblastoma multiforme (GBM) chemotherapy with temozolomide (TMZ) is often limited by chemoresistance, which is correlated with cancer immune evasion caused by the dysregulation of natural killer group 2 member D ligands (NKG2DL). While suppression of NKG2DL facilitates cancer progression, how chemoresistance regulates NKG2DL expression remains unclear. This study was conducted to examine the association of TMZ resistance with the expression and secretion of the NKG2DL subtype, MHC class I chain-related protein B (MICB), in human GBM cells.

METHODS: An experimental in vitro study was conducted using TMZ-sensitive (U87MG) and intrinsically TMZ-resistant (T98G) human GBM cell lines. MICB mRNA expression was analyzed via quantitative reverse transcription polymerase chain reaction (qRT-PCR). Surface protein expression and extracellular secretion of MICB were measured using flowcytometry and enzyme-linked immunosorbent assay (ELISA), respectively.

RESULTS: Repeated TMZ exposure successfully induced resistance in U87MG-R (+TMZ) cells, yielding significantly higher cell viability (p<0.01) and slower proliferation rates compared to parental U87MG cells. Following 2-days TMZ treatment, both intrinsically resistant T98G and adaptively resistant U87MG-R (+TMZ) cells, demonstrated a significant increase in MICB mRNA levels (p<0.05 and p<0.0001, respectively) and extracellular secretion of soluble MICB protein (p<0.01 and p<0.01, respectively), compared to TMZ-sensitive counterparts. Conversely, MICB surface protein level after 2-days TMZ treatment was significantly reduced in both T98G (p<0.0001) and U87MG-R (+TMZ) (p<0.01) cells relative to the respective U87MG cells.

CONCLUSION: GBM cell resistance to TMZ was associated with an upregulation of MICB mRNA transcription and extracellular protein secretion, and concurrently with a suppression of MICB expression on the cell surface. These conditions may lead to a cellular adaptive mechanism to develop resistance to TMZ and may represent a strategic route to evade native host immune.

KEYWORDS: glioblastoma multiforme, temozolomide, cancer chemoresistance, MHC class I chain-related protein B (MICB), natural killer group 2D receptor (NKG2D)