Scientists Uncover Potential TMZ-Resistance Reversal in Glioblastoma

TMZ-resistance in glioblastoma

A team of scientists from Virginia Tech Carilion Research Institute and clinicians from Carilion Clinic may have found a way to increase sensitivity to the front-line chemotherapeutic agent temozolomide (TMZ) in glioblastoma (GBM), even after resistance.

According to a preclinical study,¹the team found an inverse correlation between gap junction protein connexin 43 (Cx43) levels in patients and TMZ sensitivity of GBM cells, including GBM stem cells. The study states that presence of Cx43 was inversely correlated with patient survival. This survival rate includes patients with GBM, who are O-6-methylguanine-DNA methyltransferase (MGMT) deficient; MGMT is a DNA strand that allows resistance of TMZ to occur.

The study states that when given the C-terminal peptide mimetic αCT1, a selective inhibitor of Cx43 channels, with TMZ, surgically obtain GBM cells, with MGMT deficiency and TMZ resistance became sensitive to TMZ treatment. In addition to increasing sensitivity, the team found that combining αCT1 with TMZ also blocked AKT/mTOR signaling, induced autophagy, and apoptosis in TMZ-resistant GBM cells.

"A novel paradigm is proposed for development of resistance of MGMT-deficient GBM patients to TMZ. In MGMT-deficient GBM cells, Cx43 activates AKT followed by inactivation of AMPK and activation of mTOR signaling. This leads to suppression of cell death," said Murphy et al, in the study.

"Thus, these MGMT-deficient GBM cells are TMZ resistant. On the other hand, αCT1, a selective inhibitor of Cx43 channel activity, blunts this process, and sensitizes MGMT-deficient GBM cells to TMZ-induced cell death. Hence, these MGMT-deficient GBM cells become sensitive to TMZ. This new paradigm demonstrates the potentially important role of Cx43 in sensitivity to the anticancer drug TMZ. Our results presented herein also offer possible options for the diagnosis and treatment of TMZ-resistant GBM."

GBM accounts for more than 45% of all malignant brain tumors, with the 5-year survival rate after aggressive treatment of <5%. The study stated that even after surgical resection of tumors, and with the combination of TMZ and radiation therapy, median survival is only increased from 12.1 months to 14.6 months.

The preclinical study initially tested Cx43-mediated TMZ resistance with a rat malignant glioma cell line C6 that ectopically expresses Cx43. C6 cells with exogenous Cx43 showed resistance, which prompted the team to study Cx43 expression and MGMT-independent TMZ resistance in GBM cell lines.

"We found that of the MGMT-negative GBM cell lines, Cx43 was highly expressed in SF295 and U87MG cells. Intriguingly, the reported TMZ IC50s of SF295 and U87MG cells were remarkably higher than those of the other GBM cell lines" said Murphy et al in the study.

"As we could not find it documented in the literature, we assayed the TMZ IC50 of SNB75 cells and determined it to be 293 &mu;M. To validate TMZ sensitivities reported previously for other GBM cell lines, we treated U87MG, SF295, SF268, and U251 cells with different doses of TMZ. The IC50s of above cell lines were 1369, 513, 274, and 129 &mu;M, respectively. Although SF268 and U251 cells were less sensitive to TMZ in our hands, the difference in TMZ responsiveness among these GBM cell lines was more or less in line with those measured by earlier workers."

The study states regression analysis of of Cx43 and TMZ IC50s in MGMT-deficient cell lines showed a very clear relationship between TMZ responsiveness and Cx43 levels. With this trend also came the revelation that high Cx43-expressing lines SF295 and U87MG showcase the lowest cytotoxicity when treated with 100 &mu;M of TMZ for 3 days, as well as showcasing an obvious correlation between TMZ-induced reduction in cell viability and levels of Cx43.

"We next examined Cx43 expression and TMZ responses in GBM primary cells from freshly resected patient tumors. The levels of Cx43 in all GBM patient samples were approximately 6- to 14-fold higher than that in the normal brain, consistent with the results from The Protein Atlas database. Among all six primary GBM cells, only VTC-003 cells expressed no MGMT. The ratio of Cx43 to ACTB in VTC-003 cells was equivalent to that in SF295 cells. As expected, VTC-003 exhibited significant resistance to TMZ with an IC50 of approximately 3 mM," said Murphy et al in the study.

Murpy et all concluded that TMZ has historically been used to treat other types of cancers, such as neuroblastoma, Ewing's sarcoma, and hepatocarcinoma. The team said this breakthrough in resensitizing GBM patients to TMZ treatment could mean new therapies for patients with GBM, as well as for others.

References

1. Murphy S, Varghese R, Lamouille S et al. Connexin 43 inhibition sensitizes chemoresistant glioblastoma cells to temozolomide.http://cancerresaacrjournalsorg/. 2015. Available at: http://cancerres.aacrjournals.org/content/early/2015/11/05/0008-5472.CAN-15-1286.full.pdf. Accessed November 20, 2015.