Growth suppression of glioblastoma multiforme cell line response to the combination of Temozolomide and Nimotuzumab

Made Agus Mahendra Inggas; Upik Andriani Miskad; Andi Asadul Islam; Eka Julianta Wahjoepramono; Sri Maliawan

doi:10.15562/bmj.v10i3.2896

Growth suppression of glioblastoma multiforme cell line response to the combination of Temozolomide and Nimotuzumab

Made Agus Mahendra Inggas ,

Upik Andriani Miskad ,

Andi Asadul Islam ,

Eka Julianta Wahjoepramono ,

Sri Maliawan ,

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DOI: https://doi.org/10.15562/bmj.v10i3.2896 |
Published: 2021-12-30

Abstract

Background: Researches towards Glioblastoma multiforme therapy (GBM) has been performed, especially in finding the tumor’s genetic cellular expression. The overexpression of EGFR, frequently found in human malignancies, is known to significantly impact cancer cells hallmark traits, such as increased cell survival, proliferation and invasion. This research is performed to figure out the synergistic effect of temozolomide, the main chemotherapy regimen used, and the monoclonal antibody anti-EGFR nimotuzumab in inhibiting the growth of GBM cells in vitro.

Methods: We performed cell cultures using GBM U87MG cells and administered nimotuzumab 1000µg/ml and temozolomide 20 µg/ml in different sequences and timings. Inhibition of GBM cell growth is evaluated by measuring Ki-67 and ?H2AX levels using flowcytometry and tested with one-way ANOVA. Data were analyzed using SPSS version 21 for Windows.

Results: Levels of Ki-67 were in the control group (23.17±1.72), nimotuzumab monotherapy (15.43±1.70), temozolomide monotherapy (14.80±1.37), simultaneous therapy (10.73±1.19), nimotuzumab 24 hours before temozolomide (10.57±1.05), and nimotuzumab 48 hours before temozolomide (14.47±1.37). The level of ?H2AX is measured in the control group (11.90±1.25), nimotuzumab monotherapy (29.33±1.91), temozolomide monotherapy (28.13±1.58), simultaneous therapy (41.53±3.51), nimotuzumab 24 hours before temozolomide (39.56±2.06), and nimotuzumab 48 hours before temozolomide (35.93±3.56).

Conclusion: This research shows that administration of nimotuzumab simultaneously with temozolomide and nimotuzumab 24 hours before temozolomide effectively inhibits the growth of U87MG GBM cells. Inhibition of EGFR expression before temozolomide administration is able to increase DNA damage and inhibit the proliferation of glioma cells.

References

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Lee SY. Temozolomide resistance in glioblastoma multiforme. Genes & Diseases. 2016: 3: 198 – 210
Lai SW, Huang BR, Liu YS, Lin HY, Chen CC et al. Differential Characterization of Temozolomide-Resistant Human Glioma Cells. Int. J. Mol. Sci. 2018. 19: 127 - 231
Hatanpaa KJ, Burma S, Zhao D, Habib AA. Epidermal growth factor receptor in glioma: signal transduction, neuropathology, imaging, and radioresistance. Neoplasia, 2010;12(9):675-684.
Mellinghoff IK, Wang MY, Vivanco I, Haas-Kogan DA, Zhu S, Dia EQ, et al. Molecular determinants of the response of glioblastomas to EGFR kinase inhibitors. N Engl J Med. 2005;353(19):2012–24.
Wang Y, Pan L, Sheng XF, Chen S, Dai JZ. Nimotuzumab, a humanized monoclonal antibody specific for the EGFR, in combination with temolozolomide and radiation therapy for newly diagnosed glioblastoma multiforme: First results in Chinese Patient. Asia Pacific Journal of Clinical Oncology. 2016: 12: e23-e29.
Teng K, Zhang Y, Hu X, Ding Y, Gong R, Liu L. Nimotuzumab enhances radiation sensitivity of NSCLC H292 cells in vitro by blocking epidermal growth factor receptor nuclear translocation and inhibiting radiation-induced DNA damage repair. Onco Targets Ther. 2015; 8:809–18.
Akashi Y, Okamoto I, Iwasa T, Yoshida T, Suzuki M, Hatashita E, et al. Enhancement of the antitumor activity of ionising radiation by nimotuzumab, a humanised monoclonal antibody to the epidermal growth factor receptor, in non-small cell lung cancer cell lines of differing epidermal growth factor receptor status. Br J Cancer. 2008;98(4):749–55.
Garrido G, Tikhomirov IA, Rabasa A, Yang E, Gracia E, Iznaga N, et al. Bivalent binding by intermediate affinity of nimotuzumab: a contribution to explain antibody clinical profile. Cancer Biol Ther. 2011;11(4):373–82.
Perez R, Moreno E, Garrido G, Crombet T. EGFR-targeting as a biological therapy: understanding nimotuzumab’s clinical effects. Cancers. 2011;3(2):2014–31.
Diaz-Miqueli A, Martinez GS. Nimotuzumab as a radiosentizing agent in the treatment of highgrade glioma: challenge and opportunities. OncoTargets and Ther. 2013:6 931-942.
Weinstein IB, Joe A. Oncogene addiction. Cancer Res. 2008;68(9):3077–80.
Qu Y, Hu S, Xu X, Wang R, Yu H, Xu J, et al. Nimotuzumab enhances the radiosensitivity of cancer cells in vitro by inhibiting radiation-induced DNA damage repair. PLoS ONE. 2013;8(8):e70727.
Shen D, Guo CC, Wang J, Qiu ZK, Sai K et al. Interferon-?? enhances temozolomide activity against MGMT-positive glioma stem-like cell. Oncology Reports 2015: 34: 2715 – 2721.
Silber JR, Bobola MS, Blank A, Chamberlain MC. O6-methylguanine-DNA methyltransferase in glioma therapy: Promise and problems. Biochimica et Biophysica Acta. 2012;1826(1):71-82.
Chierico L, Rizzello L, Guan L, Joseph AS, Lewis A, Battaglia G. The role of the two splice variants and extranuclear pathway on Ki-67 regulation in non-cancer and cancer cells. Pal S, ed. PLoS ONE. 2017;12(2): e0171815.
Schmitz KR, Ferguson KM. Interaction of antibodies with ErbB receptor extracellular regions. Exp Cell Res. 2009;315(4):659–70.
Wee P, Wang Z. Epidermal Growth Factor Receptor Cell Proliferation Signaling Pathways. Mok SC, ed. Cancers. 2017;9(5):52.
Paul-Samojedny M, Suchanek R, Borkowska P, Pude?ko A, Owczarek A, Kowalczyk M, et al. Knockdown of AKT3 (PKB?) and PI3KCA Suppresses Cell Viability and Proliferation and Induces the Apoptosis of Glioblastoma Multiforme T98G Cells. BioMed Res Int. 2014; 2014:1–12.
Saurez-Martínez G, Bencomo-Yanes A. Nimotuzumab, effective immunotherapy for the treatment of malignant epithelial tumors. Biotecnol Apl. 2014; 31:159–67.
Cancer Genome Atlas Research Network. Comprehensive genomic characterization defines human glioblastoma genes and core pathways. Nature. 2008;455(7216):1061–8.
Crombet-Ramos T, Rak J, Pérez R, Viloria-Petit A. Antiproliferative, antiangiogenic and proapoptotic activity of h-R3: a humanized anti-EGFR antibody: a humanized anti-EGFR antibody. Int J Cancer. 2002;101(6):567–75.
Mazorra Z, Chao L, Lavastida A, Sanchez B, Ramos M. Nimotuzumab: beyond the EGFR signaling cascade inhibition. Oncology (2018): 45: 18-26
Petit AM, Rak J, Hung MC, Rockwell P, Goldstein N, Fendly B, et al. Neutralizing antibodies against epidermal growth factor and ErbB-2/neu receptor tyrosine kinases down-regulate vascular endothelial growth factor production by tumor cells in vitro and in vivo: angiogenic implications for signal transduction therapy of solid tumors. Am J Pathol. 1997;151(6):1523–30.
Hegedüs B, Czirók A, Fazekas I, Bábel T, Madarász E, Vicsek T. Locomotion and proliferation of glioblastoma cells in vitro: statistical evaluation of videomicroscopic observations. J Neurosurg. 2000;92(3):428–34.
Maier P, Hartmann L, Wenz F, Herskind. Cellular pathways in response to ionizing radiation and their targetability for tumor radiosensitization. Int J Mol Sci. 2016. 17:102
An Z, Aksoy O, Zheng T, Fan QW, Weiss WA. Epidermal growth factor resector (EGFR) and EGFRvIII in glioblastoma (GBM): signaling pathways and targeted therapy. Oncogene. 2018: 37(12): 1561 - 1575

[1] American Brain Tumor Association. About Brain Tumors: A Primer for Patients and Caregivers. In: About Brain Tumors: A Primer for Patients and Caregivers. American Brain Tumor Association; 2013. p. 76–8.

[2] Gardeck AM, Sheehan J, Low WC. Immune and Viral Therapies for Malignant Primary Brain Tumors. Exp Op on Biol Ther. 2017: 10.1080/14712598.2017.1296132.

[3] Lee SY. Temozolomide resistance in glioblastoma multiforme. Genes & Diseases. 2016: 3: 198 – 210

[4] Lai SW, Huang BR, Liu YS, Lin HY, Chen CC et al. Differential Characterization of Temozolomide-Resistant Human Glioma Cells. Int. J. Mol. Sci. 2018. 19: 127 - 231

[5] Hatanpaa KJ, Burma S, Zhao D, Habib AA. Epidermal growth factor receptor in glioma: signal transduction, neuropathology, imaging, and radioresistance. Neoplasia, 2010;12(9):675-684.

[6] Mellinghoff IK, Wang MY, Vivanco I, Haas-Kogan DA, Zhu S, Dia EQ, et al. Molecular determinants of the response of glioblastomas to EGFR kinase inhibitors. N Engl J Med. 2005;353(19):2012–24.

[7] Wang Y, Pan L, Sheng XF, Chen S, Dai JZ. Nimotuzumab, a humanized monoclonal antibody specific for the EGFR, in combination with temolozolomide and radiation therapy for newly diagnosed glioblastoma multiforme: First results in Chinese Patient. Asia Pacific Journal of Clinical Oncology. 2016: 12: e23-e29.

[8] Teng K, Zhang Y, Hu X, Ding Y, Gong R, Liu L. Nimotuzumab enhances radiation sensitivity of NSCLC H292 cells in vitro by blocking epidermal growth factor receptor nuclear translocation and inhibiting radiation-induced DNA damage repair. Onco Targets Ther. 2015; 8:809–18.

[9] Akashi Y, Okamoto I, Iwasa T, Yoshida T, Suzuki M, Hatashita E, et al. Enhancement of the antitumor activity of ionising radiation by nimotuzumab, a humanised monoclonal antibody to the epidermal growth factor receptor, in non-small cell lung cancer cell lines of differing epidermal growth factor receptor status. Br J Cancer. 2008;98(4):749–55.

[10] Garrido G, Tikhomirov IA, Rabasa A, Yang E, Gracia E, Iznaga N, et al. Bivalent binding by intermediate affinity of nimotuzumab: a contribution to explain antibody clinical profile. Cancer Biol Ther. 2011;11(4):373–82.

[11] Perez R, Moreno E, Garrido G, Crombet T. EGFR-targeting as a biological therapy: understanding nimotuzumab’s clinical effects. Cancers. 2011;3(2):2014–31.

[12] Diaz-Miqueli A, Martinez GS. Nimotuzumab as a radiosentizing agent in the treatment of highgrade glioma: challenge and opportunities. OncoTargets and Ther. 2013:6 931-942.

[13] Weinstein IB, Joe A. Oncogene addiction. Cancer Res. 2008;68(9):3077–80.

[14] Qu Y, Hu S, Xu X, Wang R, Yu H, Xu J, et al. Nimotuzumab enhances the radiosensitivity of cancer cells in vitro by inhibiting radiation-induced DNA damage repair. PLoS ONE. 2013;8(8):e70727.

[15] Shen D, Guo CC, Wang J, Qiu ZK, Sai K et al. Interferon-?? enhances temozolomide activity against MGMT-positive glioma stem-like cell. Oncology Reports 2015: 34: 2715 – 2721.

[16] Silber JR, Bobola MS, Blank A, Chamberlain MC. O6-methylguanine-DNA methyltransferase in glioma therapy: Promise and problems. Biochimica et Biophysica Acta. 2012;1826(1):71-82.

[17] Chierico L, Rizzello L, Guan L, Joseph AS, Lewis A, Battaglia G. The role of the two splice variants and extranuclear pathway on Ki-67 regulation in non-cancer and cancer cells. Pal S, ed. PLoS ONE. 2017;12(2): e0171815.

[18] Schmitz KR, Ferguson KM. Interaction of antibodies with ErbB receptor extracellular regions. Exp Cell Res. 2009;315(4):659–70.

[19] Wee P, Wang Z. Epidermal Growth Factor Receptor Cell Proliferation Signaling Pathways. Mok SC, ed. Cancers. 2017;9(5):52.

[20] Paul-Samojedny M, Suchanek R, Borkowska P, Pude?ko A, Owczarek A, Kowalczyk M, et al. Knockdown of AKT3 (PKB?) and PI3KCA Suppresses Cell Viability and Proliferation and Induces the Apoptosis of Glioblastoma Multiforme T98G Cells. BioMed Res Int. 2014; 2014:1–12.

[21] Saurez-Martínez G, Bencomo-Yanes A. Nimotuzumab, effective immunotherapy for the treatment of malignant epithelial tumors. Biotecnol Apl. 2014; 31:159–67.

[22] Cancer Genome Atlas Research Network. Comprehensive genomic characterization defines human glioblastoma genes and core pathways. Nature. 2008;455(7216):1061–8.

[23] Crombet-Ramos T, Rak J, Pérez R, Viloria-Petit A. Antiproliferative, antiangiogenic and proapoptotic activity of h-R3: a humanized anti-EGFR antibody: a humanized anti-EGFR antibody. Int J Cancer. 2002;101(6):567–75.

[24] Mazorra Z, Chao L, Lavastida A, Sanchez B, Ramos M. Nimotuzumab: beyond the EGFR signaling cascade inhibition. Oncology (2018): 45: 18-26

[25] Petit AM, Rak J, Hung MC, Rockwell P, Goldstein N, Fendly B, et al. Neutralizing antibodies against epidermal growth factor and ErbB-2/neu receptor tyrosine kinases down-regulate vascular endothelial growth factor production by tumor cells in vitro and in vivo: angiogenic implications for signal transduction therapy of solid tumors. Am J Pathol. 1997;151(6):1523–30.

[26] Hegedüs B, Czirók A, Fazekas I, Bábel T, Madarász E, Vicsek T. Locomotion and proliferation of glioblastoma cells in vitro: statistical evaluation of videomicroscopic observations. J Neurosurg. 2000;92(3):428–34.

[27] Maier P, Hartmann L, Wenz F, Herskind. Cellular pathways in response to ionizing radiation and their targetability for tumor radiosensitization. Int J Mol Sci. 2016. 17:102

[28] An Z, Aksoy O, Zheng T, Fan QW, Weiss WA. Epidermal growth factor resector (EGFR) and EGFRvIII in glioblastoma (GBM): signaling pathways and targeted therapy. Oncogene. 2018: 37(12): 1561 - 1575

Growth suppression of glioblastoma multiforme cell line response to the combination of Temozolomide and Nimotuzumab

Abstract

References

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