The simultaneous use of CRISPR/Cas9 to knock out the PI3Kca gene with radiation to enhance radiosensitivity and inhibit tumor growth in breast cancer

Document Type : Original Article

Authors

1 Department of Medical Physics, School of Medicine, Ahvaz Jundishapur University of Medical Sciences, Ahvaz, Iran

2 Cellular and Molecular Research Center, Medical Basic Sciences Research Institute, Ahvaz Jundishapur University of Medical Sciences, Ahvaz, Iran

3 Department of Virology, Faculty of Medicine, Hamedan University of Medical Sciences, Hamedan, Iran

4 Department of Virology, School of Medicine, Ahvaz Jundishapur University of Medical Sciences, Ahvaz, Iran

Abstract

Objective(s): Breast neoplasm is a malignancy that can have a poor prognosis. The PI3K/AKT signaling pathway is frequently activated in various tumor types, including breast cancer, leading to alterations in the tumor microenvironment and radioresistance. Selective inhibition of PI3Kca (p110α) has been considered an alternative approach to overcome radioresistance, owing to concerns surrounding the excessive side effects of pan-PI3K inhibitors tested in clinical trials. This investigation aimed to evaluate the efficacy of co-administering PI3Kca knocking out with radiation therapy in mitigating radioresistance and suppressing tumor growth in the MDA-MB-231 cell line. 
Materials and Methods: The present investigation utilized the CRISPR/Cas9 technique to induce a knockout of the PI3Kca gene. Subsequently, after 24 hr of transfection, gene expression, cell proliferation, apoptosis rate, and angiogenesis were assessed.
Results: We demonstrated that knocking out PI3Kca, in combination with radiation, increased apoptosis, reduced the expression of PI3Kca and AKT1 genes, and decreased cell proliferation. The CAM assay analysis has demonstrated that knocking out the PI3Kca gene and radiotherapy substantially reduced the total vessel network length and the number of junctions.
Conclusion: The findings of our investigation indicate that the integration of radiation therapy with PI3Kca yielded enhanced radiosensitivity, leading to a marked retardation of tumor progression and an increased survival rate. 

Keywords

Main Subjects

References

1. Foulkes WD, Smith IE, Reis-Filho JS. Triple-negative breast cancer. N Engl J Med 2010; 363: 1938-1948.
2. Anderson WF, Rosenberg PS, Prat A, Perou CM, Sherman ME. How many etiological subtypes of breast cancer: Two, three, four, or more? J Natl Cancer Inst  2014; 106: dju165.
3. Wahba HA, El-Hadaad HA. Current approaches in treatment of triple-negative breast cancer. Cancer Biol Med 2015; 12: 106-116.
4. Zhao L, Vogt PK. Class I PI3K in oncogenic cellular transformation. Oncogene 2008; 27: 5486-5496.
5. Saal LH, Holm K, Maurer M, Memeo L, Su T, Wang X, et al. PIK3CA mutations correlate with hormone receptors, node metastasis, and ERBB2, and are mutually exclusive with PTEN loss in human breast carcinoma. Cancer Res 2005; 65: 2554-2559.
6. Miricescu D, Totan A, Stanescu-Spinu I-I, Badoiu SC, Stefani C, Greabu M. PI3K/AKT/mTOR signaling pathway in breast cancer: from molecular landscape to clinical aspects. Int J Mol Sci 2020; 22: 173-196.
7. Martínez-Sáez O, Chic N, Pascual T, Adamo B, Vidal M, González-Farré B, et al. Frequency and spectrum of PIK3CA somatic mutations in breast cancer. Breast Cancer Res 2020; 22: 1-9.
8. Fruman DA, Chiu H, Hopkins BD, Bagrodia S, Cantley LC, Abraham RT. The PI3K pathway in human disease. Cell 2017; 170: 605-635.
9. Engelman JA. Targeting PI3K signalling in cancer: Opportunities, challenges and limitations. Nat Rev Cancer 2009; 9: 550-562.
10. Guo S, Loibl S, von Minckwitz G, Darb-Esfahani S, Lederer B, Denkert C. PIK3CA H1047R mutation associated with a lower pathological complete response rate in triple-negative breast cancer patients treated with anthracycline-taxane-based neoadjuvant chemotherapy. Cancer Res Treat 2020; 52: 689-696.
11. Marra A, Trapani D, Viale G, Criscitiello C, Curigliano G. Practical classification of triple-negative breast cancer: Intratumoral heterogeneity, mechanisms of drug resistance, and novel therapies. NPJ Breast Cancer 2020; 6: 54-69.
12. Di Cosimo S, Appierto V, Silvestri M, Pruneri G, Vingiani A, Perrone F, et al. Targeted-gene sequencing to catch triple negative breast cancer heterogeneity before and after neoadjuvant chemotherapy. Cancers 2019; 11: 1753-1766.
13. Behbahani R, Danyaei A, Teimoori A, Neisi N, Tahmasbi M. Breast cancer radioresistance may be overcome by osteopontin gene knocking out with CRISPR/Cas9 technique. Cancer Radiother 2021; 25: 222-228.
14. Lepekhin EA, Walmod PS, Berezin A, Berezin V, Bock E. Cytoskeleton Methods Protoc 2001: 85-100.
15. Wan G, Mahajan A, Lidke D, Rajput A. Bcl-2 together with PI3K p110α regulates cell morphology and cell migration. Cell Death Dis 2015; 6: e2006.
16. Alqahtani A, Ayesh HS, Halawani H. PIK3CA gene mutations in solid malignancies: Association with clinicopathological parameters and prognosis. Cancers 2019; 12: 93-110.
17.    Ghanbarnasab Behbahani R, Danyaei A, Teimoori A, Tahmasbi MJ, Neisi N. CRISPR/Cas9 mediated knocking out of OPN gene enhances radiosensitivity in MDA-MB-231 breast cancer cell line. J Cancer Res Clin Oncol 2023; 149: 4117-4130.
18.    Bai X, Zhang E, Ye H, Nandakumar V, Wang Z, Chen L, et al. PIK3CA and TP53 gene mutations in human breast cancer tumors frequently detected by ion torrent DNA sequencing. PLoS One 2014; 9: e99306.
19.    Johnson J, Chow Z, Napier D, Lee E, Weiss HL, Evers BM, et al. Targeting PI3K and AMPKα signaling alone or in combination to enhance radiosensitivity of triple negative breast cancer. Cells 2020; 9: 1253-1268.
20. Shi Q, Li Y, Li S, Jin L, Lai H, Wu Y, et al. LncRNA DILA1 inhibits cyclin D1 degradation and contributes to tamoxifen resistance in breast cancer. Nat Commun 2020; 11: 5513-5527.
21. Carpten JD, Faber AL, Horn C, Donoho GP, Briggs SL, Robbins CM, et al. A transforming mutation in the pleckstrin homology domain of AKT1 in cancer. Nature 2007; 448: 439-444.
22. Wang X, Tu W, Chen D, Fu J, Wang J, Shao C, et al. Autophagy suppresses radiation damage by activating PARP-1 and attenuating reactive oxygen species in hepatoma cells. Int J Radiat Biol 2019; 95: 1051-1057.
23. Seol MY, Choi SH, Yoon HI. Combining radiation with PI3K isoform-selective inhibitor administration increases radiosensitivity and suppresses tumor growth in non-small cell lung cancer. J Radiat Res 2022; 63: 591-601.
24. Seol MY, Choi SH, Lee IJ, Park HS, Kim HR, Kim SK, et al. Selective inhibition of PI3K isoforms in brain tumors suppresses tumor growth by increasing radiosensitivity. Yonsei Med J 2023; 64: 139-147.
25. Zhang L, Li Y, Wang Q, Chen Z, Li X, Wu Z, et al. The PI3K subunits, P110α and P110β are potential targets for overcoming P-gp and BCRP-mediated MDR in cancer. Mol Cancer 2020; 19: 1-18.
26. Sun Y, Ning X, Fan J, Hu J, Jiang Y, Hu Z, et al. Loss of tumor suppressor inositol polyphosphate 4-phosphatase type B impairs DNA double-strand break repair by destabilization of DNA tethering protein Rad50. Cell Death Dis 2020; 11: 292-301.
27. Hu H,  Zhu J, Zhong Y, Geng R, Ji Y, et al. PIK3CA mutation confers resistance to chemotherapy in triple-negative breast cancer by inhibiting apoptosis and activating the PI3K/AKT/mTOR signaling pathway. Ann Transl Med 2021; 9: 410-424.
Iranian Journal of Basic Medical Sciences
Volume 27, Issue 12
December 2024
Pages 1566-1573

Files

Share

How to cite

Statistics