Impact of AKT1 on cell invasion and radiosensitivity in a triple negative breast cancer cell line developing brain metastasis

Joanna Kempska; Leticia Oliveira-Ferrer; Astrid Grottke; Minyue Qi; Malik Alawi; Felix Meyer; Kerstin Borgmann; Fabienne Hamester; Kathrin Eylmann; Maila Rossberg; Daniel J Smit; Manfred Jücker; Elena Laakmann; Isabell Witzel; Barbara Schmalfeldt; Volkmar Müller; Karen Legler

doi:10.3389/fonc.2023.1129682

Impact of AKT1 on cell invasion and radiosensitivity in a triple negative breast cancer cell line developing brain metastasis

Front Oncol. 2023 Jul 6:13:1129682. doi: 10.3389/fonc.2023.1129682. eCollection 2023.

Authors

Joanna Kempska¹, Leticia Oliveira-Ferrer¹, Astrid Grottke¹, Minyue Qi², Malik Alawi², Felix Meyer³, Kerstin Borgmann³, Fabienne Hamester¹, Kathrin Eylmann¹, Maila Rossberg¹, Daniel J Smit⁴, Manfred Jücker⁴, Elena Laakmann¹, Isabell Witzel¹, Barbara Schmalfeldt¹, Volkmar Müller¹, Karen Legler¹

Affiliations

¹ Department of Gynecology, University Medical Center Hamburg-Eppendorf, Hamburg, Germany.
² Bioinformatics Core, University Medical Center Hamburg-Eppendorf, Hamburg, Germany.
³ Laboratory of Radiobiology & Experimental Radio Oncology, Department of Radiotherapy and Radiation Oncology, University Medical Center Hamburg-Eppendorf, Hamburg, Germany.
⁴ Institute of Biochemistry and Signal Transduction, University Medical Center Hamburg-Eppendorf, Hamburg, Germany.

Abstract

Introduction: The PI3K/AKT pathway is activated in 43-70% of breast cancer (BC)-patients and promotes the metastatic potential of BC cells by increasing cell proliferation, invasion and radioresistance. Therefore, AKT1-inhibition in combination with radiotherapy might be an effective treatment option for triple-negative breast cancer (TNBC)-patients with brain metastases.

Methods: The impact of AKT1-knockout (AKT1_KO) and AKT-inhibition using Ipatasertib on MDA-MB-231 BR cells was assessed using in vitro cell proliferation and migration assays. AKT1-knockout in MDA-MB-231BR cells was performed using CRISPR/Cas9. The effect of AKT1-knockout on radiosensitivity of MDA-MB-231BR cell lines was determined via colony formation assays after cell irradiation. To detect genomic variants in AKT1_KO MDA-MB-231BR cells, whole-genome sequencing (WGS) was performed.

Results: Pharmacological inhibition of AKT with the pan-AKT inhibitor Ipatasertib led to a significant reduction of cell viability but did not impact cell migration. Moreover, only MDA-MB-231BR cells were sensitized following Ipatasertib-treatment. Furthermore, specific AKT1-knockout in MDA-MB-231BR showed reduced cell viability in comparison to control cells, with significant effect in one of two analyzed clones. Unexpectedly, AKT1 knockout led to increased cell migration and clonogenic potential in both AKT1_KO clones. RNAseq-analysis revealed the deregulation of CTSO, CYBB, GPR68, CEBPA, ID1, ID4, METTL15, PBX1 and PTGFRN leading to the increased cell migration, higher clonogenic survival and decreased radiosensitivity as a consequence of the AKT1 knockout in MDA-MB-231BR.

Discussion: Collectively, our results demonstrate that Ipatasertib leads to radiosensitization and reduced cell proliferation of MDA-MB-231BR. AKT1-inhibition showed altered gene expression profile leading to modified cell migration, clonogenic survival and radioresistance in MDA-MB-231BR. We conclude, that AKT1-inhibition in combination with radiotherapy contribute to novel treatment strategies for breast cancer brain metastases.

Keywords: AKT1; Ipatasertib; brain metastases; breast cancer; radioresistance.

Grants and funding

The authors declare that this study received funding from Genentech, Inc. The funder was not involved in the study design, collection, analysis, interpretation of data, the writing of this article or the decision to submit it for publication. This work was also supported by the Gynecology Department of the University Medical Center Hamburg-Eppendorf, Germany.