The Role and Mechanism of CRT0066101 as an Effective Drug for Treatment of Triple-Negative Breast Cancer.

Abstract

BACKGROUND/AIMS:

Breast cancer is clinically classified into three main subtypes: estrogen receptor-positive (ER+) breast cancer, human epidermal growth factor receptor 2-positive (HER2+) breast cancer, and triple-negative breast cancer (TNBC). Without specific targeted therapies, patients with TNBC have poorer prognosis compared with those with ER+ and HER2+ breast cancer. Protein kinase D (PRKD) family members play crucial roles in cancer progression. CRT0066101, a PRKD inhibitor, has been reported to have anticancer activity in many cancer types. Nevertheless, the role and mechanism of CRT0066101 in TNBC have not been well investigated.

METHODS:

The expression level of PRKDs was analyzed in breast cancer samples and breast cancer cell lines. The effects of inhibiting PRKD activity with CRT0066101 on TNBC cell proliferation, cell cycle, apoptosis, and tumor growth were studied by Cell Counting Kit8 assay, cell cycle assay, propidium iodide/annexin-V assay, and a xenograft mouse model, respectively. To uncover the molecular mechanism of CRT0066101 in TNBC, comparative phosphoproteomic analysis using iTRAQ was employed.

RESULTS:

We found that PRKD2 and PRKD3 were preferentially expressed in breast cancers. Immunohistochemistry confirmed the overexpression of PRKD2 and PRKD3 in TNBC. CRT0066101, which inhibited the activity of PRKDs, dramatically inhibited proliferation, increased apoptosis and the G1-phase population of TNBC cells in vitro, and reduced breast tumor volume in vivo. Comparative phosphoproteomic analysis between breast cancer cells with and without CRT0066101 treatment revealed that the anti-breast cancer effects involved regulation of a complex network containing multiple enriched pathways and several hub-nodes contributing to multiple cancer-related processes, thus explaining the described effects of CRT0066101 on TNBC in vitro and in vivo. Finally, we validated several targets of PRKD inhibition by treatment with CRT0066101 and small interfering RNAs against PRKD2 and PRKD3 (siPRKD2 and siPRKD3), including p-MYC(T58/ S62), p-MAPK1/3(T202/Y204), p-AKT(S473), p-YAP(S127), and p-CDC2(T14).

CONCLUSION:

PRKD inhibitor CRT0066101 exhibits anti-TNBC effects via modulating a phosphor-signaling network and inhibiting the phosphorylation of many cancer-driving factors, including MYC, MAPK1/3, AKT, YAP, and CDC2, providing insight into the important roles as well as the molecular mechanism of CRT0066101 as an effective drug for TNBC.

• Authors: Yan Liu, Yuzhi Wang, Shiyi Yu, Yehui Zhou, Xinxing Ma, Qian Su, Li An, Feifei Wang, Aihua Shi, Jingzhong Zhang, Liming Chen
• Journal: Cellular physiology and biochemistry : international journal of experimental cellular physiology, biochemistry, and pharmacology (Cell Physiol Biochem) Vol. 52 Issue 3 Pg. 382-396 ( 2019) ISSN: 1421-9778 [Electronic] Germany
• PMID: 30845378 (Publication Type: Journal Article)
• Copyright: © Copyright by the Author(s). Published by Cell Physiol Biochem Press.

Chemical References

• CRT 0066101
• Phosphopeptides
• Proto-Oncogene Proteins c-myc
• Pyrimidines
• RNA, Small Interfering
• protein kinase D
• Proto-Oncogene Proteins c-akt
• Protein Kinase C

Topics

• Animals
• Apoptosis (drug effects)
• Cell Line, Tumor
• Cell Proliferation (drug effects)
• Female
• G1 Phase Cell Cycle Checkpoints (drug effects)
• Humans
• Mice
• Mice, Nude
• Phosphopeptides (analysis)
• Protein Kinase C (antagonists & inhibitors, genetics, metabolism)
• Proteomics
• Proto-Oncogene Proteins c-akt (genetics, metabolism)
• Proto-Oncogene Proteins c-myc (genetics, metabolism)
• Pyrimidines (metabolism, pharmacology, therapeutic use)
• RNA Interference
• RNA, Small Interfering (metabolism)
• Triple Negative Breast Neoplasms (drug therapy, metabolism, pathology)