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TrkA expression directs the anti-neoplastic activity of MLK3 inhibitors in triple-negative breast cancer

Oncogene volume 42, pages 1132–1143 (2023)Cite this article

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Abstract

Mixed Lineage Kinase 3 (MLK3) is a viable target for neoplastic diseases; however, it is unclear whether its activators or inhibitors can act as anti-neoplastic agents. We reported that the MLK3 kinase activity was higher in triple-negative (TNBC) than in hormone receptor-positive human breast tumors, where estrogen inhibited MLK3 kinase activity and provided a survival advantage to ER+ breast cancer cells. Herein, we show that in TNBC, the higher MLK3 kinase activity paradoxically promotes cancer cell survival. Knockdown of MLK3 or MLK3 inhibitors, CEP-1347 and URMC-099, attenuated tumorigenesis of TNBC cell line and Patient-Derived (PDX) xenografts. The MLK3 kinase inhibitors decreased both the expression and activation of MLK3, PAK1, and NF-kB protein and caused cell death in TNBC breast xenografts. RNA-seq analysis identified several genes downregulated by MLK3 inhibition, and the NGF/TrkA MAPK pathway was significantly enriched in tumors sensitive to growth inhibition by MLK3 inhibitors. The TNBC cell line unresponsive to kinase inhibitor had substantially lower TrkA, and overexpression of TrkA restored the sensitivity to MLK3 inhibition. These results suggest that the functions of MLK3 in breast cancer cells depend on downstream targets in TNBC tumors expressing TrkA, and MLK3 kinase inhibition may provide a novel targeted therapy.

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Fig. 1: MLK3, PAK1, and NF-ĸB activities directly correlate in human breast tumors.
Fig. 2: MLK3 is necessary for TNBC tumorigenesis.
Fig. 3: Pharmacological inhibitors of MLK3 decrease MLK3, PAK1, p-PAK1, NF-ĸB, and p-NF-ĸB protein expression and induce cell death in TNBC cell lines.
Fig. 4: Pharmacological inhibitors of MLK3 impair TNBC tumorigenesis.
Fig. 5: The NGF/TrkA MAPK pathway is suppressed in TNBC PDX tumors in response to MLK3 inhibitors.
Fig. 6: TrkA expression sensitizes MDA-MB-231 cells to MLK3 inhibitor.
Fig. 7: Both MLK3 and TrkA are necessary for MLK3 inhibitors’ action.

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Data availability

The RNA-Seq data discussed in this publication have been deposited in NCBI’s Gene Expression Omnibus (GEO) and can be accessed through accession number GSE188947. The corresponding author will fulfill additional requests for information or data upon request.

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Acknowledgements

We acknowledge funding support from National Cancer Institute to AR (CA216410 and CA176846), BR (CA219764), and the UIC-Cancer Center pilot grant award to BR. This work is partly supported by Veterans Affairs Merit Award (BX004903), and Career Scientist Award (BX004855) to AR, and VA-Merit to BR (BX003296 and BX005791). The contents of this article are the responsibility of the authors and do not represent the views of the Department of Veterans Affairs or the United States Government. The RNA sequencing and analysis were performed by the UIC Research Informatics Core, supported partly by NCATS through Grant UL1TR002003.

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Author notes

  1. These authors contributed equally: Rakesh Sathish Nair, Sandeep Kumar.

Authors and Affiliations

  1. Department of Surgery, Division of Surgical Oncology, The University of Illinois at Chicago, Chicago, IL, 60612, USA

    Rakesh Sathish Nair, Sandeep Kumar, Subhasis Das, Sunil Kumar Singh, Piush Srivastava, Gautam Sondarva, Basabi Rana & Ajay Rana

  2. University of Illinois Hospital & Health Sciences System Cancer Center, The University of Illinois at Chicago, Chicago, IL, 60612, USA

    Subhasis Das, Basabi Rana & Ajay Rana

  3. Department of Pathology and Laboratory Medicine, Baylor Scott & White Medical Center, Temple, TX, USA

    Arundhati Rao

  4. Weill Cornell Medicine, New York, NY, 10021, USA

    Subhash C. Sinha

  5. UICentre for Drug Discovery, University of Illinois at Chicago, Chicago, IL, 60612, USA

    Rui Xiong & Laura Bloem

  6. Department of Medicine, The University of Illinois at Chicago, Chicago, IL, 60612, USA

    Kent Hoskins

  7. Department of Pharmacology and Toxicology, Coit College of Pharmacy, University of Arizona, Tucson, AZ, 85721, USA

    Gregory R. J. Thatcher

  8. Jesse Brown VA Medical Center, Chicago, IL, 60612, USA

    Basabi Rana & Ajay Rana

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Contributions

AR conceived the idea; AR, SK, RSN, and SD designed the experiments. RSN, SK, SD, SKS, PS, and GS generated new reagents and performed experiments. SCS initially synthesized MLK3 inhibitors, and RX analyzed the RNA-Seq data. AR, LB, KH, GRT, and BR participated in the discussion and interpretation of data. AR and SK wrote the manuscript.

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Correspondence to Ajay Rana.

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Nair, R.S., Kumar, S., Das, S. et al. TrkA expression directs the anti-neoplastic activity of MLK3 inhibitors in triple-negative breast cancer. Oncogene 42, 1132–1143 (2023). https://doi.org/10.1038/s41388-023-02633-6

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