American Association for Cancer Research
mct-21-0783_supplementary_figure_s3_suppsf3.pdf (224.51 kB)

Supplementary Figure S3 from Targeting NANOG and FAK via Cx26-derived Cell-penetrating Peptides in Triple-negative Breast Cancer

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journal contribution
posted on 2024-01-03, 08:20 authored by Erin E. Mulkearns-Hubert, Emily Esakov Rhoades, Salma Ben-Salem, Rashmi Bharti, Nicole Hajdari, Sadie Johnson, Alex Myers, Iris Nira Smith, Smarajit Bandyopadhyay, Charis Eng, Erinn Downs, Justin D. Lathia, Ofer Reizes

SPR sensograms demonstrating no binding of Cx26 extracellular loop peptides to FAK or NANOG proteins.


Congressionally Directed Medical Research Programs (CDMRP)

National Center for Advancing Translational Sciences (NCATS)

United States Department of Health and Human Services

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Triple-negative breast cancer (TNBC) represents the most lethal and treatment-resistant breast cancer subtype with limited treatment options. We previously identified a protein complex unique to TNBC composed of the gap junction protein connexin 26 (Cx26), the pluripotency transcription factor NANOG, and focal adhesion kinase (FAK). We sought to determine whether a peptide mimetic of the interaction region of Cx26 attenuated tumor growth in preclinical models. We designed peptides based on Cx26 juxtamembrane domains and performed binding experiments with NANOG and FAK using surface plasmon resonance. Binding studies revealed that the Cx26 C-terminal tail and intracellular loop bound to NANOG and FAK with submicromolar-to-micromolar affinity and that a 5-amino acid sequence in the C-terminal tail of Cx26 (RYCSG) was sufficient for binding. Peptides with high affinity were engineered with a cell-penetrating antennapedia sequence and assessed in functional assays including cell proliferation, tumorsphere formation, and in vivo tumor growth, and downstream signaling changes were measured. The cell-penetrating Cx26 peptide (aCx26-pep) disrupted self-renewal while reducing nuclear FAK and NANOG and inhibiting NANOG target gene expression in TNBC cells but not luminal mammary epithelial cells. In vivo, aCx26-pep reduced tumor growth and proliferation and induced cell death. Here, we provide proof-of-concept that a Cx26 peptide–based strategy inhibits growth and alters NANOG activity specifically in TNBC, indicating the therapeutic potential of this targeting approach.

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