Breast cancers that disseminate to bone marrow acquire aggressive phenotypes through CX43-related tumor-stroma tunnels
Saptarshi Sinha, Brennan W. Callow, Alex P. Farfel, Suchismita Roy, Siyi Chen, Maria Masotti, Shrila Rajendran, Johanna M. Buschhaus, Celia R. Espinoza, Kathryn E. Luker, Pradipta Ghosh, Gary D. Luker
Saptarshi Sinha, Brennan W. Callow, Alex P. Farfel, Suchismita Roy, Siyi Chen, Maria Masotti, Shrila Rajendran, Johanna M. Buschhaus, Celia R. Espinoza, Kathryn E. Luker, Pradipta Ghosh, Gary D. Luker
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Research Article Oncology

Breast cancers that disseminate to bone marrow acquire aggressive phenotypes through CX43-related tumor-stroma tunnels

Abstract

Estrogen receptor-positive (ER+) breast cancer commonly disseminates to bone marrow, where interactions with mesenchymal stromal cells (MSCs) shape disease trajectory. We modeled these interactions with tumor-MSC co-cultures and used an integrated transcriptome-proteome-network-analyses workflow to identify a comprehensive catalog of contact-induced changes. Conditioned media from MSCs failed to recapitulate genes and proteins, some borrowed and others tumor-intrinsic, induced in cancer cells by direct contact. Protein-protein interaction networks revealed the rich connectome between “borrowed” and “intrinsic” components. Bioinformatics prioritized one of the borrowed components, CCDC88A/GIV, a multi-modular metastasis-related protein that has recently been implicated in driving a hallmark of cancer, growth signaling autonomy. MSCs transferred GIV protein to ER+ breast cancer cells (that lack GIV) through tunnelling nanotubes via connexin (Cx)43-facilitated intercellular transport. Reinstating GIV alone in GIV-negative breast cancer cells reproduced approximately 20% of both the borrowed and the intrinsic gene induction patterns from contact co-cultures; conferred resistance to anti-estrogen drugs; and enhanced tumor dissemination. Findings provide a multiomic insight into MSC→tumor cell intercellular transport and validate how transport of one such candidate, GIV, from the haves (MSCs) to have-nots (ER+ breast cancer) orchestrates aggressive disease states.

Authors

Saptarshi Sinha, Brennan W. Callow, Alex P. Farfel, Suchismita Roy, Siyi Chen, Maria Masotti, Shrila Rajendran, Johanna M. Buschhaus, Celia R. Espinoza, Kathryn E. Luker, Pradipta Ghosh, Gary D. Luker

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Figure 8

Expression of GIV recapitulates the MSC close contact signature, confers resistance to ER-targeted therapies, and promotes early dissemination of ER+ breast cancer cells.

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Expression of GIV recapitulates the MSC close contact signature, confers...
(A) Schematic shows creation of a stable MCF7 cell population expressing exogenous GIV (CCDC88A) by a piggyBac transposase vector. (B) Immunoblots of whole cell lysates of cells in A, confirming expression of full-length GIV. (C and D) Workflow (C) of RNA sequencing and normalized gene expression analysis of cells in B analyzed for the borrowed (n = 158) and intrinsic (n = 76) genes to perfectly classify MCF7-GIV cells from control MCF7 (ROC-AUC 1.00). Venn diagram (D) shows genes induced with GIV alone as percentage of the total borrowed and intrinsic signatures. (E and F) Heatmaps display hierarchical unsupervised clustering of MCF7 and MCF7-GIV cells by z score–normalized gene expression for the subset of genes induced among the borrowed (E) and intrinsic (F) signatures. Supplemental Figure 6 shows Reactome pathway analysis of these genes. (G–I) Graphs display viability for MCF7-GIV and control MCF7 cells exposed to increasing concentrations of tamoxifen (G), fulvestrant (H), or fulvestrant and 100 nM palpociclib (I). Data are displayed after normalization to cells treated with vehicle only. (J and K) Equal number (1 × 105) of MCF7-GIV or control MCF7 cells were injected into the left cardiac ventricle of 7- to 10-week-old female NSG mice (n = 8 per group). Representative bioluminescence images (J) show mice at days 1, 3, and 6 after injection. In G–I and K, graphs show mean values ± SEM of each group at specific data points. Statistical significance between 2 groups at each time point was computed using repeated measures ANOVA. *P < 0.1, **P ≤ 0.05, ***P ≤ 0.01 indicates corrected P values using Tukey’s method.