PIK3Cδ expression by fibroblasts promotes triple-negative breast cancer progression
Teresa Gagliano, Kalpit Shah, Sofia Gargani, Liyan Lao, Mansour Alsaleem, Jianing Chen, Vasileios Ntafis, Penghan Huang, Angeliki Ditsiou, Viviana Vella, Kritika Yadav, Kamila Bienkowska, Giulia Bresciani, Kai Kang, Leping Li, Philip Carter, Graeme Benstead-Hume, Timothy O’Hanlon, Michael Dean, Frances M.G. Pearl, Soo-Chin Lee, Emad A. Rakha, Andrew R. Green, Dimitris L. Kontoyiannis, Erwei Song, Justin Stebbing, Georgios Giamas
Teresa Gagliano, Kalpit Shah, Sofia Gargani, Liyan Lao, Mansour Alsaleem, Jianing Chen, Vasileios Ntafis, Penghan Huang, Angeliki Ditsiou, Viviana Vella, Kritika Yadav, Kamila Bienkowska, Giulia Bresciani, Kai Kang, Leping Li, Philip Carter, Graeme Benstead-Hume, Timothy O’Hanlon, Michael Dean, Frances M.G. Pearl, Soo-Chin Lee, Emad A. Rakha, Andrew R. Green, Dimitris L. Kontoyiannis, Erwei Song, Justin Stebbing, Georgios Giamas
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Research Article Cell biology Oncology

PIK3Cδ expression by fibroblasts promotes triple-negative breast cancer progression

Abstract

As there is growing evidence for the tumor microenvironment’s role in tumorigenesis, we investigated the role of fibroblast-expressed kinases in triple-negative breast cancer (TNBC). Using a high-throughput kinome screen combined with 3D invasion assays, we identified fibroblast-expressed PIK3Cδ (f-PIK3Cδ) as a key regulator of cancer progression. Although PIK3Cδ was expressed in primary fibroblasts derived from TNBC patients, it was barely detectable in breast cancer (BC) cell lines. Genetic and pharmacological gain- and loss-of-function experiments verified the contribution of f-PIK3Cδ in TNBC cell invasion. Integrated secretomics and transcriptomics analyses revealed a paracrine mechanism via which f-PIK3Cδ confers its protumorigenic effects. Inhibition of f-PIK3Cδ promoted the secretion of factors, including PLGF and BDNF, that led to upregulation of NR4A1 in TNBC cells, where it acts as a tumor suppressor. Inhibition of PIK3Cδ in an orthotopic BC mouse model reduced tumor growth only after inoculation with fibroblasts, indicating a role of f-PIK3Cδ in cancer progression. Similar results were observed in the MMTV-PyMT transgenic BC mouse model, along with a decrease in tumor metastasis, emphasizing the potential immune-independent effects of PIK3Cδ inhibition. Finally, analysis of BC patient cohorts and TCGA data sets identified f-PIK3Cδ (protein and mRNA levels) as an independent prognostic factor for overall and disease-free survival, highlighting it as a therapeutic target for TNBC.

Authors

Teresa Gagliano, Kalpit Shah, Sofia Gargani, Liyan Lao, Mansour Alsaleem, Jianing Chen, Vasileios Ntafis, Penghan Huang, Angeliki Ditsiou, Viviana Vella, Kritika Yadav, Kamila Bienkowska, Giulia Bresciani, Kai Kang, Leping Li, Philip Carter, Graeme Benstead-Hume, Timothy O’Hanlon, Michael Dean, Frances M.G. Pearl, Soo-Chin Lee, Emad A. Rakha, Andrew R. Green, Dimitris L. Kontoyiannis, Erwei Song, Justin Stebbing, Georgios Giamas

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Figure 7

Effects of fibroblast PIK3Cδ inhibition on TNBC tumor growth in vivo.

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Effects of fibroblast PIK3Cδ inhibition on TNBC tumor growth in vivo. (A...
(A) Schematic representation of the in vivo experiment using NOD CB17 PRKDC/J mice. MDA-MB-231 (groups 1 and 2) and MDA-MB-231/MRC5 (groups 3 and 4) tumor cells were implanted s.c. on day 0. After randomization on day 7, treatment with CAL-101 was initiated in groups 2 and 4, whereas groups 1 and 3 were given vehicle. During the course of the study, the growth of the subcutaneously implanted primary tumors was determined twice weekly by luminescence and caliper measurement. (B) Top: Box-and-whisker plots comparing different groups at day 14 and day 21. Significance was calculated using unpaired t test. Results are expressed as mean SEM; **P < 0.01. Bottom: Representative in vivo images of different groups, treated with vehicle or CAL-101. (C) Histological analysis of Ki-67 expression in representative tumor tissue sections of different groups. Original magnification, ×20. (D) Representative images of immunofluorescent staining of MDA-MB-231/MRC5 tumor cryosections for α-SMA and p-AKTSer473 after DMSO or CAL-101 treatment. Significance was calculated using unpaired t test. Results are expressed as mean ± SEM; *P < 0.05 vs. vehicle-treated tumors. Original magnification, ×40. (E) Representative images of immunofluorescence staining of tumor cryosections using TE-7 anti–human fibroblast antibody. Original magnification, ×20.