Direct interaction of TrkA/CD44v3 is essential for NGF-promoted aggressiveness of breast cancer cells

Abstract

Background CD44 is a multifunctional membrane glycoprotein. Through its heparan sulfate chain, CD44 presents growth factors to their receptors. We have shown that CD44 and Tropomyosin kinase A (TrkA) form a complex following nerve growth factor (NGF) induction. Our study aimed to understand how CD44 and TrkA interact and the consequences of inhibiting this interaction regarding the pro-tumoral effect of NGF in breast cancer. Methods After determining which CD44 isoforms (variants) are involved in forming the TrkA/CD44 complex using proximity ligation assays, we investigated the molecular determinants of this interaction. By molecular modeling, we isolated the amino acids involved and confirmed their involvement using mutations. A CD44v3 mimetic peptide was then synthesized to block the TrkA/CD44v3 interaction. The effects of this peptide on the growth, migration and invasion of xenografted triple-negative breast cancer cells were assessed. Finally, we investigated the correlations between the expression of the TrkA/CD44v3 complex in tumors and histo-pronostic parameters. Results We demonstrated that isoform v3 (CD44v3), but not v6, binds to TrkA in response to NGF stimulation. The final 10 amino acids of exon v3 and the TrkA H112 residue are necessary for the association of CD44v3 with TrkA. Functionally, the CD44v3 mimetic peptide impairs not only NGF-induced RhoA activation, clonogenicity, and migration/invasion of breast cancer cells in vitro but also tumor growth and metastasis in a xenograft mouse model. We also detected TrkA/CD44v3 only in cancerous cells, not in normal adjacent tissues. Conclusion Collectively, our results suggest that blocking the CD44v3/TrkA interaction can be a new therapeutic option for triple-negative breast cancers.

Expression and Signaling Pathways of Nerve Growth Factor (NGF) And Pro-NGF in Breast Cancer: A Systematic Review

“…About NGF receptors, Descamps et al [94] reported that the overall TrkA mRNA expression predicts a more favorable prognosis in a highly variable cohort of 363 primary breast carcinoma. However, as mentioned above, Davidson et al [114] showed that the levels of p-TrkA were associated with tumor progression to effusion in metastatic breast carcinoma, thus correlating the dysregulation of p-TrkA with poor prognostic outcome in a more uniform cohort of 39 patients. Furthermore, the immunohistochemical expression of NGFR/p75NTR receptor was reported as a positive and negative prognostic factor for non-TNBC and TNBC, respectively.…”

“…In addition, the authors reported a downregulation and an upregulation of NGFR/p75NTR and TrkA, respectively, compared to primary breast tumors. The levels of TrkA were also upregulated in locoregional recurrences compared to early lymph node metastases [114]. NGF strongly increased invasion, cord formation and the monolayer permeability of endothelial cells [111] and metastasis of xenografted breast cancer cells in immunodeficient mice [95].…”

“…In 2001, Aragona et al [93] observed a rapid metastatic spreading in NGFR/p75NTR-negative breast cancer patients suggesting, for the first time, the involvement of this NGF receptor in breast cancer natural history. In 2004, Davidson et al [114] characterized the expression of NGF, NGFR/p75NTR and phospho-TrkA (p-TrkA activated receptor) during the progression of breast carcinoma from primary tumor to pleural effusion in sections from 42 malignant pleural effusions from breast cancer patients and 65 corresponding solid tumors (34 primary, 31 metastatic). From the results of this study emerged that NGF expression in effusions significantly predicted a shorter time to progression (TTP).…”

“…In particular, it has been reported a transcript, protein and immunological expression of NGF in the majority of human breast tumors [91], making it a broader diagnostic potential than ER or HER-2 [17]. Noteworthy, several studies have also shown the overexpression of TrkA and NGFR/p75NTR receptors in most of the human breast cancers as compared to the expression of these receptors in normal cells [94,114]. Moreover, Islam et al [14] demonstrated that Russell's Viper Venom (RVV)-NGFa (an NGF isoform), labelled with Fluorescein Isothiocyanate (FITC), establishes strong binding to TrkA and NGFR/p75NTR receptors in breast cancer cells but not in non-cancerous cells.…”

“…Davidson et al [114] posed NGF as first molecular marker able to predict the time interval to progression. Namely, the authors found a mean of time to progression of 6.3 and of 4 years for NGF-negative and -positive effusions, respectively.…”