Combined Therapies for Neuroblastoma Based on the Activation of the Calcium- Sensing Receptor

Abstract

[eng] Neuroblastoma (NB) is the most common extracranial solid childhood tumour. Its clinical and histological manifestations range from benign tumours that spontaneously regress to highly aggressive metastatic tumours (Matthay et al -2016). In spite of the advances in treatment and the multidisciplinary approaches, almost half of high-risk NB patients do not survive (Whittle et al – 2017). The Calcium-sensing receptor (CaSR) is a G-protein coupled receptor (GPCR) that was found to be expressed in good prognosis NB tumour (de Torres et al – 2009).

Activation of this receptor using cinacalcet (CIN), a positive allosteric modulator of CaSR, in a xenograft NB animal model, reduced tumour growth. In addition, it was described that in NB cell lines with an overexpression of CaSR, CIN induced ER-stress mediated apoptosis (Rodríguez-Hernández et al – 2016). However, efficacy of CIN in the treatment of NB is limited by the low expression of CaSR in high-risk NB.

Furthermore, CIN acts mainly in the CaSR present in the parathyroid glands, inducing hypocalcemia.

This work addresses these two limitations of using CIN in the treatment of NB patients. We demonstrate that the active compound of vitamin D, 1,25-dihydroxyde vitamin D (1,25-D3) and the retinoids all-trans-retinoic acid (ATRA) and 13-cis-retinoic acid (13-cis-RA) increase the expression of CaSR in two NB cell lines.

Unfortunately, in vitro anti-tumorigenic capacities of CIN are not increased by its combination with 1,25-D3. Moreover, we show that the strong in vitro and in vivo anti-tumorigenic capacities of retinoids are unaltered by its combination with CIN.

On the other hand, we identify another positive allosteric modulator of CaSR, AC-265347, with NB anti-tumorigenic properties which does not induce hypocalcemia in mouse animal models. In these models each calcimimetic induce a differential protein and gene expression pattern, suggesting a different mechanism of action in the inhibition of tumour growth. Additionally, in vitro studies show that CIN and AC-265347 induce different stages of differentiation in NB cell lines.

Altogether, our data shows that the combination of CIN with different drugs that induce an increase in the expression levels of CaSR do not potentiate the anti-tumorigenic effect of CIN. More importantly, this work identifies a new calcimimetic that shows a potential neuroblastoma specific effect, AC-265347 inhibits NB tumour growth while maintaining plasma calcium levels. Our results strongly suggest that AC-265347 may be and effective therapeutic agent against NB, alone or in combination with other potentially synergistic treatments.