Towards the design of a bisphosphonate-based coordination complex nanocrystals formulation aimed to treat osteolytic metastases

The hydrothermal reaction between bioactive metals (Ca2+, Zn2+, and Mg2+) salts and clinically utilized bisphosphonate (BP), alendronate (ALEN), promotes the formation of bisphosphonate-based coordination complexes (BPCCs). The systematic exploration of the effect of three variables M2+/BP molar ratio, temperature, and pH on the reaction yielded materials of enough crystal quality for structural elucidation (ALEN-Ca forms I and II, ALEN-Mg, and ALEN-Zn forms I and II). Crystal structure was unveiled by single X-ray diffraction at 100 K and solid-state properties were revealed by thermogravimetry, vibrational spectroscopy, and elemental analysis. The dissolution of these BPCCs was tested and contrasted to that of the generic form of Fosamax®. Quantification of the ALEN content was performed by derivatization with Cu2+, which produced a soluble complex suitable for detection by UV-Vis spectroscopy. A phase inversion temperature (PIT) nano-emulsion method was applied to the synthesis of ALEN-Ca form II. Size distribution analysis demonstrated efficient particle size reduction of the BPCCs (from ~60 µm to ~438 d.nm). Aggregation tendency of the nano-Ca@ALEN was measured in two different dispersants (nanopure water and 10% FBS:PBS). After 48 h, the particle size distribution in 10% FBS:PBS remained constant and homogeneous. Binding assays for nano-Ca@ALEN to hydroxyapatite (HA) demonstrated 74% of nanocrystals bound in 12 days. The cytotoxicity of nano-Ca@ALEN against the MDA-MB-231 (metastatic breast cancer) and hFOB 1.19 (normal osteoblast-like cells) cell lines was investigated. Results demonstrated significant cell growth inhibition for nano-Ca@ALEN against the cancerous model, specifically at a concentration of 3.8 µM (% RCL = 44 ± 2 %). At this concentration, the nanocrystals did not present cytotoxicity against hFOB 1.19, suggesting the potential of this system to treat metastatic cancerous cells without affecting normal cell tissue at the bone microenvironment. These results provide evidence of the structure, stability, dissolution, aggregation, binding, and cytotoxicity properties of BPCCs, particularly nano-Ca@ALEN, and demonstrate its potential as a nano-formulation able to treat breast-cancer-induced osteolytic metastases (OM).