Cristian Vitale

Novel alkynylphosphonate analogue of calcitriol with potent antiproliferative effects in cancer cells and lack of calcemic activity

Here, we describe the design and synthesis of diethyl [(5Z,7E)-(1S,3R)-1,3-dihydroxy-9,10-secochola-5,7,10(19)-trien-23-in-24-yl] phosphonate (compound 10), which combines the low calcemic properties of phosphonates with the decreased metabolic inactivation due to the presence of a triple bond in C-24 and studied its in vitro effects on several cancer cell lines and its in vivo effects on blood calcium levels. We demonstrate that this compound is a potent antiproliferative vitamin D analogue, showing lack of calcemic effects in vivo.

Inhibition of p300 suppresses growth of breast cancer. Role of p300 subcellular localization

There is evidence that p300, a transcriptional co-factor and a lysine acetyl-transferase, could play a role both as an oncoprotein and as a tumor suppressor, although little is known regarding its role in breast cancer (BC). First we investigated the role p300 has on BC by performing pharmacological inhibition of p300 acetyl-transferase function and analyzing the effects on cell count, migration and invasion in LM3 murine breast cancer cell line and on tumor progression in a syngeneic murine model. We subsequently studied p300 protein expression in human BC biopsies and evaluated its correlation with clinical and histopathological parameters of the patients. We observed that inhibition of p300 induced apoptosis and reduced migration and invasion in cultured LM3 cells. Furthermore, a significant reduction in tumor burden, number of lung metastases and number of tumors invading the abdominal cavity was observed in a syngeneic tumor model of LM3 following treatment with the p300 inhibitor. This reduction in tumor burden was accompanied by a decrease in the mitotic index and Ki-67 levels and an increase in Bax expression. Moreover, the analysis of p300 expression in human BC samples showed that p300 immunoreactivity is significantly higher in the cancerous tissues than in the non-malignant mammary tissues and in the histologically normal adjacent tissues. Interestingly, p300 was observed in the cytoplasm, and the rate of cytoplasmic p300 was higher in BC than in non-tumor tissues. Importantly, we found that cytoplasmic localization of p300 is associated with a longer overall survival time of the patients. In conclusion, we demonstrated that inhibition of the acetylase function of p300 reduces both cell count and invasion in LM3 cells, and decreases tumor progression in the animal model. In addition, we show that the presence of p300 in the cytoplasm correlates with increased survival of patients suggesting that its nuclear localization is necessary for the pro-tumoral effects.

The alkynylphosphonate analogue of calcitriol EM1 has potent anti-metastatic effects in breast cancer

The active form of vitamin D3, calcitriol, plays a major role in maintaining calcium/phosphate homeostasis. In addition, it is a potent antiproliferative and prodifferentiating agent. However, when effective antitumor doses of calcitriol are employed, hypercalcemic effects are observed, thus precluding its therapeutic application. To overcome this problem, structural analogues have been designed with the aim at retaining or even increasing the antitumor effects while decreasing its calcemic activity. This report shows the biological evaluation of an alkynylphosphonate vitamin D less-calcemic analogue in a murine model of breast cancer. We demonstrate that this compound has potent anti-metastatic effects through its action over cellular migration and invasion likely mediated through the up-regulation of E-cadherin expression. Based on the current in vitro and in vivo results, EM1 is a promising candidate as a therapeutic agent in breast cancer.

Phosphonate Analogues of 1α, 25 Dihydroxyvitamin D 3 are Promising Candidates for Antitumoural Therapies

The active metabolite of vitamin D, 1α, 25 dihydroxyvitamin D3 (calcitriol) is classically known to regulate calcium and phosphate homeostasis and bone mineralization. In addition, calcitriol has also been documented to act as a potent anticancer agent in multiple cell culture and animal models of cancer. However, major side effects, such as hypercalcemia, hinder broad-spectrum therapeutic uses of calcitriol in cancer chemotherapy. Synthesis of calcitriol analogues with the same or increased antiproliferative and pro-differentiating activities, and with reduced undesired effects on calcium and bone metabolism, is getting significant attention towards rational therapeutics to treat cancer. In this regard, phosphonate analogues have been shown to display a certain degree of dissociation between the vitamin D activity in vitro and undesired hypercalcemia in vivo. However, few phosphonates have been described in the literature and fewer of them tested for antitumoral effects. Our group has synthesized a novel vitamin D analogue (EM1) bearing an alkynylphosphonate moiety that combines the low calcemic properties of phosphonates with the decreased metabolic inactivation due to the presence of a triple bond between C-23 and C-24. Biological assays demonstrated that this analogue has potent antiproliferative effects in a wide panel of tumour cell lines, even in those resistant to calcitriol treatment. Importantly, EM1 does not show toxic effects in animals, even administered at high doses and for extended periods of time. In the current review we discuss the effects and the potential application in cancer of vitamin D and its derivatives, with an emphasis on phosphonate analogues.

Antitumoral effects of the alkynylphosphonate analogue of calcitriol EM1 on glioblastoma multiforme cells

Glioblastoma multiforme (GBM) is the worst and most common brain tumor, characterized by high proliferation and invasion rates. The current standard treatment is mainly based on chemoradiotherapy and this approach has slightly improved patient survival. Thus, novel strategies aimed at prolonging the survival and ensuring a better quality of life are necessary. In the present work, we investigated the antitumoral effect of the novel analogue of calcitriol EM1 on GBM cells employing in vitro, in silico, and in vivo assays. In vitro, we demonstrated that EM1 treatment selectively decreases the viability of murine and human tumor cells without affecting that of normal human astrocytes. The analysis of the mechanisms showed that EM1 produces cell cycle arrest in the T98G cell line, which is accompanied by an increase in p21, p27, p57 protein levels and a decrease in cyclin D1, p-Akt-S473, p-ERK1/2 and c-Jun expression. Moreover, EM1 treatment also exerts in GBM cells anti-migratory effects and decreases their invasive capacity by a reduction in MMP-9 proteolytic activity. In silico, we demonstrated that EM1 is able to bind to the vitamin D receptor with greater affinity than calcitriol. Finally, we showed that EM1 treatment of nude mice administered at 50ug/kg body weight during 21days neither induces hypercalcemia nor toxicity effects. In conclusion, all the results indicate the potential of EM1 analogue as a promising therapeutic alternative for GBM treatment.

A Straightforward Methodology for the Synthesis of α,ω-Telechelic Poly(dimethylsiloxane)s

In this work we report the synthesis of α,ω-telechelic poly(dimethylsiloxane)s (α,ω-PDMS) by employing a novel bifunctional initiator obtained from a commercially available siloxane precursor, diglycidylether-terminated poly(dimethylsiloxane) (PDMS-DGE). The synthetic strategy was easily followed by different colour changes, and involved the high-vacuum reaction of sec-Bu−Li+ with 1,1′-diphenylethylene (DPE) to afford the addition adduct (bright red) that was subsequently reacted with PDMS-DGE, promoting the nuclephilic ring-opening from epoxide-end chains. The resulting bifunctional initiator (light green) was then employed to polymerize hexamethyl(cyclotrisiloxane) monomer (D3) by using conventional anionic polymerization (from light green to pale yellow). From suitable terminating agents, silane (–SiH), vinyl (–CH=CH2), hydroxy (–OH), and even methacryloyl α,ω-PDMS were obtained. 1H NMR and FT-IR analyses confirmed the presence of the targeted functional groups in the resulting polymers. However, a careful siliconization procedure should be performed over glass surfaces during the fractionation of chlorosilane ampoules in order to avoid the presence of silanol moieties that decrease end-capping efficiency. This fact was observed not only from NMR but also from size exclusion chromatography (SEC) analyses, since narrow molar masses distributions (1.15 ≤ Mw/Mn ≤ 1.3) and a good control over the resulting molar masses were observed.