RESUMEN
A green, fast and selective approach for the synthesis of mono-substituted closo-decaborate derivatives [2-B10H9COR]2- has been established via a nucleophilic addition reaction between the carbonyl derivative of closo-decaborate [2-B10H9CO]- and the corresponding Grignard reagent RMgX, where R is the ethyl, iso-propyl, pentyl, allyl, vinyl and propynyl groups. This approach is accomplished under mild conditions with 70-80% yields. The significance of these derivative is their ability to constitute building blocks for polymeric integration via the allyl, vinyl and propynyl substituents. All products were characterized by 11B, 1H and 13C NMR, elemental analysis and mass spectrometry.
RESUMEN
Glycosaminoglycans (GAG) are key elements involved in various physiological and pathological processes including cancer. Several GAG-based drugs have been developed showing significant results and potential use as cancer therapeutics. We previously reported that alginate sulfate (AlgSulf), a GAG-mimetic, reduces the proliferation of lung adenocarcinoma cells. In this study, we evaluated the preferential effect of AlgSulf on tumorigenic and nontumorigenic mammary epithelial cells in 2D, 3D, and coculture conditions. AlgSulf were synthesized with different degrees of sulfation (DSs) varying from 0 to 2.7 and used at 100 µg/mL on HMT-3522 S1 (S1) nontumorigenic mammary epithelial cells and their tumorigenic counterparts HMT-3522 T4-2 (T4-2) cells. The anti-tumor properties of AlgSulf were assessed using trypan blue and bromodeoxyuridine proliferation (BrdU) assays, immunofluorescence staining and transwell invasion assay. Binding of insulin and epidermal growth factor (EGF) to sulfated substrates was measured using QCM-D and ELISA. In 2D, the cell growth rate of cells treated with AlgSulf was consistently lower compared to untreated controls (p<0.001) and surpassed the effect of the native GAG heparin (positive control). In 3D, AlgSulf preferentially hindered the growth rate and the invasion potential of tumorigenic T4-2 nodules while maintaining the formation of differentiated polarized nontumorigenic S1 acini. The preferential growth inhibition of tumorigenic cells by AlgSulf was confirmed in a coculture system (p<0.001). In the ELISA assay, a trend of EGF binding was detected for sulfated polysaccharides while QCM-D analysis showed negligible binding of insulin and EGF to sulfated substrates. The preferential effect mediated by the mimetic sulfated GAGs on cancer cells may in part be growth factor dependent. Our findings suggest a potential anticancer therapeutic role of AlgSulf for the development of anticancer drugs.
Asunto(s)
Biomimética , Neoplasias Pulmonares , Antígenos de Diferenciación , Proliferación Celular , Células Epiteliales , Glicosaminoglicanos , HumanosRESUMEN
BACKGROUND: Targeting angiogenesis has been considered a promising treatment of choice for a large number of malignancies, including gastrointestinal cancers. Bevacizumab is an anti-vascular endothelial growth factor (anti-VEGF) being used for this purpose. However, treatment efficacy is largely questioned. Telomerase activity, responsible for cancer cell immortality, is detected in 85-95% of human cancers and is considered a potential regulator of VEGF. The aim of our study was to investigate the interrelationship between VEGF and hTERT in gastrointestinal cancers and to explore cell response to a combined inhibition of telomerase and VEGF. METHODS: AGS (gastric cancer), Caco-2 (colorectal cancer) and HepG2/C3A (hepatocellular carcinoma), were treated with telomerase inhibitors BIBR-1232 (10µM) and costunolide (10µM), with bevacizumab (Avastin® at 5 ng/ml or 100µg/ml) or with a combination of both types of inhibitors. VEGF and hTERT mRNA levels, and telomerase activity were detected by RT-PCR. VEGF levels were quantified by ELISA. Telomerase was knocked down using hTERT siRNA and hTERT was overexpressed in the telomerase negative cell line, Saos-2 (osteosarcoma), using constructs expressing either wild type hTERT (hTERT-WT) or dominant negative hTERT (hTERT-DN). Tube formation by HUVECs was assessed using ECMatrix™ (EMD Millipore). RESULTS: Our results showed that telomerase regulates VEGF expression and secretion through its catalytic subunit hTERT in AGS, Caco2, and HepG2/C3A, independent of its catalytic activity. Interestingly, VEGF inhibition with bevacizumab (100µg/ml) increased hTERT expression 42.3% in AGS, 94.1% in Caco2, and 52.5% in HepG2/C3A, and increased telomerase activity 30-fold in AGS, 10.3-fold in Caco2 and 8-fold in HepG2/C3A. A further investigation showed that VEGF upregulates hTERT expression in a mechanism that implicates the PI3K/AKT/mTOR pathway and HIF-1α. Moreover, bevacizumab treatment increased VEGFR1 and VEGFR2 expression in cancer cells and human umbilical vein endothelial cells (HUVECs) through hTERT. Thus, the combination of bevacizumab with telomerase inhibitors decreased VEGF expression and secretion by cancer cells, inhibited VEGFR1 and VEGFR2 upregulation, and reduced tube formation by HUVECs. CONCLUSIONS: Taken together, our results suggest that bevacizumab treatment activates a VEGF autoregulatory mechanism involving hTERT and VEGF receptors and that an inhibition of this pathway could improve tumor cell response to anti-VEGF treatment.