Molecular bowls for inclusion complexation of toxic anticancer drug methotrexate.

We describe the preparation and study of novel cavitands, molecular bowls 16+ and 26+, as good binders of the anticancer drug methotrexate (MTX). Molecular bowls are comprised of a curved tribenzotriquinacene (TBTQ) core conjugated to three macrocyclic pyridinium units at the top. The cavitands are easily accessible via two synthetic steps from hexabromo-tribenzotriquinacene in 25% yield. As amphiphilic molecules, bowls 16+ and 26+ self-associate in water by the nucleation-to-aggregation pathway (NMR). The bowls are preorganized, having a semi-rigid framework comprising a fixed bottom with a wobbling pyridinium rim (VT NMR and MD). Further studies, both experimental (NMR) and computational (DFT and MCMM), suggested that a folded MTX occupies the cavity of bowls wherein it forms π-π, C-H-π, and ion pairing intermolecular contacts but also undergoes desolvation to give stable binary complexes (µM) in water. Moreover, a computational protocol is introduced to identify docking pose(s) of MTX inside molecular bowls from NMR shielding data. Both molecular bowls have shown in vitro biocompatibility with liver and kidney cell lines (MTS assay). As bowl 26+ is the strongest binder of MTX reported to date, we envision it as an excellent candidate for further studies on the way toward developing an antidote capable of removing MTX from overdosed cancer patients.

On a stake-out: Mycobacterial small RNA identification and regulation.

Persistence of mycobacteria in the hostile environment of human macrophage is pivotal for its successful pathogenesis. Rapid adaptation to diverse stresses is the key aspect for their survival in the host cells. A range of heterogeneous mechanisms operate in bacteria to retaliate stress conditions. Small RNAs (sRNA) have been implicated in many of those mechanisms in either a single or multiple regulatory networks to post-transcriptionally modulate bacterial gene expression. Although small RNA profiling in mycobacteria by advanced technologies like deep sequencing, tilling microarray etc. have identified hundreds of sRNA, however, a handful of those small RNAs have been unearthed with precise regulatory mechanism. Extensive investigations on sRNA-mediated gene regulations in eubacteria like Escherichia coli revealed the existence of a plethora of distinctive sRNA mechanisms e.g. base pairing, protein sequestration, RNA decoy etc. Increasing studies on mycobacterial sRNA also discovered several eccentric mechanisms where sRNAs act at the posttranscriptional stage to either activate or repress target gene expression that lead to promote mycobacterial survival in stresses. Several intrinsic features like high GC content, absence of any homologue of abundant RNA chaperones, Hfq and ProQ, isolate sRNA mechanisms of mycobacteria from that of other bacteria. An insightful approach has been taken in this review to describe sRNA identification and its regulations in mycobacterial species especially in Mycobacterium tuberculosis.

Hydrazone-Linkage-Based Self-Healing and Injectable Xanthan-Poly(ethylene glycol) Hydrogels for Controlled Drug Release and 3D Cell Culture.

Polymeric hydrogels have been extensively explored for controlled drug-delivery applications, but there is an increasing demand for smart drug delivery combined with tunable physicochemical attributes and tissue engineering potential. In this work, novel xanthan-poly(ethylene glycol) (PEG) hydrogels were developed by cross-linking polysaccharide, oxidized xanthan, and 8-arm PEG hydrazine through dynamic, pH-responsive, and biodegradable hydrazone linkages. Aqueous solutions (pH 6.5) of oxidized xanthan and PEG hydrazine were mixed together at 37 °C to obtain hydrogels within minutes, and the formation of hydrazone linkages was ascertained using Fourier transform infrared spectroscopy. Fabrication of xanthan-PEG hydrogels using hydrazone linkages has not been reported previously. The 3% hydrogels exhibited the storage modulus of 194 Pa, which increased to 770 Pa for 5% hydrogels. When subjected to alternating cycles of varying strains of 1 and 800% (5 cycles), hydrogels demonstrated instant recovery each time the extreme strain was relieved, thus suggesting excellent self-healing capabilities. Doxorubicin (DOX), chemotherapeutic agent, was loaded onto hydrogels, and release studies were carried out at pH 5.5 (tumoral) and 7.4 (physiological). The cumulative release from 3, 4, and 5% hydrogels at pH 5.5 was 81.06, 61.98, and 41.67%, whereas the release at pH 7.4 was 47.43, 37.01, and 35.34% at 30 days. MTT assay showed that oxidized xanthan and PEG hydrazine are not toxic to mammalian cells (NIH-3T3), as the cell viabilities were found to be 84.66 and 102% for concentrations up to 1 mg/mL. The live/dead assay with encapsulated NIH-3T3 cells showed no significant dead cell population, suggesting excellent compatibility of hydrogels in 2D and 3D culture. DOX-loaded hydrogels exhibited cytotoxicity against A549 cells when exposed to media released from hydrogels. Overall, hydrogels developed in this work may have potential applications in drug delivery and 3D cell culture for cell delivery.