G1-4A, a Polysaccharide from Tinospora cordifolia Inhibits the Survival of Mycobacterium tuberculosis by Modulating Host Immune Responses in TLR4 Dependent Manner.

Rapid emergence of drug resistance in Mycobacterium tuberculosis (MTB) is a major health concern and demands the development of novel adjunct immunotherapeutic agents capable of modulating the host immune responses in order to control the pathogen. In the present study, we sought to investigate the immunomodulatory effects of G1-4A, a polysaccharide derived from the Indian medicinal plant Tinospora cordifolia, in in-vitro and aerosol mouse models of MTB infection. G1-4A treatment of MTB infected RAW264.7 macrophages significantly induced the surface expression of MHC-II and CD-86 molecules, secretion of proinflammatory cytokines (TNF-α, IL-ß, IL-6, IL-12, IFN-γ) and nitric oxide leading to reduced intracellular survival of both drug sensitive (H37Rv) as well as multi drug resistant strains (Beijing and LAM) of MTB, which was partially attributed to G1-4A induced NO production in TLR4-MyD88 dependent manner. Similarly, bacillary burden was significantly reduced in the lungs of MTB infected BALB/c mice treated with G1-4A, with simultaneous up-regulation of the expression of TNF-α, INF-γ and NOS2 in the mouse lung along with increased levels of Th1 cytokines like IFN-γ, IL-12 and decreased levels of Th2 cytokine like IL-4 in the serum. Furthermore, combination of G1-4A with Isoniazid (INH) exhibited better protection against MTB compared to that due to INH or G1-4A alone, suggesting its potential as adjunct therapy. Our results demonstrate that modulation of host immune responses by G1-4A might improve the therapeutic efficacy of existing anti-tubercular drugs and provide an attractive strategy for the development of alternative therapies to control tuberculosis.

Antifolate Activity of Plant Polyphenols against Mycobacterium tuberculosis.

With the view of exploring phytochemicals as Mycobacterium tuberculosis (Mtb) dihydrofolate reductase inhibitors, known plant polyphenols from various classes were subjected to detailed docking studies. From this in-silico screening, seven polyphenols were selected and tested against Mtb H37 Rv in whole cell assays. The phytochemicals exhibited potential activity ranging from 3 to 183 µm. These molecules were then tested against the pathogenic and human enzymes in a high-throughput microtitre assay. Epigallocatechin gallate showed the best activity and selectivity. The in-silico analysis was in agreement with the assay results. Of these 7 polyphenols, 5 exhibiting minimum inhibitory concentration values of ≤15 µm were tested for synergistic activity with first line drug Ethambutol and second line folate inhibitor para-amino salicylic acid. Epigallocatechin gallate, Magnolol and Bakuchiol exhibited moderate synergistic association by lowering the minimum inhibitory concentration of these drugs. These simple phytochemicals could hence be considered as leads for further studies, or for preparation of semi-synthetic derivatives to be used in combination therapy, for increased anti-tuberculosis activity after validation in-vivo.

Radioprotective effect of Ocimum sanctum and amifostine on the salivary gland of rats after therapeutic radioiodine exposure.

The current study investigated the radioprotective effect of Ocimum sanctum on the salivary gland of rats administered radioiodine ((131)I) and compared its efficacy with a known radioprotectant, amifostine. The experimental rats were divided in four groups and sacrificed in three different batches at 1, 3, and 6 months of time interval after 18.5 MBq/100g (i.p.) (131)I exposure. Six months duration batch received (131)I exposure twice with the gap of 3 months. Two groups of experimental rats were presupplemented with O. sanctum (40 mg/kg for 5 days, orally) and amifostine (200 mg/kg, s.c) before (131)I exposure separately. Increased Technetium-99m-pertechnetate ((99m)TcO(4)(-)) uptake at 30 minutes post injection in salivary glands of only (131)I exposed rats may imply delay in clearance at 6 months of exposure in comparison to their counterparts sacrificed at 1 month. Parotid gland histology showed atrophy with lipomatosis in only (131)I exposed rats at 3 and 6 months of duration. O. sanctum and amifostine presupplemented and subsequently exposed to (131)I rats at 3 and 6 months duration exhibited comparable histopathology with controls. Our study indicates possible radioprotective effect of O. sanctum and amifostine against high-dose (131)I exposure.

Synthesis and preliminary biological evaluation of novel N-(3-aryl-1,2,4-triazol-5-yl) cinnamamide derivatives as potential antimycobacterial agents: an operational Topliss Tree approach.

A series of novel N-(3-aryl-1,2,4-triazol-5-yl) cinnamamide derivatives were designed on basis of structural similarity to the known FAS II inhibitors. Topliss operational method was used to optimize the potency of molecules. The minimum inhibitory concentration (MIC) of all synthesized compounds was determined against Mycobacterium tuberculosis H(37)R(v) using resazurin microtitre assay (REMA) plate method. The synthesized compounds exhibit antimycobacterial activity in the range of 5-95µM with a good safety profile.

The possibility of a fully automated procedure for radiosynthesis of fluorine-18-labeled fluoromisonidazole using a simplified single, neutral alumina column purification procedure.

A novel fully automated radiosynthesis procedure for [(18)F]Fluoromisonidazole using a simple alumina cartridge-column for purification instead of conventionally used semi-preparative HPLC was developed. [(18)F]FMISO was prepared via a one-pot, two-step synthesis procedure using a modified nuclear interface synthesis module. Nucleophilic fluorination of the precursor molecule 1-(2'-nitro-1'-imidazolyl)-2-O-tetrahydropyranyl-3-O-toluenesulphonylpropanediol (NITTP) with no-carrier added [(18)F]fluoride followed by hydrolysis of the protecting group with 1M HCl. Purification was carried out using a single neutral alumina cartridge-column instead of semi-preparative HPLC. The maximum overall radiochemical yield obtained was 37.49+/-1.68% with 10mg NITTP (n=3, without any decay correction) and the total synthesis time was 40+/-1 min. The radiochemical purity was greater than 95% and the product was devoid of other chemical impurities including residual aluminum and acetonitrile. The biodistribution study in fibrosarcoma tumor model showed maximum uptake in tumor, 2h post injection. Finally, PET/CT imaging studies in normal healthy rabbit, showed clear uptake in the organs involved in the metabolic process of MISO. No bone uptake was observed excluding the presence of free [(18)F]fluoride. The reported method can be easily adapted in any commercial FDG synthesis module.

Difficulty in dislodging in vivo fixed radiostrontium.

Many trials based on the basic phenomena of isotopic dilution, adsorption, ion exchange, chelation, etc., have been attempted for the decorporation of radiostrontium, particularly Sr, after its entry in the in vivo system. We have recently demonstrated a non-isotopic carrier effect of some common calcium salts (calcium = 9 mg mL) to reduce the whole body retention of radiostrontium, if administered within 2 h after radiostrontium exposure and furthermore once daily, in rats, supplemented with calcium fortified diet. However, 25-30% of radiostrontium (compared to 50-60% in untreated animals) was still found to be retained in the animal even after 2 wk of treatment. Trial of some simple interventional measures, which would not adversely affect the animal metabolism, like pyrophosphate and magnesium sulfate, sodium citrate, chitin (a bio-absorbent), crown ether (a metal-chelator), and ammonium chloride, was therefore attempted to dislodge this remaining radiostrontium by switching over these animals to normal diet and subjecting them to different lines of treatment with these simple interventions through diet and drinking water separately for a further 4 wk. However, this remaining portion of radiostrontium is fixed in the bone and is difficult to dislodge.

An effective and better strategy for reducing body burden of radiostrontium.

In this study we have examined the effect of different calcium salts, Ca gluconate (CaG), Ca lactate (CaL), Ca carbonate (CaC) and Ca phosphate (CaP), on the clearance of radiostrontium (*Sr) administered either intraperitoneally (ip) (*Sr-ip group) or orally (*Sr-oral group) in rats. The influence of these Ca salts was examined in a group of animals administered *Sr ip, while the effect of three Ca salts (CaG, CaL and CaP) was studied in another group of rats given *Sr orally and compared with that of Ca alginate (CaA), normally advised for *Sr decorporation. Rats from both groups were subdivided into control and four experimental subgroups and were housed individually. The experimental subgroups were given the respective Ca salts (elemental Ca = 9 mg/rat/day) 2 h post 85Sr, and thereafter once daily. In the *Sr-ip group, CaG was administered ip while the other Ca salts were given orally. In the *Sr-oral group all Ca salts were administered orally. In addition, the diet of all the experimental subgroups was supplemented with the respective Ca salts to 2% elemental Ca. The whole-body retention (WBR) of *Sr in animals treated with Ca salts was found to be significantly reduced from 50-60% at 24 h to 20-30% at the end of 15 days compared with 70-80% at 24 h to 50-60% at the end of 15 days in the untreated control animals. The results strongly suggest that CaA could be replaced by any of the commonly used Ca salts for curtailing the WBR of *Sr. CaG which was administered ip, in the *Sr-ip group, was found to be more effective in reducing the WBR of *Sr.