Automated radiochemical synthesis of pharmaceutical grade [18 F]FLT using 3-N-Boc-5'-O-dimethoxytrityl-3'-O-nosyl-thymidine precursor and its Sep-Pak® purification employing selective elution from reversed phase.

Pharmaceutical grade 3'-deoxy-3'-[18 F]fluorothymidine [18 F]FLT was synthesized using 3-N-Boc-5'-O-dimethoxytrityl-3'-O-nosyl-thymidine (BOC-Nosyl) precursor, in the general purpose TRACERlab FX modules. Purification of [18 F]FLT, via solid phase extraction (SPE) after radiosynthesis, using a combination of different SPE cartridges, yielded satisfactory results, with radiochemical and chemical purity >99%. While the non-decay corrected radiochemical yield (RCY) with 20 mg (24 µmole) of BOC-Nosyl precursor was found to be 6.80 ± 0.16%, the decay corrected radiochemical yield (RCY) was 9.95 ± 0.24%. Residual acetone, acetonitrile, and ethanol levels were found to be 22.97 ± 0.76, 109.08 ± 0.93, and 7,666.45 ± 3.7 ppm, respectively. A simplified method for solid-phase purification of [18 F]FLT was developed, circumventing the need for HPLC purification. Biodistribution in C57BL/6 mice with B16F10 cell line-induced melanoma showed tumor to blood ratio of ~3.8 at 90 min. PET/CT imaging of normal rabbit injected with [18 F]FLT shows selective uptake in the bone marrow and small intestine. [18 F]FLT was found to be excreted through the kidneys and get collected in the urinary bladder, 120 min post injection. PET/CT imaging performed in rabbit model at 30, 60, 90, and 120 min post [18 F]FLT injections showed concordance with tissue distribution kinetics of mice tumor model.

Potential immunomodulatory effect of allelochemical juglone in mice vaccinated with BCG.

Juglone, a naphthoquinone is a known phytotoxic and cytotoxic allelochemical. Various anticancer studies suggest that pharmacological ROS insult and enzymes inhibited by juglone could be of therapeutic utility. In this study, we show that juglone induces immunomodulation in BCG-vaccinated mice. We have shown that juglone treatment leads to the generation of Th1 cytokines which develops an M1 phenotype in splenic macrophages. These findings were supported by a decrease in Treg population in BCG-vaccinated mice treated with juglone. Additionally, we observed juglone significantly increases CD8+ and T-helper memory population in BCG-vaccinated mice under immunomodulation. Further, BCG-challenge test showed that juglone enhances effector immune functions. Our finding was demonstrated by estimating ex-vivo Th1 and Th2 cytokines, flow cytometric analysis of Treg, T-helper memory, and CD8+ cells and determining serum IgG2a/IgG1 titer shift in BCG-vaccinated animals treated with juglone. Collectively, our findings provide evidence that juglone may act as immunomodulator when used at a low dose, metronomically.

Improved method for preparing Ni(II) complex of (S)-tyrosine Schiff base and its use in the automated synthesis of O-(2'-[18F]fluoroethyl)-l-tyrosine using solid-phase extraction purification.

O-(2'-[18F]fluoroethyl)-l-tyrosine is reportedly suitable for PET-imaging of brain tumours. We report here the synthesis of Ni(II)-complex of Schiff's base (S)-[N-2-(N'-benzylprolyl)amino]-benzophenone,((S)BPB) and alkylated l-tyrosine precursor, Ni(II)-(S)-BPB-l-Tyr-O-CH2-CH2-OTs by an improved method. A fully-automated radio-synthesis including non-HPLC purification was developed with a radio-chemical yield of 24.6 ± 2% in 70 ± 2min (n = 5). Radiochemical and enantiomeric purity was > 98% and 94% respectively. Bio-distribution and micro-PET studies in C57BL/6 mice bearing B16F10 melanoma showed tumor to brain ratio of 3.36 and 3.62 respectively at 60min post-injection.

A Novel Folate-Targeted Nanoliposomal System of Doxorubicin for Cancer Targeting.

Targeted drug delivery systems for cancer improves anti-tumor efficacy and reduces systemic toxicity by restricting availability of cytotoxic drugs within tumors. Targeting moieties, such as natural ligands (folic acid, transferrin, and biotin) which are overexpressed on tumors, have been used to enhance liposome-encapsulated drug accumulation within tumors and resulted in better control. In this report, we explored the scope of targeting ligand folic acid, which is incorporated in liposome systems using folic acid-modified cholesterol (CPF), enabled highly selective tumor-targeted delivery of liposome-encapsulated doxorubicin and resulted in increased cytotoxicity within tumors. Folate-tagged poloxamer-coated liposomes (FDL) were found to have significantly higher cellular uptake than conventional poloxamer-coated liposomes (DL), as confirmed by fluorometric analysis in B16F10 melanoma cells. Biodistribution study of the radiolabeled liposomal system indicated the significantly higher tumor uptake of FDL as compared to DL. Anti-tumor activity of FDL against murine B16F10 melanoma tumor-bearing mice revealed that FDL inhibited tumor growth more efficiently than the DL. Taken together, the results demonstrated the significant potential of the folate-conjugated nanoliposomal system for drug delivery to tumors.

Mono-guanidine heterolipid based SMEDDS: A promising tool for cytosolic delivery of antineoplastics.

In the present work, we designed and synthesized a novel mono-guanidine heterolipid (MGH) and confirmed its structure by NMR and ESI-MS. The MGH was used as cationic lipid in developing etoposide loaded cationic self-microemulsifying drug delivery system (ECS) intended to be delivered by intratumoral route. The ECS exhibited size <50 nm and zeta potential +32.6 mV on dilution with various isotonic vehicles with no phase separation or drug precipitation. The ECS could be easily sterilized by membrane filtration method and showed excellent stability for 6 months. The ECS demonstrated excellent in vitro antiproliferative activity against B16F10 cells which is attributed to its high transfection efficiency and capability to cause prolonged drug release in cytosolic space. In vivo antitumor activity of ECS was conducted in B16F10 induced melanoma tumor model. ECS at 12 mg/kg dose showed superior tumor suppression ability and exhibited 100% survival compared to other formulations. Mice treated with ECS by intratumoral route, showed neither systemic side effect nor any evidences of hepatotoxicity and nephrotoxicity. In contrast, etoposide administered by intravenous route showed remarkable systemic toxicity, hepatotoxicity and nephrotoxicity.

Anticancer activity of betulinic acid on MCF-7 tumors in nude mice.

Breast cancer is a major public health problem and the low effectiveness of conventional therapies to achieve long-term survival results in increased mortality associated with advanced breast cancers. Betulinic acid (BA) is a pentacyclic triterpene which can be isolated from number of plants grown in the tropics. It exhibits cytotoxic activity against variety of cancer cell lines. In the present study, the in vitro cytotoxic activity and in vivo antitumor activity of BA was evaluated in athymic nude mice bearing MCF-7 breast adenocarcinoma xenografts. In vitro cytotoxic activity of BA on MCF-7 cells was studied using the MTT assay and BA was cytotoxic towards MCF-7 cells with IC50 value of 13.5 microg/mL. The antitumor activity of BA was studied at concentrations of 50 and 100 mg/kg body weight in mice injected with MCF-7 cells. BA treatment delayed tumor formation and statistically significant reduction (P < 0.0001) of 52 and 77% in the tumor size at concentrations of 50 and 100 mg, respectively was observed. Histopathological analysis of tumors revealed decreased angiogenesis, proliferation and invasion in BA treated animals. This is one of the first studies demonstrating the in vivo antitumor activity of BA on MCF-7 breast cancer tumors in nude mice. The antitumor effect of BA can further be enhanced by use of combination therapy and novel drug delivery systems, thus making it a promising candidate for management of breast cancer patients.

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.