Lipid-Based Nanocarrier by Targeting with LHRH Peptide: A Promising Approach for Prostate Cancer Radio-Imaging and Therapy.

Prostate cancer is a prevalently detected malignancy with a dismal prognosis. Luteinizing-hormone-releasing-hormone (LHRH) receptors are overexpressed in such cancer cells, to which the LHRH-decapeptide can specifically bind. A lipid-polyethylene glycol-conjugated new LHRH-decapeptide analogue (D-P-HLH) was synthesized and characterized. D-P-HLH-coated and anticancer drug doxorubicin (DX)-loaded solid lipid nanoparticles (F-DX-SLN) were formulated by the cold homogenization technique and characterized by Fourier transform infrared spectroscopy, X-ray diffraction, X-ray photoelectron spectroscopy, differential scanning calorimetry, dynamic light scattering, electron microscopy, entrapment efficiency, and drug-release profile studies. F-DX-SLN allows site-specific DX delivery by reducing the side effects of chemotherapy. Cancer cells could precisely take up F-DX-SLN by targeting specific receptors, boosting the cytotoxicity at the tumor site. The efficacy of F-DX-SLN on PC3/SKBR3 cells by the MTT assay revealed that F-DX-SLN was more cytotoxic than DX and/or DX-SLN. Flow cytometry and confocal microscopic studies further support F-DX-SLNs' increased intracellular absorption capability in targeting LHRH overexpressed cancer cells. F-DX-SLN ensured high apoptotic potential, noticeably larger mitochondrial transmembrane depolarization action, as well as the activation of caspases, a longer half-life, and greater plasma concentration. F-DX-SLN/DX-SLN was radiolabeled with technetium-99m; scintigraphic imaging studies established its tumor selectivity in PC3 tumor-bearing nude mice. The efficacy of the formulations in cancer treatment, in vivo therapeutic efficacy tests, and histopathological studies were also conducted. Results clearly indicate that F-DX-SLN exhibits sustained and superior targeted administration of anticancer drugs, thus opening up the possibility of a drug delivery system with precise control and targeting effects. F-DX-SLN could also provide a nanotheranostic approach with improved efficacy for prostate cancer therapy.

Synthesis, Characterization, and Biological Evaluation of Radiolabeled Glutamine Conjugated Polymeric Nanoparticles: A Simple Approach for Tumor Imaging.

Application of nanoradiopharmaceuticals for molecular imaging has gained worldwide importance for their multifaceted potentials focusing on providing a safe and cost-effective approach. Biodistribution studies on such species are capable of bringing nanomedicine to patients. Current therapeutically available labeling strategies suffer from different limitations, including off-target cytotoxicity and radiolabel release over time. Poly(lactic-co-glycolic acid)(PLGA) nanoparticles are biodegradable carriers for a variety of contrast agents that can be employed in medicine with high loading capacity for multimodal imaging agents. Here, glutamine-conjugated PLGA polymers were used to construct polymeric nanoparticles (G-PNP) similar to unconjugated PLGA nanoparticles (PNP)s formulated for ex vivo cell labeling and in vivo tumor scintigraphy studies. G-PNP/PNP, characterized by Fourier-transform infrared, atomic-force-microscopy, particle-size, and zeta-potential studies, were biocompatible as evaluated by MTT assay. G-PNPs were radiolabeled with 99mtechnetium (99mTc) by borohydrite reduction. G-PNPs demonstrated higher cellular uptake than PNPs, with no major cytotoxicity. Radiochemical purity indicated that 99mTc labeled G-PNP (99mTc-G-PNP) can form a stable complex with substantial stability in serum with respect to time. Imaging studies showed that 99mTc-G-PNP significantly accumulated at the C6 glioma cell induced tumor-site in rats. Thus, 99mTc-G-PNP demonstrated favorable characteristics and imaging potential which may make it a promising tumor imaging nanoprobe as a nanoradiopharmaceutical.

Preparation of [68Ga]Ga-Chloride from 68Zn solid target for the synthesis of pharmaceutical grade [68Ga]Ga-PSMA-11 and [68Ga]Ga-DOTA-TATE.

68Ga is produced from enriched zinc-68 target electrodeposited on copper base material which was irradiated with 15 MeV proton energy in 30 MeV cyclotron. A modified semi-automated separation and purification module was used to obtain pharmaceutical grade [68Ga]GaCl3 in 35 ± 5 min. The quality of [68Ga]GaCl3 produced was in accordance with Pharmeuropa 30.4. The [68Ga]GaCl3 was utilized for the formulation of multiple doses of [68Ga]Ga-PSMA-11 and [68Ga]Ga-DOTATATE. The quality of [68Ga]Ga-PSMA-11 and [68Ga]Ga-DOTATATE were also in accordance with Pharmacopeia.

HYNIC and DOMA conjugated radiolabeled bombesin analogs as receptor-targeted probes for scintigraphic detection of breast tumor.

BACKGROUND: Among the many peptide receptor systems, gastrin-releasing-peptide (GRP) receptors, the mammalian equivalent of bombesin (BN) receptors, are potential targets for diagnosis and therapy of breast tumors due to their overexpression in various frequently occurring human cancers. The aim of this study was to synthesize and comparative evaluation of 99mTc-labeled new BN peptide analogs. Four new BN analogs, HYNIC-Asp[PheNle]BN(7-14)NH2, BN1; HYNIC-Pro-Asp[TyrMet]BN(7-14)NH2, BN2; HYNIC-Asp-Asn[Lys-CHAla-Nle]BN(7-14)NH2, BN3; and DOMA-GABA[Pro-Tyr-Nle]BN(7-14)NH2, BN4 were synthesized and biologically evaluated for targeted imaging of GRP receptor-positive breast-tumors. METHODS: Solid-phase synthesis using Fmoc-chemistry was adopted for the synthesis of peptides. BN1-BN4 analogs were better over the standard Gln-Trp-Ala-Val-Gly-His-Leu-Met-NH2 (BNS). Lipophilicity, serum stability, internalization, and binding affinity studies were carried out using 99mTc-labeled analogs. Biodistribution and imaging analyses were performed on MDA-MB-231 cell-induced tumor-bearing mice. BN-analogs induced angiogenesis; tumor formation and GRP-receptor-expression were confirmed by histology and immunohistochemistry analyses of tumor sections and important tissue sections. RESULTS: All the analogs displayed ≥ 97% purity after the HPLC purification. BN-peptide-conjugates exhibited high serum stability and significant binding affinity to GRP-positive tumor; rapid internalization/externalization in/from MDA-MB-231 cells were noticed for the BN analogs. BN4 and BN3 exhibited higher binding affinity, stability than BN1 and BN2. Highly specific in vivo uptakes to the tumor were clearly visualized by scintigraphy; rapid excretion from non-target tissues via kidneys suggests a higher tumor-to-background ratio. BN4, among all the analogs, stimulates the expression of angiogenic markers to a maximum. CONCLUSION: Considering its most improved pharmacological characteristics, BN4 is thus considered as most promising probes for early non-invasive diagnosis of GRP receptor-positive breast tumors.

Garcinol loaded vitamin E TPGS emulsified PLGA nanoparticles: preparation, physicochemical characterization, in vitro and in vivo studies.

Garcinol (GAR) is a naturally occurring polyisoprenylated phenolic compound. It has been recently investigated for its biological activities such as antioxidant, anti-inflammatory, anti ulcer, and antiproliferative effect on a wide range of human cancer cell lines. Though the outcomes are very promising, its extreme insolubility in water remains the main obstacle for its clinical application. Herein we report the formulation of GAR entrapped PLGA nanoparticles by nanoprecipitation method using vitamin E TPGS as an emulsifier. The nanoparticles were characterized for size, surface morphology, surface charge, encapsulation efficiency and in vitro drug release kinetics. The MTT assay depicted a high amount of cytotoxicity of GAR-NPs in B16F10, HepG2 and KB cells. A considerable amount of cell apoptosis was observed in B16f10 and KB cell lines. In vivo cellular uptake of fluorescent NPs on B16F10 cells was also investigated. Finally the GAR loaded NPs were radiolabeled with technetium-99m with >95% labeling efficiency and administered to B16F10 melanoma tumor bearing mice to investigate the in vivo deposition at the tumor site by biodistribution and scintigraphic imaging study. In vitro cellular uptake studies and biological evaluation confirm the efficacy of the formulation for cancer treatment.

Synthesis and exploration of novel radiolabeled bombesin peptides for targeting receptor positive tumor.

Increasing evidence of peptide receptor overexpression in various cancer cells, warrant the development of receptor specific radiolabeled peptides for molecular imaging and therapy in nuclear medicine. Gastrin-releasing-peptide (GRP) receptor, are overexpressed in a variety of human cancer cells. The present study report the synthesis and biological evaluation of new bombesin (BBN) analogs, HYNIC-Asp-[Phe13]BBN(7-13)-NH-CH2-CH2-CH3:BA1, HYNIC-Pro-[Tyr13Met14]BBN(7-14)NH2:BA2 as prospective tumor imaging agent with compare to BBN(7-14)NH2:BS as standard. The pharmacophores were radiolabeled in high yields with 99mTc, characterized for their stability in serum and saline, cysteine/histidine and were found to be substantially stable. Internalization/externalization and receptor binding studies were assessed using MDA-MB-231 cells and showed high receptor binding-affinity and favourable internalization. Fluorescence studies revealed that BA1 changed the morphology of the cells and could localize in the nucleus more effectively than BA2/BS. Cell-viability studies displayed substantial antagonistic and nuclear-internalization effect of BA1. BA1 also exhibited antiproliferative effect on MDA-MB-231 cell by inducing apoptosis. In vivo behaviour of the radiopeptides was evaluated in GRP receptor positive tumor bearing mice. The 99mTc-BA1/99mTc-BA2 demonstrated rapid blood/urinary clearance through the renal pathway and comparatively more significant tumor uptake image and favourable tumor-to-non-target ratios provided by 99mTc-BA1. The specificity of the in vivo uptake was confirmed by co-injection with BS. Moreover, 99mTc-BA1 provided a much clearer tumor image in scintigraphic studies than others. Thus the combination of favourable in vitro and in vivo properties renders BA1 as more potential antagonist bombesin-peptide for targeting GRP-receptor positive tumor. These properties are encouraging to carry out further experiments for non-invasive receptor targeting potential diagnostinc and therapeutic agent for tumors.

Ursolic Acid Loaded PLGA Nanoparticles: in vitro and in vivo Evaluation to Explore Tumor Targeting Ability on B16F10 Melanoma Cell Lines.

PURPOSE: Ursolic acid (UA), a pentacyclic triterpenoid extracted from plants, shows promising inhibitory effect in different tumor bearing cell lines. In the present study we fabricated UA loaded PLGA nanoparticles (UA-NPs) as the drug carrier and thoroughly evaluated in vitro and in vivo the differential tumor targeting effects of UA and UA-NPs in B16F10 melanoma cells. METHODS: Ursolic acid loaded PLGA nanoparticles were prepared by emulsion solvent evaporation technique and evaluated for particle size, polydispersity, zeta potential and drug release potency. MTT assay as well as flow cytometric and confocal microscopic analyses were done in B16F10 mouse melanoma cell lines. Formulations were labeled with technetium-99m to evaluate the biodistribution and perform scintigraphic imaging studies following intravenous administration in tumor bearing mice model. RESULTS: Single emulsification technique produced smooth spherical nanoparticles of small size with relatively narrow size distribution (154 ± 4.56 nm). On B16F10 cell line, the formulation showed higher cytotoxicity compared to the free drug due to increased in vitro cellular uptake. The formulation was successfully radiolabeled and remained substantially (>90%) stable when incubated (37°C, 6 h) separately in normal saline or freshly collected rat serum or histidine solution. The radiolabeled UA-NPs exhibited slower blood clearance and comparatively high uptake in tumor region as evidenced by biodistribution and scintigraphic studies. CONCLUSIONS: The in vitro and in vivo studies have proved the tumor targeting potential of UA-NPs in B16F10 melanoma cell lines.

Exploring the Potential of (99m)Tc(CO)3-Labeled Triazolyl Peptides for Tumor Diagnosis.

In recent years the authors have reported on (99m)Tc(CO)3-labeled peptides that serve as carriers for biomolecules or radiopharmaceuticals to the tumors. In continuation of that work they report the synthesis of a pentapeptide (Met-Phe-Phe-Gly-His; pep-1), a hexapeptide (Met-Phe-Phe-Asp-Gly-His; pep-2), and a tetrapeptide (Asp-Gly-Arg-His; pep-3) and the attachment of 3-amino-1,2,4-triazole to the ß carboxylic function of the aspartic acid unit of pep-2 and pep-3. The pharmacophores were radiolabeled in high yields with [(99m)Tc(CO)3(H2O)3](+) metal aqua ion, characterized for their stability in serum and saline, as well as in His solution, and found to be substantially stable. B16F10 cell line binding studies showed favorable uptake and internalization. In vivo behavior of the radiolabeled triazolyl peptides was assessed in mice bearing induced tumor. The (99m)Tc(CO)3-triazolyl pep-3 demonstrated rapid urinary clearance and comparatively better tumor uptake. Imaging studies showed visualization of the tumor using (99m)Tc(CO)3-triazolyl pep-3, but due to high abdominal background, low delineation occurred. Based on the results further experiments will be carried out for targeting tumor with triazolyl peptides.

Polymers derived from Xanthomonas campesteris and Cyamopsis tetragonolobus used as retardant materials for the formulation of sustained release floating matrix tablet of atenolol.

The objective of the present study was to develop, optimize, in vitro, and in vivo evaluation of floating matrix tablet of atenolol using polymer blend derived from Xanthomonas campesteris and Cyamopsis tetragonolobus that are characterized by release requirements of sustained-release product and to improve the oral bioavailability of the drug. A 3(2) full factorial design was employed to optimize the tablets, where content of polymer blend (X1) and ratio of xanthan gum-to-guar gum (X2) were considered as independent variables. The effects of independent variables on dependent variables, i.e. floating time, diffusion exponent, and time to release 50% of atenolol were evaluated. The in vivo pharmacokinetic parameters of the optimized formulation were compared with the marketed sustained release formulation of atenolol (Aten(®)). The optimized formulation containing 20% (w/w) of polymer blend and 50:50 ratio of xanthan gum-to-guar gum was able to float more than 12h and showed the desired sustained drug release from the tablets. In vivo retention studies in rabbit stomach showed the gastric residence of tablet up to 6h. The in vivo study of optimized tablets illustrated significant improvement in the oral bioavailability of atenolol in rabbits. It can be concluded that floating matrix tablet of atenolol prepared by using xanthan gum and guar gum has potential for sustained release of the drug as well as improved oral bioavailability through enhanced gastric residence time of formulation in stomach.

Preparation and evaluation of 99mTc-cefuroxime, a potential infection specific imaging agent: a reliable thin layer chromatographic system to delineate impurities from the 99mTc-antibiotic.

Technetium-99m labelled cefuroxime, a second-generation cephalosporin antibiotic and potential bacteria specific infection imaging agent was evaluated. A good radiochemical purity (95%) of the labelled product was obtained after filtering the reaction mixture through a 0.22 µm filter. Scintigraphy study of the purified product showed uptake in infectious lesions 45 min after injection and abscess-to-muscle ratios were found to be 1.80, 1.85 and 1.88 at 45 min, 1.5 hr and 3 hr, respectively. A versatile and reliable chromatographic technique to assess the radiochemical purity of (99m)Tc-cefuroxime has also been described.

Synthesis and evaluation of (99m)Tc-moxifloxacin, a potential infection specific imaging agent.

To synthesize and evaluate a (99m)Tc labeled fluroquinolone, moxifloxacin as a potential bacteria specific infection imaging agent. A radiolabeling formulation including moxifloxacin, [Moxicip(TM) injection, Cipla] (4mg), sodium pertechnetate and stannous chloride (5microg) gave the best radiolabeling efficiency and moderately stable labeled (99m)Tc moxifloxacin. Quality control analysis was performed by ITLC. Rats and rabbit with infectious intramuscular lesions induced in either thigh with E. Colli were used for studying biodistribution and scintigraphic imaging of the labeled product. Imaging of an infected thigh of a rabbit was performed with a gamma-camera at various intervals. A good radiolabeling efficiency (90-95%) was obtained within 5min. No purification of the labeled product was done. Labeled product retained its radiochemical purity upto 85% even at 3h. Scintigraphy showed uptake in infectious lesions at 30min after injection, which remains constant upto 3h study. Abscess-to-muscle ratios were 1.60, 1.62, 1.74 and 1.75 at 30min, 1, 2 and 3h, respectively. Thus, (99m)Tc moxifloxacin, a new potential radiopharmaceutical has been developed for infection imaging agent.