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.
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
99m
Tc-labeled new BN peptide analogs. Four new BN analogs, HYNIC-Asp[PheNle]BN(7-14)NH
2
, BN1; HYNIC-Pro-Asp[TyrMet]BN(7-14)NH
2
, BN2; HYNIC-Asp-Asn[Lys-CHAla-Nle]BN(7-14)NH
2
, BN3; and DOMA-GABA[Pro-Tyr-Nle]BN(7-14)NH
2
, 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-NH
2
(BNS). Lipophilicity, serum stability, internalization, and binding affinity studies were carried out using
99m
Tc-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.
Electronic supplementary material
The online version of this article (10.1186/s13550-019-0493-x) contains supplementary material, which is available to authorized users.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.