Biodegradable nanocarrier of gemcitabine and tocopherol succinate synergistically ameliorates anti-proliferative response in MIA PaCa-2 cells.

Gemcitabine (GEM) is an important chemotherapeutic agent used alone or in combination with other anticancer agents for the treatment of various solid tumors. In this study, the potential of a dietary supplement, α-tocopherol succinate (TOS) was investigated in combination with GEM by utilizing human serum albumin-based nanoparticles (HSA NPs). The developed nanoparticles were characterized using DLS, SEM and FTIR and evaluated in a panel of cell lines to inspect cytotoxic efficacy. The ratio metric selected combination of the NPs was further investigated in human pancreatic cancer cell line (MIA PaCa-2 cells) to assess the cellular death mechanism via a myriad of biochemical and bio-analytical assays including nuclear morphometric analysis by DAPI staining, ROS generation, MMP loss, intracellular calcium release, in vitro clonogenic assay, cell migration assay, cell cycle analysis, immunocytochemical staining followed by western blotting, Annexin V-FITC and cellular uptake studies. The desolvation-crosslinking method was used to prepare the NPs. The average size of TOS-HSA NPs and GEM-HSA NPs was found to be 189.47 ± 5 nm and 143.42 ± 7.4 nm, respectively. In combination, the developed nanoparticles exhibited synergism by enhancing cytotoxicity in a fixed molar ratio. The selected combination also significantly triggered ROS generation and mitochondrial destabilization, alleviated cell migration potential and clonogenic cell survival in MIA PaCa-2 cells. Further, cell cycle analysis, Annexin-V FITC assay and caspase-3 activation, up regulation of Bax and down regulation of Bcl-2 protein confirmed the occurrence of apoptotic event coupled with the G0/G1 phase arrest. Nanocarriers based this combination also offered approximately 14-folds dose reduction of GEM. Overall, the combined administration of TOS-HSA NPs and GEM-HSA NPs showed synergistic cytotoxicity accompanied with dose reduction of the gemcitabine. These encouraging findings could have implication in designing micronutrient based-combination therapy with gemcitabine and demands further investigation.

HPLC-PDA Method for Quantification of Bioactive Compounds in Crude Extract and Fractions of Aucklandia costus Falc. and Cytotoxicity Studies against Cancer Cells.

Aucklandia costus Falc. (Synonym: Saussurea costus (Falc.) Lipsch.) is a perennial herb of the family Asteraceae. The dried rhizome is an essential herb in the traditional systems of medicine in India, China and Tibet. The important pharmacological activities reported for Aucklandia costus are anticancer, hepatoprotective, antiulcer, antimicrobial, antiparasitic, antioxidant, anti-inflammatory and anti-fatigue activities. The objective of this study was the isolation and quantification of four marker compounds in the crude extract and different fractions of A. costus and the evaluation of the anticancer activity of the crude extract and its different fractions. The four marker compounds isolated from A. costus include dehydrocostus lactone, costunolide, syringin and 5-hydroxymethyl-2-furaldehyde. These four compounds were used as standard compounds for quantification. The chromatographic data showed good resolution and excellent linearity (r2 ˃ 0.993). The validation parameters, such as inter- and intraday precision (RSD < 1.96%) and analyte recovery (97.52-110.20%; RSD < 2.00%),revealed the high sensitivity and reliability of the developed HPLC method. The compounds dehydrocostus lactone and costunolide were concentrated in the hexane fraction (222.08 and 65.07 µg/mg, respectively) and chloroform fraction (99.02 and 30.21 µg/mg, respectively), while the n-butanol fraction is a rich source of syringin (37.91 µg/mg) and 5-hydroxymethyl-2-furaldehyde (7.94 µg/mg). Further, the SRB assay was performed for the evaluation of anticancer activity using lung, colon, breast and prostate cancer cell lines. The hexane and chloroform fractions show excellent IC50 values of 3.37 ± 0.14 and 7.527 ± 0.18 µg/mL, respectively, against the prostate cancer cell line (PC-3).

Recent advances in tumor microenvironment associated therapeutic strategies and evaluation models.

As per a report of the world health organization, an estimated 9.6 million people died due to cancer in 2018, globally. Most of the cancer death attributed to the lack of early detection and effective treatment. In the case of solid tumors, various factors such as leaky vasculature, angiogenesis, interstitial fluid pressure and lymphatic drainage are important in cancer chemotherapy. The poor penetration and retention of the drug/drug delivery system in tumor tissue are most critical issues in the way of effective treatment. In this scenario, the challenges are to design the specific nano-therapeutics with the potential to penetrate inside the adverse condition of tumor microenvironment (TME) including high interstitial pressure region and abnormal vasculature. The modification of nanocarriers surfaces with enzymes, peptides, pH-responsive moieties, antibodies etc. could be a promising strategy to improve the nanocarriers penetration inside the solid tumor. The priming with the drug before the administration of nanotherapeutics may also represents an efficient approach for solid tumor treatment. Further, the growth factors including fibroblast growth factor (FGF), platelet-derived growth factor (PDGF), vascular endothelial growth factor (VEGF) and their pathways could offer potential targeting opportunities for anticancer treatment. Recently, there is a surge in various approaches and formulation design directed towards abnormal TME for more effective cancer therapy. In this review, various factors related to the poor penetration, retention and specific delivery of chemotherapeutics inside tumor cells/tissues are discussed. The emerging formulations strategies directed to the TME and various methodologies for evaluation of their efficacy are also included in this review.

Development and evaluation of folate functionalized albumin nanoparticles for targeted delivery of gemcitabine.

Gemcitabine is one of the most potent anticancer agents acting on a wide range of solid tumors, however, its use is limited by short half life and high dose leading to serious side effects. The present investigation describes the development and characterization of folate functionalized gemcitabine loaded bovine serum albumin nanoparticles (Fa-Gem-BSANPs). The nanoparticles were prepared by desolvation cross-linking technique and characterized for various parameters including morphology, particle size, zeta potential, drug loading and release profile. The particle size of Gem-BSANPs and Fa-Gem-BSANPs was found to be 159.1±5.29 and 208.7±1.80 nm, respectively. DSC and XRD analysis indicated amorphous nature of the drug within the particles. The encapsulated gemcitabine exhibited less hemolytic properties as compared to native drug. The anticancer activity of Fa-Gem-BSANPs was evaluated in folate receptor over expressing cell lines (Ovcar-5 and MCF-7) and folate receptor deficient cell line (MIAPaCa-2). The Fa-Gem-BSANPs showed superior anticancer activity as compared to Gem-BSANPs in Ovcar-5 and MCF-7 cells while no significant difference in cytotoxicity was found with MIAPaCa-2 cells. Confocal microscopy indicated facilitated intracellular uptake of Fa-Gem-BSANPs in MCF-7, which in turn result in a higher potential for apoptosis. Further, Fa-Gem-BSANPs exhibited improved anti-tumor activity in Ehrlich solid tumor model in mice. In conclusion, our study indicates that folate functionalized nanoparticles confer enhance cellular uptake and cytotoxicity for gemcitabine.

Nanodecoy system: a novel approach to design hepatitis B vaccine for immunopotentiation.

The progress toward subunit vaccines has been limited by their poor immunogenicity and limited stability. To enhance the immune response, subunit vaccines universally require improved adjuvants and delivery vehicles. In the present study, we propose the use of ceramic core based nanodecoy systems for effective immunization, which seems to exhibit a broad range of surface properties. Nanodecoy systems were prepared by self-assembling of hydroxyapatite core and cellobiose and finally the hepatitis B surface antigen (HBsAg) was adsorbed over the preformed nanodecoy systems. HBsAg loaded nanodecoy systems were characterized for size, shape and antigen loading efficiency. The effect of processing steps on the stability and integrity of HBsAg was assessed by in vitro antigenicity and SDS-PAGE experiments. Nanodecoy preparations were nanometric in size range and almost spherical in shape. SDS-PAGE studies confirmed the integrity of HBsAg protein in the formulation. Vaccine efficacy was determined in female Balb/c mice and results indicated that specific anti-HBsAg antibody titers in mice receiving nanodecoy system were more efficient than the conventional adjuvant alum followed by subcutaneous immunization. Studies also indicated that nanodecoy formulations could elicit combined Th1 and Th2 immune response. It is inferred that nanodecoy systems are a class of novel carriers and hold potential as an alternative adjuvant in vaccine technology.