SubbaRao V. Madhunapantula

An overview on the role of dietary phenolics for the treatment of cancers

Plant derived phenolic compounds have been shown to inhibit the initiation and progression of cancers by modulating genes regulating key processes such as: (a) oncogenic transformation of normal cells; (b) growth and development of tumors; and (c) angiogenesis and metastasis. Recent studies focusing on identifying the molecular basis of plant phenolics-induced cancer cell death have demonstrated down-regulation of: (a) oncogenic survival kinases such as PI3K and Akt; (b) cell proliferation regulators that include Erk1/2, D-type Cyclins, and Cyclin Dependent Kinases (CDKs); (c) transcription factors such as NF-kβ, NRF2 and STATs; (d) histone deacetylases HDAC1 and HDAC2; and (e) angiogenic factors VEGF, FGFR1 and MIC-1. Furthermore, while inhibiting oncogenic proteins, the phenolic compounds elevate the expression of tumor suppressor proteins p53, PTEN, p21, and p27. In addition, plant phenolic compounds and the herbal extracts rich in phenolic compounds modulate the levels of reactive oxygen species (ROS) in cells thereby regulate cell proliferation, survival and apoptosis. Furthermore, recent studies have demonstrated that phenolic compounds undergo transformation in gut microbiota thereby acquire additional properties that promote their biological activities. In vitro observations, preclinical and epidemiological studies have shown the involvement of plant phenolic acids in retarding the cancer growth. However, to date, there is no clinical trial as such testing the role of plant phenolic compounds for inhibiting tumor growth in humans. More over, several variations in response to phenolic acid rich diets-mediated treatment among individuals have also been reported, raising concerns about whether phenolic acids could be used for treating cancers. Therefore, we have made an attempt to (a) address the key structural features of phenolic acids required for exhibiting potent anti-cancer activity; (b) review the reported findings about the mechanisms of action of phenolic compounds and their transformation by gut microbiota; and (c) update the toxicological aspects and anti-tumor properties of phenolic compounds and extracts containing phenolic compounds in animals.

Mucosal vaccines: a paradigm shift in the development of mucosal adjuvants and delivery vehicles.

Mucosal immune responses are the first‐line defensive mechanisms against a variety of infections. Therefore, immunizations of mucosal surfaces from which majority of infectious agents make their entry, helps to protect the body against infections. Hence, vaccinization of mucosal surfaces by using mucosal vaccines provides the basis for generating protective immunity both in the mucosal and systemic immune compartments. Mucosal vaccines offer several advantages over parenteral immunization. For example, (i) ease of administration; (ii) non‐invasiveness; (iii) high‐patient compliance; and (iv) suitability for mass vaccination. Despite these benefits, to date, only very few mucosal vaccines have been developed using whole microorganisms and approved for use in humans. This is due to various challenges associated with the development of an effective mucosal vaccine that can work against a variety of infections, and various problems concerned with the safe delivery of developed vaccine. For instance, protein antigen alone is not just sufficient enough for the optimal delivery of antigen(s) mucosally. Hence, efforts have been made to develop better prophylactic and therapeutic vaccines for improved mucosal Th1 and Th2 immune responses using an efficient and safe immunostimulatory molecule and novel delivery carriers. Therefore, in this review, we have made an attempt to cover the recent advancements in the development of adjuvants and delivery carriers for safe and effective mucosal vaccine production.

Lipid-based nanocarriers for breast cancer treatment – comprehensive review

Abstract Breast cancer is the second leading cancer-related disease as the most common non-cutaneous malignancy among women. Curative options for breast cancer are limited, therapeutically substantial and associated with toxicities. Emerging nanotechnologies exhibited the possibility to treat or target breast cancer. Among the nanoparticles, various lipid nanoparticles namely, liposomes, solid lipid nanoparticles, nanostructured lipid carriers and lipid polymer hybrid nanoparticles have been developed over the years for the breast cancer therapy and evidences are documented. Concepts are confined in lab scale, which needs to be transferred to large scale to develop active targeting nanomedicine for the clinical utility. So, the present review highlights the recently published studies in the development of lipid-based nanocarriers for breast cancer treatment.

Dipeptidyl peptidase-4 inhibition by Pterocarpus marsupium and Eugenia jambolana ameliorates streptozotocin induced Alzheimer's disease

Alzheimers disease (AD), the most common form of dementia, is characterized by the loss of normal functions of brain cells and neuronal death, ultimately leading to memory loss. Recent accumulating evidences have demonstrated the therapeutic potential of anti-diabetic agents, such as dipeptidyl peptidase-4 (DPP-4) inhibitors, for the treatment of Alzheimers disease (AD), providing opportunities to explore and test the DPP-4 inhibitors for treating this fatal disease. Prior studies determining the efficacy of Pterocarpus marsupium (PM, Fabaceae) and Eugenia jambolana (EJ, Myrtaceae) extracts for ameliorating type 2 diabetes have demonstrated the DPP-4 inhibitory properties indicating the possibility of using of these extracts even for the treating AD. Therefore, in the present study, the neuroprotective roles of PM and EJ for ameliorating the streptozotocin (STZ) induced AD have been tested in rat model. Experimentally, PM and EJ extracts, at a dose range of 200 and 400mg/kg, were administered orally to STZ induced AD Wistar rats and cognitive evaluation tests were performed using radial arm maze and hole-board apparatus. Following 30 days of treatment with the extracts, a dose- and time-dependent attenuation of AD pathology, as evidenced by decreasing amyloid beta 42, total tau, phosphorylated tau and neuro-inflammation with an increase in glucagon-like peptide-1 (GLP-1) levels was observed. Therefore, PM and EJ extracts contain cognitive enhancers as well as neuroprotective agents against STZ induced AD.

Design and Development of Oral Nanoparticulated Insulin in Multiple Emulsion

The present research aimed at developing an injection-free nanoparticulated formulation in multiple emulsion form, for oral delivery of insulin, which otherwise undergoes degradation in the gastric environment if administered orally. Insulin-polymeric nanoparticles were prepared using layer by layer (LbL) adsorption method and incorporated into an emulsion to form a nanoparticulated multiple emulsion. Using 0.6 M sodium chloride, the insulin nanoaggregates of 300-400 nm size were obtained about a yield of 94%. The characteristics of a representative nanoparticle were as follows: particle size - 391.9±0.41 nm, polydispersity index -0.425, zeta potential- +20.6 mv, encapsulation efficiency- 86.7±1.42% and percentage entrapment efficiency of the insulin-polymeric nanoparticles in the inner aqueous phase of emulsion was 84.6%. The FT-IR analysis confirms that there were no drug interactions with the polymers. Stability analysis carried out for 3 months at 8-40 °C, showed only minor changes at the end period. The release kinetics of the nanoparticulated multiple emulsion at pH 7.4 followed first order kinetics and obeyed the Fickian law. However, at pH 2.0 the release kinetics from nanoparticulated multiple emulsion followed zero order kinetics without obeying to the Fickian law. In conclusion, our data demonstrate that the nanoparticulated multiple emulsion formulation has good release characteristics and imparted a tolerable protection for insulin at different pH conditions, which may be exploited for oral administration.

Exploring the anti-breast cancer potential of flavonoid analogs

In the course of our search for new antitumor agents for breast cancer, novel flavone derivatives were synthesized, characterized and examined for their antitumor activities against breast cancer cell lines. In initial screening, analogs 7a [3-(5-amino-1,3,4-thiadiazol-2-yl)methoxy-2-phenyl-4H-chromen-4-one] and 7b [3-(5-amino-1,3,4-thiadiazol-2-yl)methoxy-2-(4-methoxyphenyl)-4H-chromen-4-one] were found to be effective against the estrogen receptor negative cell line (MDA-MB 453), which was followed by their evaluation in five dose assays. In addition, mechanistic studies of 7a and 7b were performed by cytometric analysis and electrophoretic studies and it was observed that apoptosis is a mechanism of cell death, confirmed morphologically by acridine orange/ethidium bromide double staining and TUNEL analysis. Further in vivo evaluation of the anti-tumor activity of compound 7a and 7b by Ehrlich Ascites Carcinoma (EAC) model and related studies confirms the anti-breast cancer potential of flavonoid analogs.

Induction of colon and cervical cancer cell death by cinnamic acid derivatives is mediated through the inhibition of Histone Deacetylases (HDAC)

Recent studies from our group and many others have shown the ability of histone deacetylase (HDAC) inhibitors for retarding the growth of carcinomas of cervix, colon and rectum in vitro. A search for naturally occurring HDAC inhibitors continues due to the adverse effects associated with known HDAC inhibitors like SAHA and TSA. Therefore in the current study, naturally occurring cinnamic acids derivatives were screened for HDAC inhibitory effect using in silico docking method which identified cinnamic acids as potential candidates. Cinnamic acids (CA) are naturally occurring phenolic compounds known to exhibit anticancer properties. However, it is not clearly known whether the anticancer properties of CA derivatives are due to the inhibition of oncogenic HDACs, if so how the efficacy varies among various CA derivatives. Hence, the HDAC inhibitory potential of CA derivatives containing increasing number of hydroxylic groups or methoxy moieties was determined using Discovery Studio software and the most potent CA derivatives tested ex vivo (biochemical assay) as well as in vitro (using cell based assay). Among CA derivatives tested, dihydroxy cinnamic acid (DHCA, commonly known as caffeic acid) exhibited better interactions with HDAC2 (compared to other isoforms) in silico and inhibited its activity ex vivo as well as in vitro. Targeted reduction of HDAC activity using DHCA induced death of cancer cells by (a) generating reactive oxygen species, (b) arresting cells in S and G2/M phases; and (c) induction of caspase-3 mediated apoptosis. In conclusion, we demonstrated that DHCA inhibited cancer cell growth by binding to HDAC followed by the induction of apoptosis.

Application of quality-by-design approach to optimize diallyl disulfide-loaded solid lipid nanoparticles.

Abstract The current work was carried out by the principles of quality-by-design approach to develop an optimized solid lipid nanoparticles (SLNs) formulation of diallyl disulfide (DADS) through systematic statistical study. And its antitumor activity of DADS was also evaluated on breast cancer cell lines. To understand the effect of formulation variables (critical parameters) on the responses (critical quality attributes) of SLN, a 3-factor, 3-level Box–Behnken design, was explored to predict the responses such as particle size (Y1) and % entrapment efficiency (EE) (Y2) when concentration of surfactant (X1), amount of lipid (X2), and volume of solvent (X3) were selected as independent variables. Particle size analysis revealed that all the batches were within the nanometer range. DADS was released from the SLN much more rapidly at pH 4.5 than at pH 7.4, which is a desirable characteristic for tumor-targeted drug delivery. The cytotoxicity, reactive oxygen species (ROS), determination revealed that the antitumor activity of DADS is enhanced with SLN compared to DADS-free drug, and apoptosis is the mechanism underlying the cytotoxicity. The present study indicated the remarkable potential of DADS-SLN in enhancing the anticancer effect of DADS in breast cancer cells in vitro.

Naturally occurring benzoic acid derivatives retard cancer cell growth by inhibiting histone deacetylases (HDAC)

ABSTRACT Histone deacetylases (HDACs), which modulate the expression of genes, are potential therapeutic targets in several cancers. Targeted inhibition of HDAC prevents the expression of oncogenes thereby help in the treatment of cancers. Hence, several pharmaceutical companies developed inhibitors of HDAC and tested them in preclinical models and in clinical trials. SAHA (suberanilohydroxamic acid) is one such HDAC inhibitor developed for treating breast and colorectal carcinomas. However, due to poor efficacy in clinical trials the utility of SAHA for treating cancers was discouraged. Similarly another HDAC inhibitor Trichostatin-A (TSA) also showed promising results in clinical trials but exhibited severe adverse effects, which dampened the interest of using this molecule for cancer treatment. Therefore, search for developing a potent HDAC inhibitor with minimal side effects still continues. Hence, in this study we have screened benzoic acid and benzoic acid derivatives with hydroxylic (-OH) groups and methoxy (-OCH3) groups for their efficacy to bind to the TSA binding site of HDAC using molecular docking studies. Molecules that showed much stronger affinity (than TSA) to HDAC were tested for inhibiting HDAC expressing cultured cancer cells. DHBA but not Dimethoxy Benzoic Acid (DMBA) inhibited HDAC activity, leading to cancer cell growth inhibition through the induction of ROS and cellular apoptosis mediated by Caspase-3. In addition, DHBA arrested cells in G2/M phase of the cell cycle and elevated the levels of sub-G0-G1 cell population. In summary, results of this study report that DHBA could be a strong HDAC inhibitor and inhibit cancer cell growth more effectively.

In vitro anticancer activity of ethanolic extracts of Piper nigrum against colorectal carcinoma cell lines

Background: Piper nigrum (PN) is well known for its cytotoxic and pharmacological benefits. However, there is minimal documented evidence about its cytotoxic efficacy against colorectal carcinoma. We therefore sought to procure a precisely quantitative and qualitative result, pertaining the efficacy of an ethanolic extract of PN (EEPN) against colorectal carcinoma. Materials and Methods: EEPN was prepared by subjecting dried PN seeds to gradient ethanol fractionation. The total phenol content (TPC), antioxidant activity (AOA), and anti-inflammatory activity (AIA) were determined using Folin-Ciocalteu assay, ferric reducing ability of plasma and 2, 2-diphenyl-1-picrylhydrazyl methods, and human red blood cells membrane stabilizing assay, respectively. Colorectal carcinoma cell lines (HCT-116, HCT-15, and HT-29) were procured from National Centre for Cell Science, Pune, and were cultured in Dulbecco′s modified eagle media supplemented with 10% fetal bovine serum and 1 mM L-glutamine. Cells were seeded into a 96-well plate, followed by treatment with increasing concentrations of EEPN. The cytotoxic efficacy was evaluated based on percentage inhibition of cells, using sulforhodamine-B assay. The IC-50 values were calculated using Prism software (Prism from GraphPad software, Inc. CA, USA). Results: Biochemical analysis revealed that 50% EEPN exhibited higher TPC, AOA, and AIA when compared to 70% and 100% EEPN at any given concentration (P = 0.041). Cytotoxic analysis revealed a dose-dependent response with maximum cellular inhibition at TPC of 6 and 3 μg/ml, using 50% EEPN. However, 50% inhibition of cellular growth using 50% EEPN was seen with TPC of 3.2, 2.9, and 1.9 μg/ml at 24, 48, and 72 h, respectively, in HCT-15 cells. Hence, time- and dose-dependent increase in the cytotoxic efficacy of 50% EEPN against colorectal carcinoma cell lines were noted (P < 0.001). Conclusion: Given the significantly positive correlations exhibited between the biochemical and the cytotoxic properties evaluated in our study, we hereby conclude PN as a novel therapeutic spice for the treatment of colorectal carcinoma.