Effect of Early pregnancy associated protein-1 on Spike protein and ACE2 interactions: Implications in SARS Cov-2 vertical transmission.

INTRODUCTION: Several factors influence transmission of 2019-nCoV from mother to fetus during pregnancy, thus the dynamics of vertical transmission is unclear. The role of cellular protective factors, namely a 90 KDa glycoprotein, Early pregnancy-associated protein (Epap-1), expressed by placental endothelial cells in women during early pregnancy would provide an insight into role of placental factors in virus transmission. Since viral spike protein binding to the ACE2 receptors of the host cells promotes virus invasion in placental tissue, an analysis of effects of Epap-1 on the Spike-ACE2 protein binding was studied. METHODS: Epap-1 was isolated from MTP placental tissue. Molecular interaction of Epap-1 and variants of the spike was analyzed in silco. The interaction of Epap-1 with Spike and RBD were analyzed using ELISA and immunofluorescence studies. RESULTS: The results in silico showed an interaction of Epap-1 with S-protein at RBD region involving K417, Y449, Y453, Y456, Y473, Q474, F486, Q498, N501 residues of spike with Y61, F287, I302, N303, N305, S334, N465, G467, N468 residues of Epap-1 leading to interference of S-protein and ACE2 interaction [1]. Further, the interaction is conserved among the variants. The studies in vitro confirm that Epap-1 affects S protein-ACE2 and RBD- ACE2 binding, thus suggesting that during early pregnancy, SARS CoV-2 infection may be protected by Epap-1 protein present in placental tissue. The results were further confirmed by pseudovirus expressing Spike and RBD in an infection assay. DISCUSSION: Epap-1 interferes with Spike and RBD interaction with ACE2, suggesting a possible mechanism of the antiviral environment during pregnancy.

The topoisomerase II ß -kinase associated with HIV-1 is a potential target for pyridine-bischalcones' anti-HIV-1 activity.

Topoisomerase II (TopoII) is a critical component of HIV-1 integration, proviral DNA synthesis, and reverse transcription. During HIV-1 infection, the TopoIIßkinase (TopoIIßKHIV-1) phosphorylates TopoIIß. Our earlier research demonstrated that the pyridine scaffold has potent anti-HIV-1 activity by specifically inhibiting TopoIIßKHIV-1 activity. 3D QSAR results showed the presence of molecular features for interaction with TopoIIßKHIV-1 requiring chemically induced proximity for potential interaction. In this study, the chalcone and methyl groups were added to the pyridine scaffold's core to achieve the desired proximity length between the pyridine scaffold and charged centers, which resulted in an inhibitory activity against TopoIIßKHIV-1 and viral replication. According to the findings, the TopoIIßKHIV-1activity was inhibited by the inclusion of the pyridine scaffold with the chalcone group, leading to better anti-HIV-1 activity. The water-soluble methylated pyridinium chalcones' showed significant TopoIIßKHIV-1 antagonism, anti-HIV-1 activity (from IC50 > 500 nM to ID50 25 nM), and reduced cytotoxicity (CC50 = 2 mM). These activities could be associated with the charge on the pyridine and extended proximity. Therefore, it is clear that within the scope of this work, altering the proximity length and charge centers of pyridine molecules are critical for the design and development of effective anti-HIV-1 leads, specifically targeting TopoIIßKHIV-1.

Synergistic action of lactoferrin in enhancing the safety and effectiveness of docetaxel treatment against prostate cancer.

BACKGROUND: Tumor metastasis is promoted by an immunosuppressive environment. Lactoferrin (Lf) is known to regulate immunological activity in tumor cells and inhibit processes associated with tumor metastasis. A delivery of lactoferrin with docetaxel (DTX) in prostate cancer cells in the form of DTX-loaded lactoferrin nanoparticles (DTX-LfNPs) would provide a dual activity wherein the lactoferrin affects metastasis and DTX chemotherapeutically inhibits mitosis and cell division. METHODS: DTX-LfNPs were prepared using sol-oil chemistry, and particles were characterized using transmission electron microscopy. Antiproliferation activity was analyzed in prostate cancer Mat Ly Lu cells. The target localization and efficacy of DTX-LfNPs were studied in an orthotopic prostate cancer induced by Mat Ly Lu cells in a rat model. Biomarkers were estimated using ELISA and biochemical reactions. RESULTS: DTX was loaded in pure Lf nanoparticles without involving any chemical modification and conjugation, thus when these nanoparticles are delivered in cancer cells both DTX and Lf will be present in biologically active forms. DTX-LfNps exhibit a spherical morphology of dimension of 60 ± 10 nm with DTX Encapsulation Efficiency of 62.06 ± 4.07%. Competition experiments using soluble Lf confirm that DTX-LfNPs enter prostate cancer cells through the Lf receptor. DTX-LfNPs exhibit an improved anti-proliferative activity by 2.5 times compared to DTX. Further, analysis of the bioavailability of the drug in the prostate showed that DTX-LfNPs increased drug bioavailability in the prostate by two times more than the DTX. The analysis of efficacy in the Mat Ly Lu cells-induced orthotopic prostate cancer model showed that DTX-LfNPs significantly enhanced the anti-cancer activity compared to DTX in terms of regression of weight and volume of prostate tissue, the efficacy was confirmed by histochemical analysis. Lf provides synergistic activity along with DTX in inhibiting metastasis as assessed by the reduction of lactate dehydrogenase, alkaline phosphatase, TNF alpha, and IFNγ. LfNPs facilitate higher DTX localization along with Lf-mediated protection from DTX-associated toxicity to neutrophils and kidneys as assessed by C-reactive protein, creatinine, and uric acid. Thus, DTX LfNPs show a dual action by enhancing DTX bioavailability in prostate along with Lf-mediated suppression of metastasis as well as DTX-associated toxicity. CONCLUSION: In conclusion, DTX-LfNPs enhance the bioavailability of DTX in the prostate along with Lf-assisted improvement in inhibition of tumor metastasis and drug-associated toxicity.

Carboplatin- and Etoposide-Loaded Lactoferrin Protein Nanoparticles for Targeting Cancer Stem Cells in Retinoblastoma In Vitro.

Purpose: Cancer stem cells (CSCs) are known to contribute to tumor relapses by virtue of their chemoresistance. With the knowledge that nanoformulations can overcome drug resistance, we evaluated the efficacy and cytotoxicity of clinical-grade carboplatin (CPT)- and etoposide (ETP)-loaded lactoferrin nanoparticles (Lf-Nps) on total, CD133-enriched (non-CSC), and CD133-depleted (CSC) populations of retinoblastoma (Rb) Y79 cells. Methods: Physicochemical properties of drug-loaded Lf-Nps were measured with transmission electron microscopy and attenuated total reflectance-Fourier transform infrared. The encapsulation efficiency, uptake, and release of drug-loaded Lf-Nps were measured using high-performance liquid chromatography and a UV-visible spectrophotometer. Cytotoxicity of the standard and drug-loaded Lf-Nps was evaluated by the MTT assay. Results: The mean (SD) size and encapsulation efficiency of Lf-CPT and Lf-ETP were 61.2 (3.94) nm, 60% and 45.15 (5.85) nm, 38%, respectively, and the drug release efficiency was highest at pH 6. The increased drug uptake and lower release of drug-loaded Lf-Nps were observed in CSC and non-CSC populations compared to their standard forms. The relative increase of drug uptake and sustained intracellular retention of the drug-loaded Lf-Nps compared to standard drugs showed an enhanced cytotoxicity up to 50%, especially in Rb Y79 CSCs (IC50: CPT, 230.3; Lf-CPT, 118.2; ETP, 198.1; and Lf-ETP, 129) compared to non-CSCs. Conclusions: Our study documents an increase in drug uptake, retention, and cytotoxicity of Lf-CPT and Lf-ETP on Y79 CSCs and non-CSCs as compared to their standard drugs in vitro. The reversal of chemoresistance in the CSC population by nanoformulation appears promising with the potential to pave the way for improved targeted therapy and better clinical outcomes.

Design, Synthesis, and Antiviral activity of 1,2,3,4-Tetrahydropyrimidine derivatives acting as novel entry inhibitors to target at "Phe43 cavity" of HIV-1 gp120.

The HIV-1 invasion is initiated with the interaction of viral glycoprotein gp120 and cellular receptor CD4. The binding mechanism reveals two major hotspots involved in gp120-CD4 interaction. The first one is a hydrophobic cavity (Phe43 cavity) on gp120 capped with phenyl ring of phe43CD4 and the second is the electrostatic interaction between positive charge of Arg59CD4 and negative charge of Asp368gp120. Targeting these hotspots, small molecules for entry inhibition and HIV-1 neutralization were designed and tested. In the process, pyrimidine derivatives were identified as potent molecules to intercept gp120-CD4 binding by targeting both the hotspots. Herein, the synthesis, characterization of 1,2,3,4-Tetrahydropyrimidine derivatives, and biological evaluation on 93IN101, a clade C virus are presented. The paper presents a novel set of entry inhibitors to target dual hotspots on gp120 to inhibit protein-protein interactions.

Design, synthesis, and evaluation of HIV-1 entry inhibitors based on broadly neutralizing antibody 447-52D and gp120 V3loop interactions.

The third variable loop region (V3 loop) on gp120 plays an important role in cellular entry of HIV-1. Its interaction with the cellular CD4 and coreceptors is an important hallmark in facilitating the bridging by gp41 and subsequent fusion of membranes for transfer of viral genetic material. Further, the virus phenotype determines the cell tropism via respective co- receptor binding. Thus, coreceptor binding motif of envelope is considered to be a potent anti-viral drug target for viral entry inhibition. However, its high variability in sequence is the major hurdle for developing inhibitors targeting the region. In this study, we have used an in silico Virtual Screening and "Fragment-based" method to design small molecules based on the gp120 V3 loop interactions with a potent broadly neutralizing human monoclonal antibody, 447-52D. From the in silico analysis a potent scaffold, 1,3,5-triazine was identified for further development. Derivatives of 1,3,5-triazine with specific functional groups were designed and synthesized keeping the interaction with co-receptor intact. Finally, preliminary evaluation of molecules for HIV-1 inhibition on two different virus strains (clade C, clade B) yielded IC50 < 5.0 µM. The approach used to design molecules based on broadly neutralizing antibody, was useful for development of target specific potent antiviral agents to prevent HIV entry. The study reported promising inhibitors that could be further developed and studied.

Topoisomerase II ß Gene Specific siRNA Delivery by Nanoparticles Prepared with c-ter Apotransferrin and its Effect on HIV-1 Replication.

Topoisomerase II beta (Topo IIß) is one of the two isoforms of type II topoisomerases present in higher eukaryotes. This 180 kDa nuclear protein involves in different cellular processes like transcription, recombination, etc., apart from its normal topological functions. Previously, we have reported the association of this isoform along with the other isoform topoisomerase II alpha (Topo IIα) with HIV-1 reverse transcription complex and the downregulation of Topo IIß expression resulted in incomplete reverse transcription. In this study, we have tested the Topo IIß specific siRNA delivery using protein nanoparticles prepared with c-terminal domine of transferrin (c-ter) for the first time. Results show that, c-ter nanoparticles resemble apotransferrin nanoparticles in drug holding capability and drug delivery but with small in size. Topo IIß specific siRNA delivered in the form of c-ter nanoformulation resulted in knockdown of Topo IIß expression for the prolonged periods and which intern resulted in decreased viral replication of HIV-1.

Lactoferrin nanoparticles coencapsulated with curcumin and tenofovir improve vaginal defense against HIV-1 infection.

Aim: We report here the development of tenofovir- and curcumin-loaded lactoferrin nanoparticles (TCNPs) as an HIV-microbicide. Materials & methods: TCNPs were subjected to various physicochemical characterization experiments, followed by in vitro and in vivo experiments to assess their efficacy. Results: TCNPs had a diameter of 74.31 ± 2.56 nm with a gross encapsulation of more than 61% for each drug. Nanoparticles were effective against HIV-1 replication, with an IC50 of 1.75 µM for curcumin and 2.8 µM for tenofovir. TCNPs provided drug release at the application site for up to 8-12 h, with minimal leakage into the systemic circulation. TCNPs showed spermicidal activity at ≥200 µM and induced minimal cytotoxicity and inflammation in the vaginal epithelium as revealed by histopathological and ELISA studies. Conclusion: We demonstrated that TCNPs could serve as a novel anti-HIV microbicidal agent in rats. [Formula: see text].

Clinical and whole genome characterization of SARS-CoV-2 in India.

We report clinical profile of hundred and nine patients with SARS CoV-2 infection, and whole genome sequences (WGS) of seven virus isolates from the first reported cases in India, with various international travel histories. Comorbidities such as diabetes, hypertension, and cardiovascular disease were frequently associated with severity of the disease. WBC and neutrophil counts showed an increase, while lymphocyte counts decreased in patients with severe infection suggesting a possible neutrophil mediated organ damage, while immune activity may be diminished with decrease in lymphocytes leading to disease severity. Increase in SGOT, SGPT and blood urea suggests the functional deficiencies of liver, heart, and kidney in patients who succumbed to the disease when compared to the group of recovered patients. The WGS analysis showed that these isolates were classified into two clades: I/A3i, and A2a (four according to GISAID: O, L, GR, and GH). Further, WGS phylogeny and travel history together indicate possible transmission from Middle East and Europe. Three S protein variants: Wuhan reference, D614G, and Y28H were identified predicted to possess different binding affinities to host ACE2.

Targeting cancer with lactoferrin nanoparticles: recent advances.

Lactoferrin, an iron storage protein, is known for its microbicidal activity and its ability to modulate the immune system, mediated through specific interactions with receptors on cell surfaces for internalization. These activities confer a significant versatility to lactoferrin, presenting it as a targeting ligand to disease-bearing cells. Early efforts in developing targeted delivery systems have focused on nano- and microcomposites comprised of metal and polymeric materials. These can be targeted through conjugation or adsorption of lactoferrin to achieve recognition to receptor-expressing cells. More recently, efforts are underway to utilize lactoferrin itself as a medium in loading the therapeutic agent. The functional efficiency of drug-loaded lactoferrin nanoparticles has been evaluated in different disease conditions such as cancer, HIV, Parkinson's disease, etc. This review will present the details of composition and performance of various delivery systems designed and developed using lactoferrin as targeting agent for the treatment of cancer.

Sulfonate modified Lactoferrin nanoparticles as drug carriers with dual activity against HIV-1.

Intriguing properties and structural dynamics of Lactoferrin have been exploited in numerous applications, including its use as self-assembling, pH sensitive nanoparticles to deliver intended cargo at the disease site. In this study, we explore the possibility of surface modification of Lactoferrin nanoparticles to hone its specificity to target HIV-1 infected cells. Existence of free cysteine groups on Lactoferrin nanoparticles available for reaction with external molecules facilitates conjugation on the surface with Sodium 2-mercaptoethanesulfonate (MES). Conjugation with MES is used to edge a negative charge that can mimic CCR5 and Heparan sulfate (initial point of contact of HIV-1 env to host cell surface) electrostatic charge (Sulfate group). A simple sono-chemical irradiation method was employed for self-assembly of Nanoparticles and for surface modification. The nanoparticles serve dual purpose to abrogate extracellular entry and to target viral enzymes, when loaded with ART drugs. The morphology and size distribution of the formed particles were explored using Transmission Electron Microscope (TEM), Scanning Electron Microscope (SEM) and Dynamic Light Scattering. Raman SERS was employed to understand the difference in the protein upon surface modification. The anti-HIV property of the particles was confirmed in-vitro. The modified device demonstrated acceptable nanoparticle properties with controlled release and higher effective concentration in the area of infection.

Discovery of dual cation-π inhibitors of acetylcholinesterase: design, synthesis and biological evaluation.

BACKGROUND: Alzheimer's disease (AD) is a widespread dementia-related disease affecting mankind worldwide. A cholinergic hypothesis is considered the most effective target for treating mild to moderate AD. Present study aims to identify new scaffolds for inhibiting acetylcholinesterase activity. METHODS: To find Acetylcholinesterase (AChE) inhibitors, we computationally designed and chemically synthesized a series of cation-π inhibitors based on novel scaffolds that potentially block AChE. The cytotoxic effect of inhibitors were determined by MTT. AChE inhibition experiment was performed by Ellman and the Amplex red method in the SH-SY5Y cell line. Further, the experimental data on designed compounds corroborate with various computational studies that further elucidate the binding mode of interactions and binding affinity. RESULTS: The inhibitors were designed to promote dual binding and were incorporated with groups that may facilitate any of the cation- π, hydrophobic and hydrogen-bonding interactions with the conserved and hot-spot residues in the binding site. The inhibitors possessing pyridine-N-methylated pyridinium group and thereby involved in cation- π interactions are highly active relative to the marketed drug Donepezil as well as the designed analogs that lack the group. In vitro enzymatic Ellman assay and Amplex red assay on SH-SY5Y cell line estimated IC50 of the designed compounds in nM range with one having binding affinity higher than Donepezil. Compounds exhibit no significant toxicity up to µM range. CONCLUSIONS: Compounds possessing methylidenecyclohexanone scaffolds, with characteristic dual-binding and involving strong cation-π interactions, serves as new leads for AChE and opens a new direction for drug discovery efforts.

Aurora kinase B siRNA-loaded lactoferrin nanoparticles potentiate the efficacy of temozolomide in treating glioblastoma.

AIM: To investigate the efficacy of lactoferrin nanoparticles (LfNPs) in delivering siRNA across the blood-brain barrier to treat glioblastoma multiforme (GBM) and with an additional objective of potentiation of conventional temozolomide (TMZ) chemotherapy. METHODS: Aurora kinase B (AKB) siRNA-loaded nanoparticles (AKB-LfNPs) were prepared with milk protein, lactoferrin, by water in oil emulsion method. AKB-LfNPs were tested in cell lines and in GBM orthotopic mouse model with and without TMZ treatment. RESULTS: AKB silencing, cytotoxicity and cell cycle arrest by these LfNPs were shown to be effective on GL261 cells. Tumor growth was significantly lower in AKB-LfNPs alone and in combination with TMZ treated mice and increased the survival by 2.5-times. CONCLUSION: Treatment of AKB-LfNPs to GBM mice improves life expectancy and has potential to combine with conventional chemotherapy.

Development of pyridine dicoumarols as potent anti HIV-1 leads, targeting HIV-1 associated topoisomeraseIIß kinase.

AIM: A structural study of a series of pyridine dicoumarol derivatives with potential activity against a novel Topoisomerase IIß kinase which was identified in the HIV-1 viral lysate, compounds were designed and synthesized based on a 3D-QSAR study. MATERIALS & METHODS: Based on QSAR model we have designed and synthesized a series of pyridine dicoumarol derivatives and characterized by spectral studies, all the molecules are biologically evaluated by kinase assay, cytotoxicity assay, ELISA and PCR method. RESULT: We demonstrated the achievement of water soluble disodium pyridine dicoumarate derivatives showing high anti-HIV-1 activity (IC50 <25 nM) which provides a crucial point for further development of pyridine dicoumarol series as HIV-1-associated topoisomerase IIß kinase inhibitors for clinical application against AIDS. CONCLUSION: A new class of anti-HIV-1 lead compounds have been designed and tested. Further studies would result in development of  novel and potential drugs.

Overcoming blood brain barrier with a dual purpose Temozolomide loaded Lactoferrin nanoparticles for combating glioma (SERP-17-12433).

Targeted delivery of drugs to the brain is challenging due to the restricted permeability across the blood brain barrier (BBB). Gliomas are devastating cancers and their positive treatment outcome using Temozolomide (TMZ) is limited due to its short plasma half-life, systemic toxicity and limited access through the blood-brain barrier (BBB). Nanoparticles made of Lactoferrin (Lf) protein, have been shown to enhance the pharmacological properties of drugs. Here, we report the specific ability of Lf nanoparticles to cross BBB and target over-expressed Lf receptors on glioma for enhanced TMZ delivery. TMZ-loaded Lf nanoparticles (TMZ-LfNPs) were prepared by our previously reported sol-oil method. While the Lf protein in the NP matrix aids in transcytosis across the BBB and preferential tumor cell uptake, the pH responsiveness leads to TMZ release exclusively in the tumor microenvironment. Delivery through LfNPs results in an enhanced and sustained intracellular concentration of TMZ in GL261 cells in vitro along with improving its in vivo pharmacokinetics and brain accumulation. TMZ-LfNPs treatment results in a significant reduction of tumor volume, higher tumor cell apoptosis and improved median survival in glioma bearing mice. These results demonstrate that LfNPs present an efficient TMZ delivery platform for an effective treatment of gliomas.

Topoisomerase IIß and its role in different biological contexts.

Topoisomerase IIß is a type II DNA topoisomerase that was reported to be expressed in all mammalian cells but abundantly expressed in cells that have undergone terminal differentiation to attain a post mitotic state. Enzymatically it catalyzes ATP-dependent topological changes of double stranded DNA, while as a protein it was reported to be associated with several factors in promoting cell growth, migration, DNA repair and transcription regulation. The cellular roles of topoisomerase IIß are very less understood compared to its counterpart topoisomerase IIα. This review discusses origin of Topoisomerase II beta, its structure, activities reported in vitro and in vivo along with implications in cellular processes namely transcription, DNA repair, neuronal development, aging, HIV-infection and cancer.

Triple Drug Combination of Zidovudine, Efavirenz and Lamivudine Loaded Lactoferrin Nanoparticles: an Effective Nano First-Line Regimen for HIV Therapy.

PURPOSE: To enhance efficacy, bioavailability and reduce toxicity of first-line highly active anti-retroviral regimen, zidovudine + efavirenz + lamivudine loaded lactoferrin nanoparticles were prepared (FLART-NP) and characterized for physicochemical properties, bioactivity and pharmacokinetic profile. METHODS: Nanoparticles were prepared using sol-oil protocol and characterized using different sources such as FE-SEM, AFM, NanoSight, and FT-IR. In-vitro and in-vivo studies have been done to access the encapsulation-efficiency, cellular localization, release kinetics, safety analysis, biodistribution and pharmacokinetics. RESULTS: FLART-NP with a mean diameter of 67 nm (FE-SEM) and an encapsulation efficiency of >58% for each drug were prepared. In-vitro studies suggest that FLART-NP deliver the maximum of its payload at pH5 with a minimum burst release throughout the study period with negligible toxicity to the erythrocytes plus improved in-vitro anti-HIV activity. FLART-NP has improved the in-vivo pharmacokinetics (PK) profiles over the free drugs; an average of >4fold increase in AUC and AUMC, 30% increase in the Cmax, >2fold in the half-life of each drug. Biodistribution data suggest that FLART-NP has improved the bioavailability of all drugs with less tissue-related inflammation as suggested with histopathological evaluation CONCLUSIONS: The triple-drug loaded nanoparticles have various advantages against soluble (free) drug combination in terms of enhanced bioavailability, improved PK profile and diminished drug-associated toxicity.

Lactoferrin nanoparticle mediated targeted delivery of 5-fluorouracil for enhanced therapeutic efficacy.

Malignant melanoma is an aggressive form of skin cancer with high mortality rates. Common treatments for malignant melanoma involve a combination of radiotherapy and chemotherapy with fluorouracil (5-FU). A major challenge with melanoma treatment is active resistance to chemotherapeutic drugs. Superior treatment outcome lies on balance involving optimum therapeutic doses and the side effects associated with dose escalation. The study aimed to efficiently entrap 5-FU in lactoferrin nanoparticles (LfNPs) in an attempt to enhance its therapeutic efficacy. 5-FU loaded lactoferrin nanoparticles (5-FU-LfNPs) were prepared by sol-oil method with a narrow size distribution of 150±20nm 5-FU-LfNPs exhibits high encapsulation efficiency (64±2.3%) and increased storage stability at 4°C. Competitive ligand binding and lysosomal colocalization studies suggested a receptor mediated uptake for LfNPs internalization in B16F10 cells. Moreover, 5-FU-LfNPs show a pH dependent drug release similar to endosomal pH (pH 5 and 6). In addition compared to free 5-FU, 5-FU- LfNPs showed a higher intracellular uptake, prolonged retention and improved cytotoxicity (2.7-fold) in melanoma cells (B16F10). To conclude, LfNPs represent a superior nano-carrier for the targeted delivery of 5-FU in melanoma cells intended for the efficient treatment of melanoma though detailed in vivo investigations are warranted.

Triple combination MPT vaginal microbicide using curcumin and efavirenz loaded lactoferrin nanoparticles.

We report that a combination of anti-HIV-1 drug efavirenz (EFV), anti-microbial-spermicidal curcumin (Cur) and lactoferrin nanoparticles (ECNPs) act as MPT formulation. These nanoparticles are of well dispersed spherical shape with 40-70 nm size, with encapsulation efficiency of 63 ± 1.9% of Cur &61.5% ± 1.6 of EFV, significantly higher than that of single drug nanoparticles (Cur, 59 ± 1.34%; EFV: 58.4 ± 1.79). ECNPs were found to be sensitive at pH 5 and 6 and have not effected viability of vaginal micro-flora, Lactobacillus. Studies in rats showed that ECNPs delivers 88-124% more drugs in vaginal lavage as compared to its soluble form, either as single or combination of EFV and Cur. The ECNPs also shows 1.39-4.73 fold lower concentration of absorption in vaginal tissue and plasma compared to soluble EFV + Cur. Furthermore, ECNPs show significant reduction in inflammatory responses by 1.6-3.0 fold in terms of IL-6 and TNF-α in vaginal tissue and plasma compared to soluble EFV + Cur. ECNPs showed improved pharmacokinetics profiles in vaginal lavage with more than 50% of enhancement in AUC, AUMC, Cmax and t1/2 suggesting longer exposure of Cur and EFV in vaginal lavage compared to soluble EFV + Cur. Histopathological analysis of vaginal tissue shows remarkably lower toxicity of ECNPs compared to soluble EFV + Cur. In conclusion, ECNPs are significantly safe and exhibit higher bioavailability thus constitute an effective MPT against HIV.

TopoisomeraseIIß in HIV-1 transactivation.

TopoisomeraseIIß, an isoform of type II topoisomerase, was found to be functional in various viral infections. Its plausible role in HIV life cycle was also suggested earlier, but not clearly established. In the present study, we have investigated the role of TopoIIß in HIV-1 infection by its gain and loss of function. Overexpression of TopoIIß lead to an increase in viral replication, resulting in enhanced virion production. HIV-1 replication was impaired when TopoIIß was down regulated by siRNA and inhibited by ICRF-193 and merbarone. The role of TopoIIß in HIV-1 transcription was shown through its interaction with Tat and recruitement to long terminal repeat (LTR) region by co-immunoprecipitation and ChIP assays. Involvement of TopoIIß in transactivation of HIV-1 LTR was confirmed by luciferase assay in reporter cell line, TZM bl and also by transfection of reporter exogenously. It was also observed that LTR transactivation commensurated with the expression of TopoIIß in the presence of Tat. In addition, a decreased viral gene expression on treatment with merbarone exemplifies the importance of catalytic activity of TopoIIß in viral replication. These observations indicate that TopoIIß is involved in the cascade of coactivator complexes that are recruited to LTR for regulation of HIV-1 transcription.