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Breast cancer is one of the most commonly diagnosed cancers in American women. Triple negative breast cancer is among the most advanced and aggressive forms of breast cancer. Treatment options are limited for such cancers, making chemotherapy a convenient and effective treatment. Although these therapies can reduce morbidity and mortality, it is often followed by systemic side effects or relapse. Nanoparticles (NPs) have been considered for drug delivery approaches due to their ability to target various disease sites. Herein, we aim to develop a biomimetic NP construct (cell membrane-cloaked NPs) that exhibits specific affinity with triple negative breast cancer cells. In this regard, we designed biomimetic supramolecular nanoconstructs composed of a poly(vinyl pyrrolidone)-tannic acid (PVP-TA NPs/ PVT NPs) core and biofunctionalized with neutrophil cell membranes (PVT-NEU NPs). In this study, we have synthesized a PVT-NEU NP construct, characterized it, and evaluated it for improved targeting and therapeutic benefits in

The management of aggressive breast cancer, particularly, triple negative breast cancer (TNBC) remains a formidable challenge, despite treatment advancement. Although newer therapies such as atezolizumab, olaparib, and sacituzumab can tackle the breast cancer prognosis and/or progression, but achieved limited survival benefit(s). The current research efforts are aimed to develop and implement strategies for improved bioavailability, targetability, reduce systemic toxicity, and enhance therapeutic outcome of FDA-approved treatment regimen. This review presents various nanoparticle technology mediated delivery of chemotherapeutic agent(s) for breast cancer treatment. This article also documents novel strategies to employ cellular and cell membrane cloaked (biomimetic) nanoparticles for effective clinical translation. These technologies offer a safe and active targeting nanomedicine for effective management of breast cancer, especially TNBC., Graphical abstract Image 1, Highlights • The management of aggressive breast cancer remains challenge. • Various nanoparticle technology mediated delivery of chemotherapeutic agent(s) for breast cancer treatment. • Novel strategies to employ cellular and cell membrane cloaked nanoparticles for effective clinical translation.

Background: Adequate bioimaging is crucial in cancer management in many ways including screening, detection, characterization, staging and grading, therapy response, surgical guidance, and margins assessment. Indocyanine green (ICG) is one of the FDA-approved near infra-red fluorescent (NIRF) probe for cancer imaging and image-guided surgery in clinical setting. However, limitations of ICG includes poor photostability, high concentration toxicity, short circulation time, and poor cancer cell specificity. To overcome these hurdles, we engineered a nanoconstruct composed of poly(vinyl pyrrolidone) (PVP)-indocyanine green that is cloaked self-assembled with tannic acid (termed as ICG-Glow NPs) for the cancer cells/tissues specific targeting. Methods: Pursuing the novel nanotherapy approach, our lab has developed PVP-TA based ICG (PVT-ICG) fluorescent nanoparticles via self-assembly process. Our optimized PVT-ICG nanoformulation was further characterized for its physicochemical properties. An IVIS imaging system was further used to measure NIR fluorescence of novel PVT-ICG. Moreover, Human cancer (Breast, Pancreatic, Liver and Prostate) tissue microarrays (TMAs) were histochemically stained to assess cancer cell targeting/specificity of PVT-ICG. Results: PVT-ICG indicated particle size and surface charge ideal for cancer cell/tissue delivery. PVT-ICG, further, demonstrated improved photostability and fluorescent intensity. Additionally, TMA studies exhibited enhanced internalization and cancer targeting/specificity of PVT-ICG nanoparticles compared to free ICG dye in all cancers. Conclusion: Collectively, our findings suggest that this NIR fluorescent probe PVT-ICG has great potential for becoming a novel and safe imaging modality for various types of cancer cells and tumors which can result in early cancer diagnosis leading to improved disease management. Citation Format: Neeraj Chauhan, Marco Cabrera, Pallabita Chowdhury, Prashanth K. Nagesh, Anupam Dhasmana, Meena Jaggi, Subhash C. Chauhan, Murali M. Yallapu. Illuminating cancer cells with a novel nano fluorescent NIR probe for bioimaging [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 826.

Cisplatin is a commonly used chemotherapeutic agent to treat a wide array of cancers that is frequently associated with toxic injury to the kidney due to oxidative DNA damage and perturbations in cell cycle progression leading to cell death. In this study, we investigated whether thyroid receptor interacting protein 13 (TRIP13) plays a central role in the protection of the tubular epithelia following cisplatin treatment by circumventing DNA damage. Following cisplatin treatment, double-stranded DNA repair pathways were inhibited using selective blockers to proteins involved in either homologous recombination or non-homologous end joining. This led to increased blood markers of acute kidney injury (AKI) (creatinine and neutrophil gelatinase-associated lipocalin), tubular damage, activation of DNA damage marker (γ-H2AX), elevated appearance of G2/M blockade (phosphorylated histone H3 Ser10 and cyclin B1), and apoptosis (cleaved caspase-3). Conditional proximal tubule-expressing Trip13 mice were observed to be virtually protected from the cisplatin nephrotoxicity by restoring most of the pathological phenotypes back toward normal conditions. Our findings suggest that TRIP13 could circumvent DNA damage in the proximal tubules during cisplatin injury and that TRIP13 may constitute a new therapeutic target in protecting the kidney from nephrotoxicants and reduce outcomes leading to AKI.

Cellular senescence is one of the prevailing issues in cancer therapeutics that promotes cancer relapse, chemoresistance, and recurrence. Patients undergoing persistent chemotherapy often develop drug-induced senescence. Docetaxel, an FDA-approved treatment for prostate cancer, is known to induce cellular senescence which often limits the overall survival of patients. Strategic therapies that counter the cellular and drug-induced senescence are an unmet clinical need. Towards this an effort was made to develop a novel therapeutic strategy that targets and removes senescent cells from the tumors, we developed a nanoformulation of tannic acid−docetaxel self-assemblies (DSAs). The construction of DSAs was confirmed through particle size measurements, spectroscopy, thermal, and biocompatibility studies. This formulation exhibited enhanced in vitro therapeutic activity in various biological functional assays with respect to native docetaxel treatments. Microarray and immunoblot analysis results demonstrated that DSAs exposure selectively deregulated senescence associated TGFβR1/FOXO1/p21 signaling. Decrease in β-galactosidase staining further suggested reversion of drug-induced senescence after DSAs exposure. Additionally, DSAs induced profound cell death by activation of apoptotic signaling through bypassing senescence. Furthermore, in vivo and ex vivo imaging analysis demonstrated the tumor targeting behavior of DSAs in mice bearing PC-3 xenograft tumors. The antisenescence and anticancer activity of DSAs was further shown in vivo by inhibiting TGFβR1 proteins and regressing tumor growth through apoptotic induction in the PC-3 xenograft mouse model. Overall, DSAs exhibited such advanced features due to a natural compound in the formulation as a matrix/binder for docetaxel. Overall, DSAs showed superior tumor targeting and improved cellular internalization, promoting docetaxel efficacy. These findings may have great implications in prostate cancer therapy.

Introduction: Antiretroviral therapy (ART) has led to a significant reduction in HIV-1 morbidity and mortality. Many antiretroviral drugs (ARVs) are metabolized by cytochrome P450 (CYP) pathway, and the majority of these drugs are also either CYP inhibitors or inducers and few possess both activities. These CYP substrates, when used for HIV treatment in the conventional dosage form, have limitations such as low systemic bioavailability, potential drug-drug interactions, and short half-lives. Thus, an alternative mode of delivery is needed in contrast to conventional ARVs. Areas covered: In this review, we summarized the limitations of conventional ARVs in HIV treatment, especially for ARVs which are CYP substrates. We also discussed the preclinical and clinical studies using the nanotechnology strategy to overcome the limitations of these CYP substrates. The preclinical studies and clinical studies published from 2000 to February 2019 were discussed. Expert opinion: Since preclinical and clinical studies for prevention and treatment of HIV using nanotechnology approaches have shown considerable promise in recent years, nanotechnology could become an alternative strategy for daily oral therapy as a future treatment.

Paclitaxel (PTX) is a gold standard chemotherapeutic agent for breast, ovarian, pancreatic and non-small cell lung carcinoma. However, in clinical use PTX can have adverse side effects or inadequate pharmacodynamic parameters, limiting its use. Nanotechnology is often employed to reduce the therapeutic dosage required for effective therapy, while also minimizing the systemic side effects of chemotherapy drugs. However, there is no nano formulation of paclitaxel with chemosensitization motifs built in. With this objective, we screened eleven pharmaceutical excipients to develop an alternative paclitaxel nanoformulation using a self-assembly method. Based on the screening results, we observed tannic acid possesses unique properties to produce a paclitaxel nanoformulation, i.e., tannic acid-paclitaxel (TAP). This stable TAP nanoformulation, referred to as TAP nanoparticles (TAP NPs), showed a spherical shape of 102.22±14.05 nm and negative zeta potential of −8.85±0.44 mV. The presence of PTX in TAP NPs was confirmed by Fourier Transform Infrared (FTIR) spectra, thermogravimetric analyzer (TGA) scans, and X-ray diffraction (XRD). Encapsulation efficiency of PTX in TAP NPs was determined to be > 96.49±0.43 %. Intracellular drug uptake of plain drug PTX on breast cancer cells (MDA-MB-231) shows more or less constant drug concentration from 2–6 hours, suggesting drug efflux by the P-gp transporters, over TAP NPs, in which PTX uptake was more than 95.52+11.01% in 6 hours, as analyzed by liquid chromatography-tandem mass spectrometry (LC-MS/MS). Various biological assays such as proliferation, clonogenic formation, invasion, and migration confirm superior anticancer effects of TAP NPs over plain PTX at all tested concentrations. P-gp expression Rhodamine-123 (RH123), beta-tubulin stabilization, Western blot, and microarray analysis further confirm the improved therapeutic potential of TAP NPs. These results suggest that the TAP nanoformulation provides an important reference for developing a therapeutic nanoformulation affording pronounced, enhanced effects in breast cancer therapy.

Bone metastasis occurs in the majority of cancer patients, which hampers quality of life and significantly decreases survival. Aggressive chemotherapy is a traditional treatment regimen that induces severe systemic toxicities. Therefore, bone-directed therapies are highly warranted. We report a novel nanoparticle formulation that is composed of poly(vinylpyrrolidone) and tannic acid core nanoparticles (PVT NPs) that forms self-assembly with zoledronic acid (ZA@PVT NPs). The construction of ZA@PVT NPs was confirmed by particle size, zeta potential, transmission electron microscopy, and spectral analyses. An optimized bone-targeted ZA@PVT NPs formulation showed greater binding and internalization in in vitro with metastasis prostate and breast cancer cells. ZA@PVT NPs were able to deliver ZA more efficiently to tumor cells, which inhibited proliferation of human prostate and breast cancer cells. In addition, ZA@PVT NPs were capable of targeting mouse bones and prostate tumor microarray tissues (ex vivo) while sparing all other vital organs. More importantly, ZA@PVT NPs induce chemo sensitization to docetaxel treatment in cancer cells. Overall, the study results confirm that ZA-based, bone-targeted NPs have great potential for the treatment of bone metastasis in the near future.

The use of antiretroviral therapy (ART) has remarkably decreased the morbidity associated with HIV-1 infection, however, the prevalence of HIV-1-associated neurocognitive disorders (HAND) is still increasing. The blood-brain barrier (BBB) is the major impediment for penetration of antiretroviral drugs, causing therapeutics to reach only suboptimal level to the brain. Conventional antiretroviral drug regimens are not sufficient to improve the treatment outcomes of HAND. In our recent report, we have developed a poloxamer-PLGA nanoformulation loaded with elvitegravir (EVG), a commonly used antiretroviral drug. The nanoformulated EVG is capable of elevating intracellular drug uptake and simultaneously enhance viral suppression in HIV-1-infected macrophages. In this work, we identified the clinical parameters including stability, biocompatibility, protein corona, cellular internalization pathway of EVG nanoformulation for its potential clinical translation. We further assessed the ability of this EVG nanoformulation to cross the in vitro BBB model and suppress the HIV-1 in macrophage cells. Compared with EVG native drug, our EVG nanoformulation demonstrated an improved BBB model penetration cross the in vitro BBB model and an enhanced HIV-1 suppression in HIV-1-infected human monocyte-derived macrophages after crossing the BBB model without altering the BBB model integrity. Overall, this is an innovative and optimized treatment strategy that has a potential for therapeutic interventions in reducing HAND.

Prostate cancer (PCa) cells exploit the aberrant lipid signaling and metabolism as their survival advantage. Also, intracellular storage lipids act as fuel for the PCa proliferation. However, few studies were available that addressed the topic of targeting lipid metabolism in PCa. Here, we assessed the tannic acid (TA) lipid-targeting ability and its capability to induce endoplasmic reticulum (ER) stress by reactive oxygen species (ROS) in PCa cells. TA exhibited dual effects by inhibiting lipogenic signaling and suppression of lipid metabolic pathways. The expression of proteins responsible for lipogenesis was down regulated. The membrane permeability and functionality of PCa were severely affected and caused nuclear disorganization during drug exposure. Finally, these consolidated events shifted the cell’s survival balance towards apoptosis. These results suggest that TA distinctly interferes with the lipid signaling and metabolism of PCa cells.

The goal of this investigation was to develop and demonstrate a polymer/paclitaxel self-assembly (PTX-SA) formulation. Polymer/PTX-SAs were screened based on smaller size of formulation using dynamic light scattering analysis. Additionally, fluorescence microscopy and flow cytometry studies exhibited that polyvinylpyrrolidone (PVP)-based PTX-SAs (PVP/PTX-SAs) had superior cellular internalization capability in MCF7 and MDA-MB-231 breast cancer cells. The optimized PVP/PTX-SAs exhibited less toxicity to human red blood cells indicating a suitable formulation for reducing systemic toxicity. The formation of PVP and PTX self-assemblies was confirmed using fluorescence quenching and transmission electron microscopy which indicated that the PVP/PTX-SAs were spherical in shape with an average size range of 53.81 nm as detected by transmission electron microscopy (TEM). FTIR spectral analysis demonstrates incorporation of polymer and paclitaxel functional groups in PVP/PTX-SAs. Both proliferation (MTS) and clonogenic (colony formation) assays were used to validate superior anticancer activity of PVP/PTX-SAs in breast cancer cells over paclitaxel. Such superior anticancer activity was also demonstrated by downregulation of the expression of pro-survival protein (Bcl-xL), upregulation of apoptosis-associated proteins (Bid, Bax, cleaved caspase 7, and cleaved PARP) and β-tubulin stabilization. These results support the hypothesis that PVP/PTX-SAs improved paclitaxel delivery to cancer cells., Graphical abstract An optimized polyvinylpyrrolidone/paclitaxel self-assembly formulation was established and tested for its improved cellular binding and anti-cancer activity against breast cancer cells.fx1

Purpose Monocytes serve as sanctuary sites for HIV-1 from which virus is difficult to be eliminated. Therefore, an effective viral suppression in monocytes is critical for effective antiretroviral therapy (ART). This study focuses on a new strategy using nanoformulation to optimize the efficacy of ART drugs in HIV-infected monocytes. Methods Poly(lactic- co -glycolic acid) (PLGA)-based elvitegravir nanoparticles (PLGA-EVG) were prepared by nano-precipitation technique. The physicochemical properties of PLGA-EVG were characterized using transmission electron microscopy, dynamic light scattering, and Fourier-transform infrared spectroscopy. Cellular uptake study was performed by fluorescence microscopy and flow cytometry. All in vitro experiments were performed by using HIV-infected monocytic cell lines U1 and HIV-infected primary macrophages. Elvitegravir quantification was performed using LC-MS/MS. HIV viral replication was assessed by using p24 ELISA. Results We developed a PLGA-EVG nanoparticle formulation with particle size of ~ 47 nm from transmission electron microscopy and zeta potential of ~ 6.74 mV from dynamic light scattering. These nanoparticles demonstrated a time- and concentration-dependent uptakes in monocytes. PLGA-EVG formulation showed a ~ 2 times higher intracellular internalization of EVG than control group (EVG alone). PLGA-EVG nanoparticles also demonstrated superior viral suppression over control for a prolonged period of time. Conclusions PLGA-based EVG nanoformulation increased the intracellular uptake of EVG, as well as enhanced viral suppression in HIV-infected macrophages, suggesting its potential for improved HIV treatment in monocytic cells.

INTRODUCTION: Drugs used in HIV treatment; all protease inhibitors, some non-nucleoside reverse transcriptase inhibitors, and pharmacoenhancers ritonavir and cobicistat can inhibit cytochrome P450 (CYP) enzymes. CYP inhibition can cause clinically significant drug-drug interactions (DDI), leading to increased drug exposure and potential toxicity. AREAS COVERED: A complete understanding of pharmacodynamics and CYP-mediated DDI is crucial to prevent adverse side effects and to achieve optimal efficacy. We summarized the pharmacodynamics of all the CYP inhibitors used for HIV treatment, followed by a discussion of drug interactions between these CYP inhibitors and other drugs, and a discussion on the effect of CYP polymorphisms. We also discussed the potential advancements in improving the pharmacodynamics of these CYP inhibitors by using nanotechnology strategy. EXPERT OPINION: The drug-interactions in HIV patients receiving ARV drugs are complicated, especially when patients are on CYP inhibitors-based ART regimens. Therefore, evaluation of CYP-mediated drug interactions is necessary prior to prescribing ARV drugs to HIV subjects. To improve the treatment efficacy and minimize DDI, novel approaches such as nanotechnology may be the potential alternative approach. However, further studies with large cohort need to be conducted to provide strong evidence for the use of nano-formulated ARVs to effectively treat HIV patients.

Background The management of pancreatic cancer (PanCa) is exceptionally difficult due to poor response to available therapeutic modalities. Tubulins play a major role in cell dynamics, thus are important molecular targets for cancer therapy. Among various tubulins, βIII and βIV-tubulin isoforms have been primarily implicated in PanCa progression, metastasis and chemo-resistance. However, specific inhibitors of these isoforms that have potent anti-cancer activity with low toxicity are not readily available. Methods We determined anti-cancer molecular mechanisms and therapeutic efficacy of a novel small molecule inhibitor (VERU-111) using in vitro (MTS, wound healing, Boyden chamber and real-time xCELLigence assays) and in vivo (xenograft studies) models of PanCa. The effects of VERU-111 treatment on the expression of β-tubulin isoforms, apoptosis, cancer markers and microRNAs were determined by Western blot, immunohistochemistry (IHC), confocal microscopy, qRT-PCR and in situ hybridization (ISH) analyses. Results We have identified a novel small molecule inhibitor (VERU-111), which preferentially represses clinically important, βIII and βIV tubulin isoforms via restoring the expression of miR-200c. As a result, VERU-111 efficiently inhibited tumorigenic and metastatic characteristics of PanCa cells. VERU-111 arrested the cell cycle in the G2/M phase and induced apoptosis in PanCa cell lines via modulation of cell cycle regulatory (Cdc2, Cdc25c, and Cyclin B1) and apoptosis - associated (Bax, Bad, Bcl-2, and Bcl-xl) proteins. VERU-111 treatment also inhibited tumor growth (P

Even with an efficient combination of antiretroviral therapy (ART), which significantly decreases viral load in human immunodeficiency virus type 1 (HIV-1)-positive individuals, the occurrence of HIV-1-associated neurocognitive disorders (HAND) still exists. Microglia have been shown to have a significant role in HIV-1 replication in the brain and in subsequent HAND pathogenesis. However, due to the limited ability of ART drugs to cross the blood&ndash, brain barrier (BBB) after systemic administration, in addition to efflux transporter expression on microglia, the efficacy of ART drugs for viral suppression in microglia is suboptimal. Previously, we developed novel poly (lactic-co-glycolic acid) (PLGA)-based elvitegravir nanoparticles (PLGA-EVG NPs), which showed improved BBB penetration in vitro and improved viral suppression in HIV-1-infected primary macrophages, after crossing an in vitro BBB model. Our objective in the current study was to evaluate the efficacy of our PLGA-EVG NPs in an important central nervous system (CNS) HIV-1 reservoir, i.e., microglia. In this study, we evaluated the cyto-compatibility of the PLGA-EVG NPs in microglia, using an XTT (2,3-bis-(2-methoxy-4-nitro-5-sulfophenyl)-2H-tetrazolium-5-carboxanilide) assay and cellular morphology observation. We also studied the endocytosis pathway and the subcellular localization of PLGA NPs in microglia, using various endocytosis inhibitors and subcellular localization markers. We determined the ability of PLGA-EVG NPs to suppress HIV-1 replication in microglia, after crossing an in vitro BBB model. We also studied the drug levels in mouse plasma and brain tissue, using immunodeficient NOD scid gamma (NSG) mice, and performed a pilot study, to evaluate the efficacy of PLGA-EVG NPs on viral suppression in the CNS, using an HIV-1 encephalitic (HIVE) mouse model. From our results, the PLGA-EVG NPs showed ~100% biocompatibility with microglia, as compared to control cells. The internalization of PLGA NPs in microglia occurred through caveolae-/clathrin-mediated endocytosis. PLGA NPs can also escape from endo-lysosomal compartments and deliver the therapeutics to cells efficiently. More importantly, the PLGA-EVG NPs were able to show ~25% more viral suppression in HIV-1-infected human-monocyte-derived microglia-like cells after crossing the in vitro BBB compared to the EVG native drug, without altering BBB integrity. PLGA-EVG NPs also showed a ~two-fold higher level in mouse brain and a trend of decreasing CNS HIV-1 viral load in HIV-1-infected mice. Overall, these results help us to create a safe and efficient drug delivery method to target HIV-1 reservoirs in the CNS, for potential clinical use.

Pancreatic cancer (PanCa) is a lethal disease. Conventional chemotherapies for PanCa offer severe systemic toxicities. Thus, the development of a successful nanomedicine-based therapeutic regimen with augmented therapeutic efficacy is highly sought. Naturally occurring pectin and modified pectin-based drug delivery systems exhibit remarkable self-targeting ability via galactose residues to various cancer cells. Herein, we developed and used an innovative approach of highly stable nanocomplexes based on modified pectin and tannic acid (MPT-NCs). The nanocomplex formation was enabled by strong intermolecular interactions between pectin and tannic acid under very mild conditions. These nanocomplexes were characterized by particle size and morphology (DLS, TEM, and SEM), FT-IR spectroscopy, and zeta potential measurements. Additionally, MPT-NCs were capable of encapsulating anticancer drugs (5-fluorouracil, gemcitabine, and irinotecan) through tannic acid binding. The in vitro bioactivity of these drug MPT-NCs were evaluated in pancreatic cancer adenocarcinoma (PDAC) cell lines (HPAF-II and PANC-1). A dose-dependent internalization of nanocomplexes was evident from microscopy and flow cytometry analysis. Both proliferation and colony formation assays indicated the anticancer potential of pectin drug nanocomplexes against PDAC cells compared to that of free drug treatments. Together, the pectin-based nanocomplexes could be a reliable and efficient drug delivery strategy for cancer therapy.

The therapeutic application of microRNA(s) in the field of cancer has generated significant attention in research. Previous studies have shown that miR-205 negatively regulates prostate cancer cell proliferation, metastasis, and drug resistance. However, the delivery of miR-205 is an unmet clinical need. Thus, the development of a viable nanoparticle platform to deliver miR-205 is highly sought. A novel magnetic nanoparticle (MNP)-based nanoplatform composed of an iron oxide core with poly(ethyleneimine)-poly(ethylene glycol) layer(s) was developed. An optimized nanoplatform composition was confirmed by examining the binding profiles of MNPs with miR-205 using agarose gel and fluorescence methods. The novel formulation was applied to prostate cancer cells for evaluating cellular uptake, miR-205 delivery, and anticancer, antimetastasis, and chemosensitization potentials against docetaxel treatment. The improved uptake and efficacy of formulations were studied with confocal imaging, flow cytometry, proliferation, clonogenicity, Western blot, q-RT-PCR, and chemosensitization assays. Our findings demonstrated that the miR-205 nanoplatform induces significant apoptosis and enhancing chemotherapeutic effects in prostate cancer cells. Overall, these study results provide a strong proof-of-concept for a novel nonviral-based nanoparticle protocol for effective microRNA delivery to prostate cancer cells.

Endoplasmic reticulum (ER) stress is an intriguing target with significant clinical importance in chemotherapy. Interference with ER functions can lead to the accumulation of unfolded proteins, as detected by transmembrane sensors that instigate the unfolded protein response (UPR). Therefore, controlling induced UPR via ER stress with natural compounds could be a novel therapeutic strategy for the management of prostate cancer. Tannic acid (a naturally occurring polyphenol) was used to examine the ER stress mediated UPR pathway in prostate cancer cells. Tannic acid treatment inhibited the growth, clonogenic, invasive, and migratory potential of prostate cancer cells. Tannic acid demonstrated activation of ER stress response (Protein kinase R-like endoplasmic reticulum kinase (PERK) and inositol requiring enzyme 1 (IRE1)) and altered its regulatory proteins (ATF4, Bip, and PDI) expression. Tannic acid treatment affirmed upregulation of apoptosis-associated markers (Bak, Bim, cleaved caspase 3, and cleaved PARP), while downregulation of pro-survival proteins (Bcl-2 and Bcl-xL). Tannic acid exhibited elevated G1 population, due to increase in p18INK4C and p21WAF1/CIP1 expression, while cyclin D1 expression was inhibited. Reduction of MMP2 and MMP9, and reinstated E-cadherin signifies the anti-metastatic potential of this compound. Altogether, these results demonstrate that tannic acid can promote apoptosis via the ER stress mediated UPR pathway, indicating a potential candidate for cancer treatment.

Drug delivery to the eye still is a challenge due to its intricate anatomical structure. Posterior segment delivery is much more challenging due to the acellular nature of the vitreous humor and the longer diffusion distance to the retina. Iontophoresis is a noninvasive technique that facilitates the movement of charged drug molecules into tissues by an electric field. Using iontophoresis, it is possible to achieve therapeutic concentrations faster by modulating the intensity and duration of the applied current. Transscleral iontophoretic delivery is gaining pace and is considered as an alternative for a safe and more effective treatment to retinal disorders. This chapter intends to highlight various aspects of iontophoresis, with a special emphasis on transscleral delivery of drugs and drug-loaded nanocarrier systems for treating disorders in the back of the eye. Finally, a section on toxic effects of iontophoresis to various ocular tissues is included.

This study aims at the development and preliminary evaluation of dexamethasone nanomicelles for treating posterior uveitis. Nanomicelles were formulated using polyoxyl 40 stearate (P40S) and polysorbate 80 (P80), which are approved by the FDA for ocular use.Dexamethasone nanomicelles were prepared and characterized for critical micellar concentration, solubility of dexamethasone, particle size, surface charge, morphology, in vitro drug release, clarity, stability, filtration efficiency, and sterility. Ocular tolerance and the tissue drug distribution of dexamethasone were assessed in rabbits after single and multiple topical administration.Dexamethasone nanomicelles (0.1% w/v) were successfully developed and characterized with an optimized composition of P40S/P80=7/3 by weight. The mean diameter of blank and drug-loaded nanomicelles was 13.3±0.4 and 14.5±0.4 nm, respectively. Transmission electron microscopy images revealed the spherical structure of nanomicelles. Nanomicelles were found to be stable with respect to clarity, size and drug content at 4°C and 25°C for up to 6 months. No irritation or redness was observed in the treated eyes as compared with the untreated control rabbit eyes. Therapeutic concentrations of dexamethasone were observed in the retina and choroid after single and multiple topical application in rabbits.In conclusion, the nanomicelles of P40S and P80 could efficiently solubilize 0.1% dexamethasone in their cores. The results also indicate that mixed nanomicelles could be utilized as a potential delivery system for delivering dexamethasone to treat the back of the eye diseases such as posterior uveitis after topical application.

Chemotherapy is one of the most conventionally used therapeutic interventions for treating various diseases. Chances of acquiring multidrug resistance in response to chemotherapeutic agents are exceedingly common among patients. Drug resistance arises mainly due to overexpression of efflux transporters such as P-glycoprotein and multidrug resistance-associated protein of the ATP-binding cassette superfamily of proteins, which significantly limits intracellular drug accumulation and drug activity. Although many approaches exist to overcome drug resistance, their uses are significantly limited in clinical practice. In this chapter, we demonstrate the superior functions of Pluronic-based technologies to overcome drug resistance. The present chapter highlights various aspects of Pluronic polymers, Pluronic conjugates, Pluronic nanotechnology, as well as their therapeutic implications for effective treatment strategies. We include the role of Pluronic polymers as a pharmaceutic excipient and drug delivery vehicle in this review. In addition, we highlight examples of Pluronic nanosystems that are currently in preclinical development, clinical trials, and clinically translatable formulations. Furthermore, a number of innovative Pluronic nano-designs of advanced therapeutics for future medicinal applications are presented. Collectively, the use of Pluronic-based nanoformulations discussed in this chapter suggests sensitization and prevention of drug resistance. Such an approach not only minimizes the dose required for treatment, but also minimizes the number of treatment cycles, which is useful in a clinical scenario.

Magnetic nanoparticles (MNPs) play a vital role for improved imaging applications. Recently, a number of studies demonstrate MNPs can be applied for targeted delivery, sustained release of therapeutics, and hyperthermia. Based on stable particle size and shape, biocompatibility, and inherent contrast enhancement characteristics, MNPs have been encouraged for pre-clinical studies and human use. As a theranostic platform development, MNPs need to balance both delivery and imaging aspects. Thus, this review provides significant insight and advances in the theranostic role of MNPs through the documentation of unique magnetic nanoparticles used in prostate cancer, their interaction with prostate cancer cells, in vivo fate, targeting, and biodistribution. Specific and custom-made applications of various novel nanoformulations in prostate cancer are discussed.

Background Lung cancer is the leading cause of cancer deaths across globe. Statistically, 70-80% of lung cancer cases fall under non-small cells lung cancer (NSCLC) category. Recently, usage of natural compounds (e.g. EGCG, curcumin, resveratrol, apigenin, etc.,) for cancer therapy has gained lots of attention. However, poor dissolution profiles of these natural compounds lead to unfavorable pharmacological doses and administration. For this purpose, we have candidate, a water-soluble natural polyphenol, Tannic acid (TA, C76H52O46), which, a natural polyphenol that is richly found in plants such as green tea. This molecule exhibits anti-oxidant and anti-proliferative activities, which is being used as an alternative therapeutic option in cancer therapy. However, role of TA in NSCLC was not examined. Therefore, the aim of present study is to assess the molecular effects of TA in NSCLC cells. Methods For this study, two NSCLC cell lines (A549 and H1299) and one normal lung cell line (BEAS-2B) were used. We investigated the anti-proliferative effects of TA on NSCLC cells using 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide) tetrazolium (MTT), and colony formation assays. The anti-invasiveness and anti-migratory potential of TA was evaluated through Matrigel and Boyden chamber studies, respectively. Cell cycle and apoptotic effects of TA was evaluated through flow cytometry and western blot studies. The quantification of secreted VEGF was assessed by ELISA. Also, for affirmation results of gene expression profiles we performed q-PCR studies. Results Cell proliferation study demonstrated a dose dependent anti-cancer effects of TA on NSCLC cells. However, TA had no significant toxicity effects on human bronchial epithelial cells. Similarly, colony formation results exhibited dose dependence reduction in response to escalated dose of TA exposure. Both the invasion and migration results confirm its anti-metastatic role. Extracellular VEGF quantification showed lower VEGF secretion with TA treatments compared to the control cells. The cell cycle analysis evidently showed TA efficiently induced G1 phase cell arrest in NSCLC cells which was further confirmed through western blotting studies of relevant proteins (Cyclin D1, P53, p21, p18, BAX, BCL-2). We also achieved to show the decreased expression of VEGFR2, p-AKT, and cleaved caspase 7 indicating TA induces apoptosis. Conclusion Overall, these results demonstrate illustrate anti-cancer activity of TA by targeting the VEGF related pathways in NSCLC. This approach can lead to an alternative therapeutic strategy for lung cancer, with ideal PK/PD profiles. Citation Format: Elham Hatami, Prashanth K.B. Nagesh, Pallabita Chowdhury, Advit Bhaskar Shetty, Manish K. Tripathi, Subhash Chauhan, Meena Jaggi, Murali Yallapu. Tannic acid: A natural anticancer agent for non-small cell lung cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 1871.

Purpose: Prostate cancer (PrCa) is one of the leading causes of cancer-related deaths among men in the United States. Chemotherapy with docetaxel holds great promise and important therapeutic option for PrCa treatment. However, chemotherapy is often associated with incomplete cell death and leads to persistent senescent phenotype. Such phenotypic cancer cells survive and promotes tumor development, recurrence, and drug resistance. Altogether, targeting these cells can enhance chemotherapy outcomes. In the current study, we established a tannic acid-docetaxel scaffold nanoparticles (TDS NP) platform that allows for facile targeting and inhibition of drug induced senescence in prostate cancer. Methods: TDS NP were prepared by solvent evaporation and extrusion process using a series of tannic acid and docetaxel weight ratios. The scaffold self-assembly formation was determined using a steady-state fluorescence quenching, FT-IR, XRD, DSC, and TGA. The surface and morphological properties of TDS NPs were determined by dynamic light scattering and transmission electron microscopy. The biocompatibility assessment was characterized by hemolysis assay. C4-2 and PC-3 cell lines were used as PrCa model systems for in vitro and in vivo studies. The cellular internalization (in vitro) and accumulation (in vivo) was examined using dye-tagged TDS NPs. The superior in vitro anti-cancer and metastatic potential of TDS NPs were evaluated using proliferation, colony formation, migration, invasion, and immunoblotting assays. The anti-senescence ability of TDS NPs was examined through β-Galactosidase Staining Kit. A PC-3 xenograft mouse model was used to examine its superior therapeutic activity over free docetaxel treatment. Results: A series of physico-chemical analyses confirmed the integrity of TDS NP. An optimized formulation exhibited a spherical shape particle formulation with 124.13 ± 2.16 nm with a negative zeta potential of -14.0 mV. TDS NPs formulation exhibited superior internalization capacity in PrCa cells in a dose- and time-dependent manner. In vitro functional studies confirm superior therapeutic activity of TDS NPs over free docetaxel treatment. Enhanced pro-apoptotic while downregulating anti-apoptotic effects with treatment of TDS NPs in comparison to native drug. A profound inhibition of β-galactosidase activity was noticed with TDS NPs treatment. In vivobiodistribution studies confirm efficient accumulation of TDS NPs in mice. In addition, TDS NPs showed robust antitumor activity against PC-3 xenografts. Such improved activity was correlated with increased inhibition of senescence and induced apoptosis. Conclusion: In summary, we developed a novel tannic acid-docetaxel scaffold nanoparticle formulation that is capable of inhibiting senescence, minimizing drug resistance, and inducing apoptosis in prostate cancer. Citation Format: Prashanth Kumar Bhusetty Nagesh, Pallabita Chowdhury, Elham Hatami, Sonam Kumari, Vivek Kumar Kashyap, Bilal Hafeez, Sheema Khan, Manish Kumar Tripathi, Meena Jaggi, Subhash C. Chauhan, Murali M. Yallapu. Tannic acid-docetaxel scaffold nanoparticles for improved prostate cancer therapy [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 3634.

Background: Although, surgical resection and chemotherapy are the gold standard for treating pancreatic cancer (PanCa), poor patient survival remains the problem. Despite being one of the most common oncogenes in human cancer, to date, no success has been achieved to inhibit KRAS. Targeting KRAS has been shown to synergize anti-cancer activity of gemcitabine in pancreatic cancer. Herein, we have developed a supermagnetic iron oxide (SP) nanoparticles for the sustained delivery of KRAS siRNA to the tumor and simultaneous sensitization of gemcitabine in PanCa. Methods: PanCa cells, AsPC1 and Panc-1 were used in the study. A precipitation approach was employed to develop the SP formulation. These particles were further conjugated with siKRAS (12D; most common KRAS mutation) and investigated for its anticancer efficacy alone and in combination with gemcitabine. Particles were investigated for size, physico-chemical characterization (Dynamic light scattering), hemocompatiblity (hemolysis assay) and the complexation of siKRAS (gel retardation assay). Cellular internalization and uptake of the particles were investigated using FAM labelled siRNA and Prussian blue assay. kRAS silencing was confirmed at both mRNA and protein levels using quantitative reverse-transcription PCR and Western blotting, respectively. Anti-cancer efficacy of SP-siKRAS particles alone or in combination with gemcitabine treatment was determined using in vitro functional assays for cell viability (MTT), migration (Boyden chambers), invasion (Matrigel), colonogenicity and tumor spheroid formation. Results: Our results demonstrate optimal particle size (190.137 nm) and zeta potential (18.73mV) of SP-siKRAS formulation. SP-siKRAS efficiently internalized in PanCa cells and suppressed KRAS G12D expression as well as its downstream targets, YAP and PDL-1. SP-siKRAS improved gemcitabine response as observed through enhanced inhibition of cell proliferation, clonogenicity, migration, and invasion of pancreatic cancer cells. Additionally, SP-siKRAS together with gemcitabine resulted in the activation of death related mechanisms in PanCa cells, such as Bax, bcl-2, PARP cleavage. Interestingly, SP-siKRAS inhibited the secondary tumorsphere formation in combined PanCa and cancer associated fibroblast (CAFs) cells, alone as well as in combination with gemcitabine. At day 14, analysis of secondary tumorspheres, treated with SP-siKRAS revealed diminished levels of KRAS G12D through PCR. This further provides a clinical validation demonstrating potential of SP-siKRAS particles to efficiently silence KRAS expression. SP-siKRAS also exhibited haemocompatibility, suggesting its potential of silencing KRAS without being toxic to the body. Conclusion: Therefore, SP-siKRAS provide a highly efficient and safe platform for silencing KRAS and improving gemcitabine therapy in pancreatic cancer. Citation Format: Nirnoy Dan, Sheema Khan, Saini Setua, Sonam Kumari, Pallabita Chowdhury, Kamalika Samanta, Meena Jaggi, Murali Yallappu, Subhash Chauhan. Nanodelivery platform for targeting mutant-KRAS and improving response to gemcitabine therapy [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 3625.

Organogels have emerged as an alternative carrier for small and macromolecules via transdermal, oral, rectal and ophthalmic routes. Pluronic lecithin organogels (PLO gels) are lecithin-based organogels widely used in compounding pharmacies as a vehicle for enhancing the transdermal permeability of many therapeutic drugs. However, the scientific and systematic evidence in support of how well PLO gels help in transdermal delivery is scanty. Recently, some clinical studies have reported nearly complete lack of bioavailability of certain topically administered drugs from PLO gels. The present review aims at summarizing gels and organogels, with a focus on the use of PLO gels in transdermal drug delivery. A special emphasis is placed on controversies looming over the use of PLO gels as a delivery platform for drugs via transdermal route. This article is open to POST-PUBLICATION REVIEW. Registered readers (see “For Readers”) may comment by clicking on ABSTRACT on the issue’s contents page.

Docetaxel (Dtxl) is currently the most common therapeutic option for prostate cancer (PC). However, adverse side effects and problems associated with chemo-resistance limit its therapeutic outcome in clinical settings. A targeted nanoparticle system to improve its delivery to and activity at the tumor site could be an attractive strategy for PC therapy. Therefore, the objective of this study was to develop and determine the anti-cancer efficacy of a novel docetaxel loaded, prostate specific membrane antigen (PSMA) targeted superparamagnetic iron oxide nanoparticle (SPION) (J591-SPION-Dtxl) formulation for PC therapy. Our results showed the SPION-Dtxl formulation exhibits an optimal particle size and zeta potential, which can efficiently be internalized in PC cells. SPION-Dtxl exhibited potent anti-cancer efficacy via induction of the expression of apoptosis associated proteins, downregulation of anti-apoptotic proteins, and inhibition of chemo-resistance associated protein in PC cell lines. J591-SPION-Dtxl exhibited a profound uptake in C4-2 (PSMA(+)) cells compared to PC-3 (PSMA(-)) cells. A similar targeting potential was observed in ex-vivo studies in C4-2 tumors but not in PC-3 tumors, suggesting its tumor specific targeting. Overall, this study suggests that a PSMA antibody functionalized SPION-Dtxl formulation can be highly useful for targeted PC therapy.

Objectives: Endoplasmic reticulum (ER) is an intricate organelle that is crucial for cellular function and survival. Cellular environments that interfere with ER functioning can lead to the accumulation of unfolded proteins, which are sensed by transmembrane sensors that instigate the unfolded protein response (UPR) to reinstate ER proteostasis. When the UPR is perturbed or not sufficient to deal with the stress conditions, apoptotic cell death is ensued. Often, prostate tumor cells (PrCa) are exposed to intrinsic and external factors that alter protein homeostasis producing endoplasmic reticulum (ER) stress. IRE1, PERK, and ATF6 are key signaling stems of the UPR during ER stress. At molecular level, dimerization of PERK kinase phosphorylates eIF2α, resulting in translation attenuation. This selectively enhances translation of the ATF4 transcription factor. Also, activated IRE1 stimulates the nuclear translocation of XBP1. Both ATF4 and XBP1 induces expression of UPR genes. The UPR stimulates and protects tumor cells against stressful conditions within the tumor microenvironment. Therapeutic intervention on this aspect ER Stress mediated apoptosis is much needed. Here, we examined the mechanistic role of tannic acid (TA) through novel targeting inhibition of UPR via modulation of ER stress in PrCa cell death. Methods: In vitro therapeutic perspective of tannic acid was evaluated using clinically relevant human prostate cancer cell line models (C4-2, PC3 and DU145) through cell proliferation and clonogenic assays. The anti-metastatic potential of TA in PrCa cells was determined using invasive and migration studies. The phase arrest and concomitant apoptosis in PrCa cells were examined using Propidium Iodide based flow cytometric studies. TA induced ER stress-mediated UPR cellular apoptotic activity was confirmed using Western blot, real-time studies, and nuclear distortion studies. Results: Protein profiling results have showed yin approach of inhibition ER stress during TA treatments with the inhibition of CHOP protein. Also, we observed TA treated PrCa cells was able to induce ER stress response proteins, such as PERK and IRE1. Its downstream signaling events include the induction of XBP1, EIF2α expression key mediators by alteration of Bcl2/Bax ratio from shifting survival towards apoptosis. With this line we assessed the expression profile of apoptosis-associated markers such as Bid and Bim (upregulated), and Bcl-2 and Bcl-xL which ratifies the induction of apoptosis during treatments. TA exhibited prominent growth arrest at G1 phase and increase of concentration led to elevated sub G1, annotating the increase of apoptotic cells. The expression of P18 and P21 were increased during TA treatments and cyclin D1 expression was inhibited supporting the G1 phase cell arrest. The expression of MMP2 and MMP9 were reduced, signifying the superior anti-metastatic ability of this compound. Cell proliferation studies showed a dose-dependent inhibitory profile of TA. Distinct results of anti-migratory and anti-metastatic effects of TA was observed during treatments. Conclusion: This study suggests that TA can efficiently induce UPR and promote ER stress signaling in cells that renders apoptosis in PrCa cells by the over expression of CHOP. We believe this novel therapeutic paradigm have clinical significance in advanced PrCa. Citation Format: Elham Hatami, Prashanth Kumar Bhusetty Nagesh, Pallabita Chowdhury, Vivek K. Kashyap, Sheema Khan, Bilal Hafeez, Meena Jaggi, Subhash C. Chauhan, Murali M. Yallapu. Tannic acid induces prostate cancer cell death via unfolded protein response (UPR) and modulation of CHOP [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr LB-400.

Objectives: Paclitaxel (Ptxl) is currently the most common first-line therapeutic option for breast cancer (BrCa). However, adverse side effects and problems associated with chemo-resistance, limits its use in clinical settings. Nanoparticle mediated delivery can improve Ptxl delivery and its activity at the tumor site, and considered to be an attractive strategy for BrCa therapy. Therefore, we aim to generate a novel Ptx medical nanoparticle formulation (PMNPs) using a polyphenol as a carrier, and evaluate its efficacy against BrCa cells. Methods: Physico-chemical characterization of this PMNPs was performed using TEM, DLS, FT-IR, TGA, X-RD, and SAXS methods. Its internalization and cellular availability was examined using HPLC method. Cell proliferation and colony formation assays were utilized to evaluate therapeutic efficacy of this unique nanoformulation in clinically relevant cell line models (MCF7 and MDA-MB-231). Additionally, molecular effects of this formulation on apoptosis, anti-apoptosis, and drug resistant associated proteins were evaluated using immunoblotting assays. A tubulin stabilization study was performed using confocal immunofluorescence microscopy analysis. Results: The PMNPs formulation showed optimal particle size and zeta potential, which can be efficiently internalized in BrCa cells. PMNPs exhibited potent anti-cancer efficacy via induction of the expression of apoptosis associated proteins (Bax and Bad, cleaved PARP, and caspase 3) and downregulation of anti-apoptotic proteins (Bcl-2 and Bcl-xL) in BrCa ell lines. Further, the expression of chemoresistance-associated proteins were also decreased with the treatment of PMNPs. Conclusion: PMNPs formulation can efficiently be internalized in BrCa cells and induce enhanced therapeutic potential of Ptxl. Development of a targeted drug delivery system would not only reduce the dose of paclitaxel required to kill cancer cells but also minimize long-term paclitaxel associated systemic toxicity and drug-resistance. Citation Format: Pallabita Chowdhury, Sumeet S. Chauhan, Prashanth K. Nagesh, Elham Hatami, Bilal B. Hafeez, Sheema Khan, Meena Jaggi, Murali M. Yallapu. Novel paclitaxel medical nanoparticle formulation for breast cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 4658.

Objectives: Docetaxel (Dtx) is the primary choice for the treatment of castrate-resistant prostate cancer (PrCa). Besides its therapeutic benefits, many men with castrate-resistant PCa fail to respond to treatment due to Dtx resistance. There are multiple signaling mechanisms that play pivotal role in the development of Dtx resistance. Seminal reports stated that Tau, highly soluble microtubule-associated protein play a crucial role in Dtx resistance development. Disruption Tau signaling affects its chemoresistance. To revert the drug-resistance and target specific pathway, development of novel and strategic therapies are much needed. In this line, we employed magnetic nanoparticle (MNP) loaded Dtx (MNP-Dtx) to revert the chemo resistance of Dtx in PrCa. Methods: Docetaxel loaded magnetic nanoparticle (MNP-Dtx) formulation is composed of an iron oxide core coated with cyclodextrin (for drug loading) and F127 polymer (for particle stability and chemosensitization). Dtx-resistant PrCa cell line models (PC-3R and DU145R) were developed in our laboratory by repeatedly treating cancer cells in a dose escalation manner for over 24 months. Intracellular drug levels of MNP-Dtx in wild type and drug resistant cancer cells was determined by flow cytometry, TEM and HPLC methods. The superior in vitro anti-cancer activity of MNP-Dtx was examined by MTS, proliferation kinetics, colony formation, immunoblotting and qPCR studies in both wild type and drug resistant PrCa cell lines. The interaction between tau and tubulin studies were assessed through immunoprecipitation studies. Results: MNP-Dtx formulation showed optimal particle size and zeta potential which can efficiently be internalized in PrCa/Dtx resistant PrCa cells. MNP-Dtx exhibited enhanced cellular uptake in drug resistant cell lines over wild type cells in a dose and time dependent manner, with 2 to 3 folds increase. Immunoblot and q PCR studies showed the expression of Tau, whole β tubulin, β tubulin III, β tubulin IV and MDR1 were found to be deregulated in both wild type and resistant cells. To delineate the mechanism of chemosensitization by MNP-Dtx on resistant cells, we performed immunoprecipitation studies, found the expression of Tau was significantly reduced during MNP-Dtx treatments w.r.t control and Dtx. HPLC studies have shown higher amounts of Dtx levels were detected in cells after MNP-Dtx treatments, annotating the sustain drug release ability of MNP nanoformulation. Further, these results were confirmed through tubulin polymerization studies. Conclusion: Overall, sustained drug release of Dtx aided in high intracellular levels of drug, thus facilitating high probability of Dtx-tubulin interaction instead of Tau-tubulin interactions resulting in tubulin fibers stabilization. This promotes Dtx induced apoptosis in wild type and Dtx resistant PrCa cells. This novel therapeutic modality might be effective to treat patients with Dtx resistant PrCa. Citation Format: Prashanth K. B. Nagesh, Pallabita Chowdhury, Elham Hatami, Vivek K. Kashyap, Bilal B. Hafeez, Sheema Khan, Subhash C. Chauhan, Meena Jaggi, Murali M. Yallapu. Docetaxel nanoformulation reverts drug resistance in prostate cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 4657.

Objectives: Breast cancer (BC) is second leading cause of cancer-related deaths in the United States. Paclitaxel (Ptx) is an FDA-approved and frequently used chemotherapeutic agent against various cancers, including BC. However its adverse side effects and chemoresistance against it, limits its use in the clinic. Minimizing the toxicity issues of Ptx through nanoparticle technology (such as Ptx bound to human serum albumin nanoformulation, i.e., Abraxane®) is feasible and has displayed encouraging outcomes. With this background, we aim to generate Ptx self-assemblies (PSAs) using various biocompatible polymers and surfactants, and to evaluate its efficacy against BC cells. Methods: PSAs composed of Ptx dispersion or a core formation with a polymer at a weight ratio of 1:50. The extent of Ptx assembly/binding efficiency was determined using a fluorescence quenching study. FT-IR spectral study was employed to confirm the presence of Ptx in PSAs. The optimal polymers for forming PSAs were identified through measurement of particle size, zeta potential and TEM. Another check point of generating a better PSAs was evaluated by its extent of cellular internalization in BC cells and through hemolytic assay. Finally, the finalized PSAs were examined for in vitro activity in BC cells using proliferation, colony formation, and immunoblotting assays. Results: We screened 22 biocompatible polymers for PSA formation, out of which 8 were finalized due to excellent Ptx binding profiles, appropriate particle size ranges (40-300 nm), zeta potentials (-14.0 to -4.0 mV), and superior internalization in BC cells. The optimized PSAs exhibited enhanced anti-cancer capability in MCF-7 and MDA-MB-231 BC cells in proliferation and colony formation assays, compared to free Ptx. This was further affirmed through tubulin stabilization studies. Further, PSA treatment in BC cells demonstrates a distinct induction of the expression of apoptosis-associated proteins and distinct downregulation of anti-apoptotic proteins. Conclusion: Overall this study suggests a simple and feasible Ptxl self-assembly approach for achieving superior anti-cancer activity with Ptx. Citation Format: Prashanth Kumar Bhusetty Nagesh, Pallabita Chowdhury, Sumeeth S. Chauhan, Elham Hatami, Sheema Khan, Bilal Hafeez, Subhash C. Chauhan, Meena Jaggi, Murali M. Yallapu. Nano self-assemblies of paclitaxel for breast cancer treatment [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr LB-070. doi:10.1158/1538-7445.AM2017-LB-070

Introduction Prostate cancer (PrCa) is the most common male malignancy among men in the United States. Recent studies suggest that low expression of miR-205 is seen in PrCa cell lines and tumors in comparison to normal prostatic epithelial cells. A numer of studies have shown that restoration of miR-205 in PrCa cells resulted in supression of cell growth, epithelial-to-mesenchymal transition, and chemosensitization. However, due to the poor pharmacological kinetics and low in vivo stability of miR-205, serious limitations are being experienced at the clinical level. Therefore, we have chosen a novel nanoparticle-based approach to deliver miR-205, for improved therapeutic benefits in PrCa. Methods A novel miR-205 nanoparticle formulation (named miR-MPG) was generated which is composed of an iron oxide core layered with polyethylene imine (PEI), and NHS-PEG-NHS (PEG) polymer. The miR-205 withholding and release characteristic of miR-PEG were examined through fluorescence quenching and agarose gel electrophoresis. Hemocompatibility of this formulation was examined using a hemolysis assay. Cellular uptake of miR-MPG formulation was evaluated using flow cytometry and confocal studies. Further, therapeutic and chemosensitization activity of miR-205 were assessed using cell-culture based assays. Molecular effects associated with the PrCa cells growth inhibiton were evaluated through protein profiling and qRT-PCR analyses. Results and Dissusion miR-MPG formulation exhibited optimal particle size and zeta potential, which are suitable for cancer therapeutics. Agarose gel electrophoresis binding studies suggested 5 µg of nanoparticle formulation is optimum to hold 1 µg of miR-205 mimic. Release of miR-205 from miR-MPG was determined with respect to concentration of anionic molecules and in a time-dependent manner. We observed no hemolysis during miR-MPG interaction with the red blood cells indicating its hemocompatibility. In addition, miR-MPG particles exhibited superior internalization and endosmal escape in PrCa cells. This formulation displayed enhanced sensitization of PrCa cells to docetaxel. Additionally, it induced the expression of apoptotic proteins (Bax, Bim, cleaved PARP, and caspase 3), and downregulated the anti-apoptotic proteins (Bcl-2 and survivin). Moreover, the expression of the chemoresistance-associated proteins (PSMA and MDR1) was profoundly inhibited in cells treated with miR-MPG in the presence of docetaxel. Further dataset of qRT-PCR studies showed induced expression of the miR-205 and affected the expression of its downstream genes. These results suggest that miR-MPG formulation may serve as an ideal delivery vehicle to deliver miR-205. Conclusion Results from this study suggests that successful delivery of miR-205 through miR-MPG nanoparticles can induce sensitization potential for docetaxel treatment. This novel therapeutic modality might be effective for PrCa patients undergoing chemotherapy. Citation Format: Prashanth K. Nagesh, Pallabita Chowdhury, Vijayakumar N. Boya, Vivek K. Kashyap, Sheema Khan, Bilal B. Hafeez, Nadeem Zafar, Stephen W. Behrman, Subhash C. Chauhan, Meena Jaggi, Murali M. Yallapu. miR-205 replenishment in prostate cancer cells: A novel nanoparticle approach [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 1206. doi:10.1158/1538-7445.AM2017-1206

Objectives: Prostate cancer (PrCa) is the second most leading cause of cancer-related death in men in the United States. Chemotherapy (Docetaxel, Dox) is currently the most common first-line therapeutic option. However, adverse side effects and chemo-resistance of docetaxel limit its clinical use. Improving docetaxel targeted delivery and its activity at the tumor site using a targeted nanoparticle system could be an attractive strategy for PrCa therapy. Prostate Specific Membrane Antigen (PSMA) is highly overexpressed in PrCa cells, thus is a highly attractive molecular target for PrCa therapy. In this study, we developed and determined anti-cancer efficacy of a novel docetaxel loaded, PSMA targeted magnetic nanoparticle (PSMA-MNP-Dox) formulation for PrCa therapy. Methods: Docetaxel loaded magnetic nanoparticle (MNP-Dox) formulation is composed of an iron oxide core coated with cyclodextrin (for drug loading) and F127 polymer (for particle stability and chemosensitization). Therapeutic efficacy of this unique nanoparticle formulation was evaluated using clinically relevant cell line models (C4-2, PC-3, and DU-145) through cell proliferation and colony formation assays. Molecular effects of this formulation on apoptosis, anti-apotosis, and drug resistance associated proteins were evaluated using immunoblotting assays. Contrast imaging property of MNP-Dox formulation was examined using Phantom Gel MR imaging model. For active targeting, PSMA antibody conjugation to this formulation was achieved through N-hydroxysuccinimide group containing PEG polymer. Active targeting potential of this formulation was evaluated in PSMA+ (C4-2) and PSMA- (PC-3) cell lines, C4-2 generated tumor xenografts. Results: MNP-Dox formulation showed optimal particle size and zeta potential which can efficiently internalized in PrCa cells. Our formulation showed anti-cancer efficacy in prostate cancer cell lines. Additionally, it induces the expression of apoptosis associated proteins, Bax and Bad, cleaved PARP, and caspase 3, and downregulated the expression of anti-apoptotic proteins, Bcl-2 and Bcl-xL. Moreover, it also inhibited the expression of chemoresistance associated proteins (PSMA and MDR1). Our PSMA antibody targeted MNPs-Dox formulation exhibited a profound uptake pattern in PSMA+ cells (C4-2) compared to PSMA null (PC-3)- cells, suggesting its targeting potential. A similar targeting potential was also observed in ex-vivo studies while using C4-2 tumor xenografts, however, no intense targeting was observed in normal tissues due to lack of PSMA expression. Conclusion: PSMA antibody functionalized MNP-Dox formulation can efficiently target PSMA + PrCa cells and deliver docetaxel into prostate tumors. This targeted drug delivery system could reduce the dose of docetaxel required to kill cancer cells, thus minimizing long-term docetaxel associated systemic toxicity and drug-resistance. Citation Format: Prashanth Kumar Bhusetty Nagesh, Nia Johnson, Vijaya K.N. Boya, Pallabita Chowdhury, Aditya Ganju, Bilal Hafeez, Sheema Khan, Meena Jaggi, Subhash C. Chauhan, Murali M. Yallapu. PSMA antibody functionalized docetaxel-loaded magnetic nanoparticles for prostate cancer therapy. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 1312.

Table S1. List of Human β-Tubulin Isotypes qRT- PCR Primers used in this study. aIn a number of locations Inosine is inserted to break up runs of four or more Gs that might lead to unusually stable secondary structure.

Table S1. List of Human β-Tubulin Isotypes qRT- PCR Primers used in this study. aIn a number of locations Inosine is inserted to break up runs of four or more Gs that might lead to unusually stable secondary structure.