Local Lung Targeting of Tumor Associated Macrophages Combined with Cytoreductive Therapy Decrease Tumor Burden in a Secondary Lung Cancer Model

Backgrounds. The efficacy of a locally administered small molecule colony stimulating factor 1 receptor inhibitor (CSF-1Ri), PLX3397, alone or in combination with cytoreductive therapy (paclitaxel) in reducing the tumor burden of an in vivo model of secondary lung cancer was investigated in this work. Local administration of immunotherapy to the lungs may enhance lung biodistribution of such therapies and reduce potential unwanted off-target toxicity. In addition, combination of such therapy with low dose standard of care chemotherapy may offer improved anti-tumor effects. Methods. Murine breast cancer cells (4T1, known to be highly metastasized to the lungs) were transduced to express the genes luciferase and tdTomato, and cells were injected to female Balb/c mice before being treated with PLX (intratracheally administered), paclitaxel (intravenously given), or the combination therapy. Both ex vivo bioluminescent imaging and lung weights were used to evaluate tumor burden. Western blot was performed using lung tumors to assess the effect of PLX3397 on its molecular target (phosphorylated CSF-1R). Immunofluorescence and Flowcytometry were utilized to examine the impact of treatment on tumorigenic tumor associated macrophages (M2 TAM). Results. Single-agent treatment partially decreased tumor burden, while combination therapy led to a significant reduction in tumor burden. PLX3397 significantly inhibited the expression of phosphorylated CSF-1R and reduced the number of M2 TAM without affecting the total macrophage population, thereby increasing the anti-tumorigenic (M1)/M2 ratio. Conclusion. Tumor burden reduction upon local administration of PLX3397 to the lungs correlates with the marked inhibition of the molecular target and the decrease in M2 TAM.https://scholarscompass.vcu.edu/gradposters/1065/thumbnail.jp

Dendrimer Conjugation Enhances Tumor Penetration and Cell Kill of Doxorubicin in 3D Coculture Lung Cancer Models

Background: Doxorubicin (DOX) is a potent chemotherapeutic widely used for solid tumors (1). Despite high efficacy in 2D cell culture, DOX efficacy does not translate to in vivo lung cancer models (2). Major side effects such as cardiotoxicity may be alleviated with nano-based drug delivery systems (nanoDDS). However, tumor penetration of DOX and DOX-nanoDDS is largely unknown and is an additional barrier to effective clinical therapy (3). Here we describe a nanoDDS capable of enhancing the penetration of DOX. Methods: DOX was conjugated to generation 4 poly(amido-amine) dendrimers through (GFLG) tumor- liable bond. G4SA-GFLG-DOX was synthesized/characterized. spheroids were formed of (A549) lung adenocarcinoma cells and (3T3) fibroblasts. Spheroids were characterized for ECM components with immunohistochemistry. Confocal microscopy was used to evaluate the penetration, internalization, and colocalization of DOX and G4SA-GFLG-DOX. MTT assay and Caspase 3/7 to assess 2D and 3D cytotoxicity. Flow cytometry to determine cells uptake. Results: DOX conjugation to dendrimer resulted in G4SA-GFLG-DOX with ~5.5 DOX, 10±1 nm hydrodynamic diameter, and a -17±3 mV zeta-potential. Spheroids of (A549:3T3) were ECM- rich, developed ECM containing collagen-I, hyaluronan, laminin, and fibronectin. While DOX and G4SA-GFLG-DOX had similar toxicities in 2D model, G4SA-GFLG-DOX demonstrated a 3.1-fold greater penetration into spheroids compared to DOX and correlated to a greater efficacy as measured by caspase 3/7 activity. Also, flow cytometry showed higher uptake of G4SA- GFLG-DOX in cancer cells compared to fibroblasts. Conclusion: The work demonstrates enhanced penetration of DOX, via dendrimer conjugation, into an ECM- rich 3D lung cancer model. The enhanced penetration of G4SA-GFLG-DOX correlated with greater antitumor efficacy. Acknowledgements: We acknowledge partial financial support from the Center for Pharmaceutical Engineering and Sciences - School of Pharmacy at VCU. This study was supported by VCU Quest for Distinction and NSF (DRM #1508363). Microscopy was performed at the VCU Microscopy Facility, supported, in part, by funding from NIH-NCI Cancer Center Support Grant P30 CA016059. RA would like to acknowledge King Faisal University (KFU) and Saudi Arabian Cultural Mission (SACM) for a scholarship.https://scholarscompass.vcu.edu/gradposters/1091/thumbnail.jp

Overcoming Mechanisms of Acquired and Innate Doxorubicin Resistance Using Nanomedicine and Tumor-Associated Macrophage Immunotherapy

ABSTRACT Overcoming Mechanisms of Acquired and Innate Doxorubicin Resistance Using Nanomedicine and Tumor-Associated Macrophage Immunotherapy By Rashed M. Almuqbil, Pharm.D. Director: Sandro R.P. da Rocha, PhD Cancer is a leading cause of death worldwide. The available treatment for solid tumors may include surgery, radiotherapy, immunotherapy, and chemotherapy or a combination of all those regimens. Chemotherapy is widely used in the treatment of both lung cancers and breast cancer, including metastases, irrespective of type and stage. The poor rates of survival can be attributed to late diagnosis, and due to the development of acquired and innate chemoresistance. Lung tumors are intrinsically resistant to chemotherapy, thus requiring the development of predictive biomarkers to personalize chemotherapeutic agents. In breast cancer patients, continuous exposure to a single chemotherapeutic agent can lead to (acquired resistance) such as multidrug resistance. Here, we formed a realistic in vitro model for the screening of chemotherapy, as the tumor stroma is composed of extracellular matrix, fibroblast, as well as how it is able to grow in a 3D manner. We described the formulation of a 4th generation PAMAM dendrimer and evaluated its efficacy and penetration through a 3D model of single and coculture spheroids, as a strategy to overcome doxorubicin’s resistance. Also, the efficacy of DOX in vivo as a single and in combination with the immunotherapy PLX3397 was evaluated, to modulate the tumor microenvironment in a syngeneic orthotopic Balb/c mice model by repolarizing M2 macrophages toward more M1 like macrophages, a potential strategy to enhance the innate immunity to fight cancer cells and overcome resistance and diminish distant metastasis

Formulation-by-Design of Efinaconazole Spanlastic Nanovesicles for Transungual Delivery Using Statistical Risk Management and Multivariate Analytical Techniques

As regulatory and technical landscapes for pharmaceutical formulation development are rapidly evolving, a risk-management approach using multivariate analysis is highly essential for designing a product with requisite critical quality attributes (CQA). Efinaconazole, a newly approved poorly water-soluble antifungal triazole drug has poor permeability. Spanlastics, new-generation surfactant nanovesicles, being fluidic, help improve the permeability of drugs. Therefore, we optimized efinaconazole spanlastics using the concepts of Formulation-by-Design (FbD) and explored the feasibility of transungual delivery for the management of onychomycosis. Using the Ishikawa fishbone diagram, the risk factors that may have an impact on the CQA of efinaconazole spanlastic vesicles were identified. Application of the Plackett–Burman experimental design facilitated the screening of eight different formulation and process parameters influencing particle size, transmittance, relative deformability, zeta potential, entrapment efficiency, and dissolution efficiency. With the help of Pareto charts, the three most significant factors were identified, viz., vesicle builder (Span), edge activator (Tween), and mixing time. The levels of these three critical variables were optimized by FbD to reduce the particle size and maximize the transparency, relative deformability, encapsulation efficiency, and dissolution efficiency of efinaconazole spanlastic nanovesicles. Bayesian and Lenth’s analysis and mathematical modeling of the experimental data helped to quantify the critical formulation attributes required for getting the formulation with optimum quality features. The optimized efinaconazole-loaded spanlastic vesicles had a particle size of 197 nm, transparency of 91%, relative deformability of 12.5 min, and dissolution efficiency of 81.23%. The spanlastic formulation was incorporated into a gel and explored ex vivo for transungual delivery. This explorative study provides an example of the application of principles of risk management, statistical multivariate analysis, and the FbD approach in developing efinaconazole spanlastic nanovesicles

Tocotrienol in the Treatment of Topical Wounds: Recent Updates

Healing wounds is an important attempt to keep the internal higher organs safe. Complications in topical wound healing may lead to the formation of scars, which can affect the patient’s quality of life. Although several approaches are ongoing in parallel in the exploration of natural compounds via advanced delivery, in this article, an attempt has been made to highlight tocotrienol. Tocotrienol is a natural form of vitamin E and has shown its potential in certain pharmacological activities better than tocopherol. Its antioxidant, anti-inflammatory, cell signal-mediating effects, angiogenic properties, management of scar, and promotion of wound environment with essential factors have shown potential in the management of topical wound healing. Therefore, this review has aimed to focus on recent advances in topical wound healing through the application of tocotrienols. Challenges in delivering tocotrienols to the topical wound due to its large molecular weight and higher logP have also been explored using nanotechnological-based carriers, which has made tocotrienol a potential tool to facilitate the closure of wounds. Exploration of tocotrienol has also been made in human volunteers for biopsy wounds; however, the results are yet to be reported. Overall, based on the current findings in the literature, it could be inferred that tocotrienol would be a viable alternative to the existing wound dressing components for the management of topical wounds

Intranasal Administration of Dolutegravir-Loaded Nanoemulsion-Based In Situ Gel for Enhanced Bioavailability and Direct Brain Targeting

Dolutegravir’s therapeutic effectiveness in the management of neuroAIDS is mainly limited by its failure to cross the blood–brain barrier. However, lipid-based nanovesicles such as nanoemulsions have demonstrated their potential for the brain targeting of various drugs by intranasal delivery. Thus, the purpose of this study was to develop a Dolutegravir-loaded nanoemulsion-based in situ gel and evaluate its prospective for brain targeting by intranasal delivery. Dolutegravir-loaded nanoemulsions were prepared using dill oil, Tween® 80, and Transcutol® P. Optimization of the nanoemulsion particle size and drug release was carried out using a simplex lattice design. Formulations (F1–F7 and B1–B6) were assessed for various pharmaceutical characteristics. Ex vivo permeation and ciliotoxicity studies of selected in situ gels (B1) were conducted using sheep nasal mucosa. Drug targeting to the brain was assessed in vivo in rats following the nasal delivery of B1. The composition of oil, surfactant, and cosurfactant significantly (p p max (~six-fold, p 0−α (~five-fold, p max (1 h) values in the brain, compared to intravenous administration. Meantime, the drug level in the plasma was relatively low, suggesting less systemic exposure to Dolutegravir through intranasal delivery. In summary, the promising data observed here signifies the prospective of B1 to enhance the brain targeting of Dolutegravir by intranasal delivery and it could be used as a feasible and practicable strategy for the management of neuroAIDS

Intranasal Administration of Dolutegravir-Loaded Nanoemulsion-Based In Situ Gel for Enhanced Bioavailability and Direct Brain Targeting

Dolutegravir’s therapeutic effectiveness in the management of neuroAIDS is mainly limited by its failure to cross the blood–brain barrier. However, lipid-based nanovesicles such as nanoemulsions have demonstrated their potential for the brain targeting of various drugs by intranasal delivery. Thus, the purpose of this study was to develop a Dolutegravir-loaded nanoemulsion-based in situ gel and evaluate its prospective for brain targeting by intranasal delivery. Dolutegravir-loaded nanoemulsions were prepared using dill oil, Tween® 80, and Transcutol® P. Optimization of the nanoemulsion particle size and drug release was carried out using a simplex lattice design. Formulations (F1–F7 and B1–B6) were assessed for various pharmaceutical characteristics. Ex vivo permeation and ciliotoxicity studies of selected in situ gels (B1) were conducted using sheep nasal mucosa. Drug targeting to the brain was assessed in vivo in rats following the nasal delivery of B1. The composition of oil, surfactant, and cosurfactant significantly (p < 0.05) influenced the dependent variables (particle size and % of drug release in 8 h). Formulation B1 exhibits pharmaceutical characteristics that are ideal for intranasal delivery. The mucosal steady-state flux noticed with BI was significantly greater (p < 0.005) than for the control gel. A histopathology of nasal mucosa treated with BI showed no signs of toxicity or cellular damage. Intranasal administration of B1 resulted in greater Cmax (~six-fold, p < 0.0001) and AUC0−α (~five-fold, p < 0.0001), and decreased Tmax (1 h) values in the brain, compared to intravenous administration. Meantime, the drug level in the plasma was relatively low, suggesting less systemic exposure to Dolutegravir through intranasal delivery. In summary, the promising data observed here signifies the prospective of B1 to enhance the brain targeting of Dolutegravir by intranasal delivery and it could be used as a feasible and practicable strategy for the management of neuroAIDS

Formulation, Characterization, Anti-Inflammatory and Cytotoxicity Study of Sesamol-Laden Nanosponges

Sesamol (SES) possesses remarkable chemotherapeutic activity, owing to its anti-inflammatory and antioxidant potential. However, the activity of SES is mainly hampered by its poor physicochemical properties and stability issues. Hence, to improve the efficacy of this natural anti-inflammatory and cytotoxic agent, it was loaded into β-cyclodextrin nanosponges (NS) prepared using different molar ratios of polymer and crosslinker (diphenyl carbonate). The particle size of SES-laden NS (SES-NS) was shown to be in the nano range (200 to 500 nm), with a low polydispersity index, an adequate charge (−17 to −26 mV), and a high payload. Field emission scanning electron microscopy, thermogravimetric analysis, and Fourier transform infrared spectroscopy were used to characterize the bioactive-loaded selected batch (SES-NS6). This batch of nanoformulations showed improved solubilization efficacy (701.88 µg/mL) in comparison to bare SES (244.36 µg/mL), polymer (β-CD) (261.43 µg/mL), and other fabricated batches. The drug release data displayed the controlled release behavior of SES from NS. The findings of the egg albumin denaturation assay revealed the enhanced anti-inflammatory potential of SES-NS as compared to bare SES. Further, the cytotoxicity assay showed that SES-NS was more effective against B16F12 melanoma cell lines than the bioactive alone. The findings of this assay demonstrated a reduction in the IC50 values of SES-NS (67.38 μg/mL) in comparison to SES (106 μg/mL). The present investigation demonstrated the in vitro controlled release pattern and the enhanced anti-inflammatory and cytotoxic activity of SES-NS, suggesting its potential as a promising drug delivery carrier for topical delivery

Development of Novel Unfolding Film System of Itopride Hydrochloride Using Box-Behnken Design—A Gastro Retentive Approach

Currently, gastro-retentive dosage forms achieved a remarkable position among the oral drug delivery systems. This is a broadly used technique to hold the drug delivery systems for a long duration in the gastro intestine (GI) region, slow drug delivery, and overcome other challenges related to typical oral delivery such as low bioavailability. The current work aimed to formulate and characterize a new expandable gastro-retentive system through Itopride Hydrochloride (IH)’s unfolding process for controlled release. The IH-loaded unfolding film formulation was optimized using the Box-Behnken design for folding endurance and length of tested layer (LTL). Initially, the formulation was made using several anti-adhesive additives to promote the unfolding mechanism. Citric acid and sodium bicarbonate were selected as anti-adhesives based on these results. The enfolded film in a capsule shell was shown to unroll in the stomach fluids and render drug delivery up to 12 h in acidic conditions. A fabricated system should have dimensions more than the size of the relaxed pyloric sphincter, and as required, >20 mm LTL was identified. This further confirms that the residence period in the stomach is irrelevant to the fed or fasted condition. Based on desirability criteria, the formulation containing 143.83, 0.7982, and 14.6096 Eudragit L100, PEG, and sodium bicarbonate are selected as optimized formulations (O-IH-UF). The optimized formulation was further analyzed for various parameters such as tensile strength, mechanical strength, unfolding nature, degradability, and in vitro release studies. The pharmacokinetic study revealed greater AUC (area under the curve) and long half-life with the designed O-IH-UF formulation, confirming that the unfolding film type can be a favorable drug system for enhancing the bioavailability of low soluble drugs. The results showed that unfolding types of gastro retentive systems could potentiate the drugs with stability issues in an alkaline medium or those with absorption in acidic conditions