A Comprehensive Review of mRNA Vaccines.

mRNA vaccines have been demonstrated as a powerful alternative to traditional conventional vaccines because of their high potency, safety and efficacy, capacity for rapid clinical development, and potential for rapid, low-cost manufacturing. These vaccines have progressed from being a mere curiosity to emerging as COVID-19 pandemic vaccine front-runners. The advancements in the field of nanotechnology for developing delivery vehicles for mRNA vaccines are highly significant. In this review we have summarized each and every aspect of the mRNA vaccine. The article describes the mRNA structure, its pharmacological function of immunity induction, lipid nanoparticles (LNPs), and the upstream, downstream, and formulation process of mRNA vaccine manufacturing. Additionally, mRNA vaccines in clinical trials are also described. A deep dive into the future perspectives of mRNA vaccines, such as its freeze-drying, delivery systems, and LNPs targeting antigen-presenting cells and dendritic cells, are also summarized.

Octreotide-Targeted Lcn2 siRNA PEGylated Liposomes as a Treatment for Metastatic Breast Cancer.

Lcn2 overexpression in metastatic breast cancer (MBC) can lead to cancer progression by inducing the epithelial-to-mesenchymal transition and enhancing tumor angiogenesis. In this study, we engineered a PEGylated liposomal system encapsulating lipocalin 2 (Lcn2) small interfering RNA (Lcn2 siRNA) for selective targeting MBC cell line MCF-7 and triple-negative breast cancer cell line MDA-MB-231. The PEGylated liposomes were decorated with octreotide (OCT) peptide. OCT is an octapeptide analog of somatostatin growth hormone, having affinity for somatostatin receptors, overexpressed on breast cancer cells. Optimized OCT-targeted Lcn2 siRNA encapsulated PEGylated liposomes (OCT-Lcn2-Lipo) had a mean size of 152.00 nm, PDI, 0.13, zeta potential 4.10 mV and entrapment and loading efficiencies of 69.5% and 7.8%, respectively. In vitro uptake and intracellular distribution of OCT-Lcn2-Lipo in MCF-7 and MDA-MB-231 and MCF-12A cells demonstrated higher uptake for the OCT-targeted liposomes at 6 h by flow cytometry and confocal microscopy. OCT-Lcn2-lipo could achieve approximately 55-60% silencing of Lcn2 mRNA in MCF-7 and MDA-MB-231 cells. OCT-Lcn2-Lipo also demonstrated in vitro anti-angiogenic effects in MCF-7 and MDA-MB-231 cells by reducing VEGF-A and reducing the endothelial cells (HUVEC) migration levels. This approach may be useful in inhibiting angiogenesis in MBC.

Drug Resistance in Metastatic Breast Cancer: Tumor Targeted Nanomedicine to the Rescue.

Breast cancer, specifically metastatic breast, is a leading cause of morbidity and mortality in women. This is mainly due to relapse and reoccurrence of tumor. The primary reason for cancer relapse is the development of multidrug resistance (MDR) hampering the treatment and prognosis. MDR can occur due to a multitude of molecular events, including increased expression of efflux transporters such as P-gp, BCRP, or MRP1; epithelial to mesenchymal transition; and resistance development in breast cancer stem cells. Excessive dose dumping in chemotherapy can cause intrinsic anti-cancer MDR to appear prior to chemotherapy and after the treatment. Hence, novel targeted nanomedicines encapsulating chemotherapeutics and gene therapy products may assist to overcome cancer drug resistance. Targeted nanomedicines offer innovative strategies to overcome the limitations of conventional chemotherapy while permitting enhanced selectivity to cancer cells. Targeted nanotheranostics permit targeted drug release, precise breast cancer diagnosis, and importantly, the ability to overcome MDR. The article discusses various nanomedicines designed to selectively target breast cancer, triple negative breast cancer, and breast cancer stem cells. In addition, the review discusses recent approaches, including combination nanoparticles (NPs), theranostic NPs, and stimuli sensitive or "smart" NPs. Recent innovations in microRNA NPs and personalized medicine NPs are also discussed. Future perspective research for complex targeted and multi-stage responsive nanomedicines for metastatic breast cancer is discussed.

Hyaluronic Acid-Targeted Stimuli-Sensitive Nanomicelles Co-Encapsulating Paclitaxel and Ritonavir to Overcome Multi-Drug Resistance in Metastatic Breast Cancer and Triple-Negative Breast Cancer Cells.

Active targeting and overcoming multi-drug resistance (MDR) can be some of the important attributes of targeted therapy for metastatic breast cancer (MBC) and triple-negative breast cancer (TNBC) treatment. In this study, we constructed a hyaluronic acid (HA)-decorated mixed nanomicelles-encapsulating chemotherapeutic agent paclitaxel (PTX) and P-glycoprotein inhibitor ritonavir (RTV). HA was conjugated to poly (lactide) co-(glycolide) (PLGA) polymer by disulfide bonds (HA-ss-PLGA). HA is a natural ligand for CD44 receptors overexpressed in breast cancer cells. Disulfide bonds undergo rapid reduction in the presence of glutathione, present in breast cancer cells. The addition of RTV can inhibit the P-gp and CYP3A4-mediated metabolism of PTX, thus aiding in reversing MDR and sensitizing the cells toward PTX. An in vitro uptake and cytotoxicity study in MBC MCF-7 and TNBC MDA-MB-231 cell lines demonstrated the effective uptake of the nanomicelles and drug PTX compared to non-neoplastic breast epithelium MCF-12A cells. Interestingly, in vitro potency determination showed a reduction in mitochondrial membrane potential and reactive oxygen species in breast cancer cell lines, indicating effective apoptosis of cancer cells. Thus, stimuli-sensitive nanomicelles along with HA targeting and RTV addition can effectively serve as a chemotherapeutic drug delivery agent for MBC and TNBC.

Self-Assembling Tacrolimus Nanomicelles for Retinal Drug Delivery.

Neovascular age-related macular degeneration (AMD) is characterized by an increase in reactive oxygen species (ROS) and pro-inflammatory cytokines in the retinal pigment epithelium cells. The primary purpose of this study was the development of a clear, tacrolimus nanomicellar formulation (TAC-NMF) for AMD. The optimized formulation had a mean diameter of 15.41 nm, a zeta potential of 0.5 mV, and an entrapment efficiency of 97.13%. In-vitro cytotoxicity studies revealed the dose-dependent cytotoxicity of TAC-NMF on various ocular cell lines, such as human retinal pigment epithelium (D407), monkey retinal choroidal endothelial (RF/6A) cells, and human corneal epithelium (CCL 20.2) cells. Cellular uptake and in-vitro distribution studies using flow cytometry and confocal microscopy, respectively, indicated an elevated uptake of TAC-NMF in a time-dependent manner. Biocompatibility assay using macrophage RAW 264.7 cell line resulted in low production of inflammatory cytokines such as IL-6, IL-1ß and TNF-α after treatment with TAC-NMF. There was a decrease in ROS in D407 cells pre-treated with sodium iodate (ROS inducing agent) after treating with TAC-NMF and tacrolimus drug. Similarly, there was a reduction in the pro-inflammatory cytokines and VEGF-A in D407 cells pretreated with sodium iodate. This indicates that TAC-NMF could lower pro-inflammatory cytokines and ROS commonly seen in AMD.

Preparation and characterization of lutein loaded folate conjugated polymeric nanoparticles.

AIM: To prepare and characterise lutein-loaded polylactide-co-glycolide-polyethylene glycol-folate (PLGA-PEG-FOLATE) nanoparticles and evaluate enhanced uptake in SK-N-BE(2) cells. METHODS: Nanoparticles were prepared using O/W emulsion solvent evaporation and characterised using DLS, SEM, DSC, FTIR and in-vitro release. Lutein-uptake in SK-N-BE(2) cells was determined using flow-cytometry, confocal-microscopy and HPLC. Control was lutein PLGA nanoparticles. RESULTS: The size of lutein-loaded PLGA and PLGA-PEG-FOLATE nanoparticles were 189.6 ± 18.79 nm and 188.0 ± 4.06 nm, respectively. Lutein entrapment was ∼61%(w/w) and ∼73%(w/w) for PLGA and PLGA-PEG-FOLATE nanoparticles, respectively. DSC and FTIR confirmed encapsulation of lutein into nanoparticles. Cellular uptake studies showed ∼1.6 and ∼2-fold enhanced uptake of lutein from PLGA-PEG-FOLATE nanoparticles compared to PLGA nanoparticles and lutein, respectively. Cumulative release of lutein was higher in PLGA nanoparticles (100% (w/w) within 24 h) compared to PLGA-PEG-FOLATE nanoparticles (∼80% (w/w) in 48 h). CONCLUSION: Lutein-loaded PLGA-PEG-FOLATE nanoparticles could be a potential treatment for hypoxic ischaemic encephalopathy.

Lutein-Loaded, Biotin-Decorated Polymeric Nanoparticles Enhance Lutein Uptake in Retinal Cells.

Age related macular degeneration (AMD) is one of the leading causes of visual loss and is responsible for approximately 9% of global blindness. It is a progressive eye disorder seen in elderly people (>65 years) mainly affecting the macula. Lutein, a carotenoid, is an antioxidant, and has shown neuroprotective properties in the retina. However, lutein has poor bioavailability owing to poor aqueous solubility. Drug delivery to the posterior segment of the eye is challenging due to the blood-retina barrier. Retinal pigment epithelium (RPE) expresses the sodium-dependent multivitamin transporter (SMVT) transport system which selectively uptakes biotin by active transport. In this study, we aimed to enhance lutein uptake into retinal cells using PLGA-PEG-biotin nanoparticles. Lutein loaded polymeric nanoparticles were prepared using O/W solvent-evaporation method. Particle size and zeta potential (ZP) were determined using Malvern Zetasizer. Other characterizations included differential scanning calorimetry, FTIR, and in-vitro release studies. In-vitro uptake and cytotoxicity studies were conducted in ARPE-19 cells using flow cytometry and confocal microscopy. Lutein was successfully encapsulated into PLGA and PLGA-PEG-biotin nanoparticles (<250 nm) with uniform size distribution and high ZP. The entrapment efficiency of lutein was ≈56% and ≈75% for lutein-loaded PLGA and PLGA-PEG-biotin nanoparticles, respectively. FTIR and DSC confirmed encapsulation of lutein into nanoparticles. Cellular uptake studies in ARPE-19 cells confirmed a higher uptake of lutein with PLGA-PEG-biotin nanoparticles compared to PLGA nanoparticles and lutein alone. In vitro cytotoxicity results confirmed that the nanoparticles were safe, effective, and non-toxic. Findings from this study suggest that lutein-loaded PLGA-PEG-biotin nanoparticles can be potentially used for treatment of AMD for higher lutein uptake.

Prodrugs and nanomicelles to overcome ocular barriers for drug penetration.

INTRODUCTION: Ocular barriers hinder drug delivery and reduce drug bioavailability. This article focuses on enhancing drug absorption across the corneal and conjunctival epithelium. Both, transporter targeted prodrug formulations and nanomicellar strategy is proven to enhance the drug permeation of therapeutic agents across various ocular barriers. These strategies can increase aqueous drug solubility and stability of many hydrophobic drugs for topical ophthalmic formulations. AREAS COVERED: The article discusses various ocular barriers, ocular influx, and efflux transporters. It elaborates various prodrug strategies used for enhancing drug absorption. Along with this, the article also describes nanomicellar formulation, its characteristic and advantages, and applications in for anterior and posterior segment drug delivery. EXPERT OPINION: Prodrugs and nanomicellar formulations provide an effective strategy for improving drug absorption and drug bioavailability across various ocular barriers. It will be exciting to see the efficacy of nanomicelles for treating back of the eye disorders after their topical application. This is considered as a holy grail of ocular drug delivery due to the dynamic and static ocular barriers, restricting posterior entry of topically applied drug formulations.

Long-term delivery of protein and peptide therapeutics for cancer therapies.

Introduction: Proteins and peptides are prominent therapeutic agents, which are effective in number of ailments. Long-term delivery of protein and peptide therapeutics requires polymeric encapsulation to protect from degradation and for its sustained release. However, results from encapsulation of protein macromolecules in dynamic delivery systems report unreliable clinical outcome, indicating ease of degradation, low permeability, and serious immune responses. A specifically targeted delivery system as tumor or cancer theranostics may surpass these limitations. Areas covered: This review covers recent advancements in approaches involving conjugated protein nano-formulations as targeting delivery technology for various ailments encompassing mostly cancer treatment options. Progressions in targeted chemotherapeutics, protein nanoparticles, peptide nanoparticles, lipidation, and antibody drug-conjugates are discussed. Expert opinion: Significant expansions have been made in forming new generation of antitumor-recombinant proteins, which proves a milestone of advancements for more potent and explicit cancer therapies. However, transformation of biologics from laboratory to clinical trials is an immense challenge, because of drop in efficiency of drug-loading, poor reproducibility of nanoparticles, inadequate information regarding long-term toxicity and insufficient pharmacokinetics data. Hence, early stage tumor diagnosis with précised drug delivery to tumor site is crucial for protein- and peptide- based therapeutics for cancer.

Ocular Drug Delivery: Present Innovations and Future Challenges.

Ocular drug delivery has always been a challenge for ophthalmologists and drug-delivery scientists due to the presence of various anatomic and physiologic barriers. Inimitable static and dynamic ocular barriers not only exclude the entry of xenobiotics but also discourage the active absorption of therapeutic agents. Designing an ideal delivery scheme should include enhanced drug bioavailability and controlled release of drug at the site of action, which can overcome various ocular barriers. Conventional ophthalmic medications include the use of topical eye drops and intravitreal injections of anti-vascular endothelial growth factor agent for treatment of anterior and posterior segment disorders, respectively. Current inventions for anterior ocular segment disorders such as punctum plugs, ocular implants, drug-eluting contact lenses, and ocular iontophoresis represent state-of-the-art inventions for sustained and controlled drug release. Parallel efforts for ocular drug delivery technologies for back of the eye disorders have resulted in the approval of various intravitreal implants. Novel drug-delivery technologies, including nanoparticles, nanomicelles, dendrimers, microneedles, liposomes, and nanowafers, are increasingly studied for anterior and posterior disorders. To achieve patient compliance for back of the eye disorders, novel approaches for noninvasive delivery of potent therapeutic agents are on the rise. In this review article, we discuss past successes, present inventions, and future challenges in ocular drug-delivery technologies. This expert opinion also discusses the future challenges for ocular drug-delivery systems and the clinical translatable potential of nanotechnology from benchtop to bedside.

Ocular Pharmacokinetics of a Topical Ophthalmic Nanomicellar Solution of Cyclosporine (Cequa®) for Dry Eye Disease.

Cequa®, a unique and first-in-class preservative free cyclosporine-A (CsA) nanomicellar topical formulation was recently approved by US FDA for treatment of dry eye disease or keratoconjuntivitis sicca (KCS). Being highly hydrophobic, CsA is currently available as an oil based emulsion, which has its own shortcomings. Developing an aqueous and clear formulation of CsA is imperative yet a challenging need in the quest for a safe and better drug product. In this regard, a novel, clear, aqueous nanomicellar solution of CsA was developed which has the potential to deliver therapeutic concentrations of CsA with minimal discomfort to patients. Highly promising pre-clinical results of Cequa® (OTX-101), has led to its advancement to the clinical trials. Phase III clinical trials have demonstrated that OTX-101 is highly effective, safe, and has a rapid onset of action in treating KCS. This review presents a comprehensive insight on formulation development, preclinical and clinical pharmacokinetic results of Cequa®. Additionally, the translational development of Cequa® from the laboratory benchtop to patient bedside has been discussed.