Enhanced Therapeutic Efficacy Against Melanoma through Exosomal Delivery of Hesperidin.

Melanoma, characterized as the most aggressive and metastatic form of skin cancer, currently has limited treatment options, predominantly chemotherapy and radiation therapy. However, the drawbacks associated with parenterally administered chemotherapy underscore the urgent need for alternative compounds to combat melanoma effectively. Hesperidin (HES), a flavonoid present in various citrus fruits, exhibits promising anticancer activity. Nevertheless, the clinical utility of HES is hindered by challenges such as poor water solubility, a short half-life, and low oral bioavailability. In response to these limitations, we introduced a novel approach by formulating HES-loaded exosomes (Exo-HES). Isolation of exosomes was achieved through the ultracentrifugation method, and HES was efficiently loaded using the sonication method. The resulting formulations displayed a desirable particle size (∼106 nm) and exhibited a spherical morphology, as confirmed by scanning electron and atomic force microscopy. In vitro studies conducted on B16F10 cell lines demonstrated higher cytotoxicity of Exo-HES compared to free HES, supported by enhanced cellular uptake validated through coumarin-6-loaded exosomes. This superior cytotoxicity was further evidenced by DNA fragmentation, increased generation of free radicals (ROS), loss of mitochondrial membrane potential, and effective inhibition of colony formation. The antimetastatic properties of Exo-HES were confirmed through wound healing and transwell migration assays. Oral pharmacokinetics studies revealed a remarkable increase of approximately 2.5 times in oral bioavailability and half-life of HES when loaded into exosomes. Subsequent in vivo experiments utilizing a B16F10-induced melanoma model in Swiss mice established that Exo-HES exhibited superior anticancer activity compared to HES after oral administration. Importantly, no biochemical, hematological, or histological toxicities were observed in tumor-bearing mice treated with Exo-HES. These findings suggest that exosomes loaded with HES represent a promising nanocarrier strategy to enhance the therapeutic effectiveness of hesperidin in melanoma treatment.

Bacteriophage entrapped chitosan microgel for the treatment of biofilm-mediated polybacterial infection in burn wounds.

Staphylococcus aureus (S. aureus) and Pseudomonas aeruginosa (P. aeruginosa) bacteria are most commonly present in burn wound infections. Multidrug resistance (MDR) and biofilm formation make it difficult to treat these infections. Bacteriophages (BPs) are proven as an effective therapy against MDR as well as biofilm-associated wound infections. In the present work, a naturally inspired bacteriophage cocktail loaded chitosan microparticles-laden topical gel has been developed for the effective treatment of these infections. Bacteriophages against MDR S. aureus (BPSAФ1) and P. aeruginosa (BPPAФ1) were isolated and loaded separately and in combination into the chitosan microparticles (BPSAФ1-CHMPs, BPPAФ1-CHMPs, and MBP-CHMPs), which were later incorporated into the SEPINEO™ P 600 gel (BPSAФ1-CHMPs-gel, BPPAФ1-CHMPs-gel, and MBP-CHMPs-gel). BPs were characterized for their morphology, lytic activity, burst size, and hemocompatibility, and BPs belongs to Caudoviricetes class. Furthermore, BPSAФ1-CHMPs, BPPAФ1-CHMPs, and MBP-CHMPs had an average particle size of 1.19 ± 0.11, 1.42 ± 0.21, and 2.84 ± 0.28 µm, respectively, and expressed promising in vitro antibiofilm eradication potency. The ultrasound and photoacoustic imaging in infected burn wounds demonstrated improved wound healing reduced inflammation and increased oxygen saturation following treatment with BPs formulations. The obtained results suggested that the incorporation of the BPs in the MP-gel protected the BPs, sustained the BPs release, and improved the antibacterial activity.

A Bacteriophage-Loaded Microparticle Laden Topical Gel for the Treatment of Multidrug-Resistant Biofilm-Mediated Burn Wound Infection.

Klebsiella pneumoniae is regarded as one of the most profound bacteria isolated from the debilitating injuries caused by burn wounds. In addition, the multidrug resistance (MDR) and biofilm formation make treating burn patients with clinically available antibiotics difficult. Bacteriophage therapy has been proven an effective alternative against biofilm-mediated wound infections caused by MDR bacterial strains. In the current study, the bacteriophage (BPKPФ1) against MDR Klebsiella pneumoniae was isolated and loaded into the chitosan microparticles (CHMPs), which was later incorporated into the Sepineo P 600 to convert into a gel (BPKPФ1-CHMP-gel). BPKPФ1 was characterized for lytic profile, morphological class, and burst size, which revealed that the BPKPФ1 belongs to the family Siphoviridae. Moreover, BPKPФ1 exhibited a narrow host range with 128 PFU/host cell of burst size. The BPKPФ1-loaded CHMPs showed an average particle size of  1.96 ± 0.51 µm, zeta potential 32.16 ± 0.41 mV, and entrapment efficiency in the range of 82.44 ± 1.31%. Further, the in vitro antibacterial and antibiofilm effectiveness of BPKPФ1-CHMPs-gel were examined. The in vivo potential of the BPKPФ1-CHMPs-gel was assessed using a rat model with MDR Klebsiella pneumoniae infected burn wound, which exhibited improved wound contraction (89.22 ± 0.48%) in 28 days with reduced inflammation, in comparison with different controls. Data in hand suggest the potential of bacteriophage therapy to be developed as personalized therapy in case of difficult-to-treat bacterial infections.

Impact of the Drug Loading Method on the Drug Distribution and Biological Efficacy of Exosomes.

Exosomes are biological nanovesicles that are intrinsically loaded with thousands of biomacromolecules and are principally responsible for cell-to-cell communication. Inspired by the natural payload, they have been extensively investigated as drug delivery vehicles; however, the drug distribution, whether into or onto exosomes, is still debatable. In the present work, we have tried to investigate it systemically by selecting 5-fluorouracil (5-FU) (hydrophilic) and paclitaxel (PAC) (hydrophobic), drugs with very different physicochemical characteristics, for the loading to the exosomes. Exosomes were obtained from bovine milk, and the drugs were loaded using three different methods: incubation, sonication, and triton x-100. The particle size was found to be approximately 100 nm in all the cases; however, the highest drug loading was found in the sonication method. Fluorescence spectrophotometer, EDX analysis, EDX mapping, XPS, and XRD analysis indicated the possible presence of more drugs over the surface in the case of the incubation method. Drugs loaded by the sonication method had more controlled release than simple incubation and triton x-100. The method of drug loading had an insignificant effect on the cytotoxicity while in line with our previous observation, the combination (PAC and 5-FU) exhibited synergism as evidenced by ROS assay, colony formation assay, and mitochondrial membrane potential assay.

A Bacteriophage Microgel Effectively Treats the Multidrug-Resistant Acinetobacter baumannii Bacterial Infections in Burn Wounds.

Multidrug-resistant (MDR) Acinetobacter baumannii (A. baumannii) is one of the major pathogens present in burn wound infections. Biofilm formation makes it further challenging to treat with clinically available antibiotics. In the current work, we isolated the A. baumannii-specific bacteriophages (BPABΦ1), loaded into the chitosan microparticles followed by dispersion in gel, and evaluated therapeutic efficacy against MDR A. baumannii clinical strains. Isolated BPABΦ1 were found to belong to the Corticoviridae family, with burst size 102.12 ± 2.65 PFUs per infected host cell. The BPABΦ1 loaded chitosan microparticles were evaluated for quality attributes viz. size, PDI, surface morphology, in vitro release, etc. The developed formulation exhibited excellent antibiofilm eradication potential in vitro and effective wound healing after topical application.

Endorsement of TNBC Biomarkers in Precision Therapy by Nanotechnology.

Breast cancer is a heterogeneous disease which accounts globally for approximately 1 million new cases annually, wherein more than 200,000 of these cases turn out to be cases of triple-negative breast cancer (TNBC). TNBC is an aggressive and rare breast cancer subtype that accounts for 10-15% of all breast cancer cases. Chemotherapy remains the only therapy regimen against TNBC. However, the emergence of innate or acquired chemoresistance has hindered the chemotherapy used to treat TNBC. The data obtained from molecular technologies have recognized TNBC with various gene profiling and mutation settings that have helped establish and develop targeted therapies. New therapeutic strategies based on the targeted delivery of therapeutics have relied on the application of biomarkers derived from the molecular profiling of TNBC patients. Several biomarkers have been found that are targets for the precision therapy in TNBC, such as EGFR, VGFR, TP53, interleukins, insulin-like growth factor binding proteins, c-MET, androgen receptor, BRCA1, glucocorticoid, PTEN, ALDH1, etc. This review discusses the various candidate biomarkers identified in the treatment of TNBC along with the evidence supporting their use. It was established that nanoparticles had been considered a multifunctional system for delivering therapeutics to target sites with increased precision. Here, we also discuss the role of biomarkers in nanotechnology translation in TNBC therapy and management.

Recent advancements in nanotechnology-based bacteriophage delivery strategies against bacterial ocular infections.

Antibiotic resistance is growing as a critical challenge in a variety of disease conditions including ocular infections leading to disastrous effects on the human eyes. Staphylococcus aureus (S. aureus) mediated ocular infections are very common affecting different parts of the eye viz. vitreous chamber, conjunctiva, cornea, anterior and posterior chambers, tear duct, and eyelids. Blepharitis, dacryocystitis, conjunctivitis, keratitis, endophthalmitis, and orbital cellulitis are some of the commonly known ocular infections caused by S. aureus. Some of these infections are so fatal that they could cause bilateral blindness like panophthalmitis and orbital cellulitis, which is caused by methicillin-resistant S. aureus (MRSA) and vancomycin-resistance S. aureus (VRSA). The treatment of S. aureus infections with known antibiotics is becoming gradually difficult because of the development of resistance against multiple antibiotics. Apart from the different combinations and formulation strategies, bacteriophage therapy is growing as an effective alternative to treat such infections. Although the superiority of bacteriophage therapy is well established, yet physical factors (high temperatures, acidic pH, UV-rays, and ionic strength) and pharmaceutical barriers (poor stability, low in-vivo retention, controlled and targeted delivery, immune system neutralization, etc.) have the greatest influence on the viability of phage virions (also phage proteins). A variety of Nanotechnology based formulations such as polymeric nanoparticles, liposomes, dendrimers, nanoemulsions, and nanofibres have been recently reported to overcome the above-mentioned obstacles. In this review, we have compiled all these recent reports and discussed bacteriophage-based nanoformulations techniques for the successful treatment of ocular infections caused by multidrug-resistant S. aureus and other bacteria.

Lipid-Based Nanoparticles as a Pivotal Delivery Approach in Triple Negative Breast Cancer (TNBC) Therapy.

Triple-negative breast cancer is considered the most aggressive type of breast cancer among women and the lack of expressed receptors has made treatment options substantially limited. Recently, various types of nanoparticles have emerged as a therapeutic option against TNBC, to elevate the therapeutic efficacy of the existing chemotherapeutics. Among the various nanoparticles, lipid-based nanoparticles (LNPs) viz. liposomes, nanoemulsions, solid lipid nanoparticles, nanostructured lipid nanocarriers, and lipid-polymer hybrid nanoparticles are developed for cancer treatment which is well confirmed and documented. LNPs include various therapeutic advantages as compared to conventional therapy and other nanoparticles, including increased loading capacity, enhanced temporal and thermal stability, decreased therapeutic dose and associated toxicity, and limited drug resistance. In addition to these, LNPs overcome physiological barriers which provide increased accumulation of therapeutics at the target site. Extensive efforts by the scientific community could make some of the liposomal formulations the clinical reality; however, the relatively high cost, problems in scaling up the formulations, and delivery in a more targetable fashion are some of the major issues that need to be addressed. In the present review, we have compiled the state of the art about different types of LNPs with the latest advances reported for the treatment of TNBC in recent years, along with their clinical status and toxicity in detail.

Combination Therapy Comprising Paclitaxel and 5-Fluorouracil by Using Folic Acid Functionalized Bovine Milk Exosomes Improves the Therapeutic Efficacy against Breast Cancer.

Paclitaxel (PAC) has been approved by FDA for clinical use (Taxol®), yet dose-dependent severe toxicity due to the adjuvant Cremophor EL® in combination with ethanol is a major drawback. The drawbacks of the current therapy can be overcome by (i) finding a suitable vehicle that cannot only bypass the above adjuvant but also be used to deliver drugs orally and (ii) combining the PAC with some other chemotherapeutics to have the enhanced therapeutic efficacy. In the current work, we have used folic acid (FA) functionalized bovine milk-derived exosomes for oral delivery of PAC in combination with 5-fluorouracil (5-FU). Exosomes before and after the drug loading were found to have a particle size in the range of 80-100 nm, polydispersity index (PDI ~0.20), zeta potential (~-25 mV), entrapment efficiency (~82%), practical drug loading (~28%) and sustained drug release for 48 h. Significant decreases in IC50 were observed in the case of exosomes loaded drugs which further improved following the FA functionalization. FA functionalized coumarin-6-loaded exosomes showed remarkably higher cellular uptake in comparison with free coumarin-6. Moreover, FA-functionalized drug-loaded exosomes showed a higher apoptotic index with better control over cell migration. Collectively, data suggested the enhanced efficacy of the combination following its loading to the folic acid functionalized exosomes against breast cancer.

Emergence of Nanotechnology as a Powerful Cavalry against Triple-Negative Breast Cancer (TNBC).

Triple-negative breast cancer (TNBC) is considered one of the un-manageable types of breast cancer, involving devoid of estrogen, progesterone, and human epidermal growth factor receptor 2 (HER 2) receptors. Due to their ability of recurrence and metastasis, the management of TNBC remains a mainstay challenge, despite the advancements in cancer therapies. Conventional chemotherapy remains the only treatment regimen against TNBC and suffers several limitations such as low bioavailability, systemic toxicity, less targetability, and multi-drug resistance. Although various targeted therapies have been introduced to manage the hardship of TNBC, they still experience certain limitations associated with the survival benefits. The current research thus aimed at developing and improving the strategies for effective therapy against TNBC. Such strategies involved the emergence of nanoparticles. Nanoparticles are designated as nanocavalries, loaded with various agents (drugs, genes, etc.) to battle the progression and metastasis of TNBC along with overcoming the limitations experienced by conventional chemotherapy and targeted therapy. This article documents the treatment regimens of TNBC along with their efficacy towards different subtypes of TNBC, and the various nanotechnologies employed to increase the therapeutic outcome of FDA-approved drug regimens.

Exosomes as Emerging Drug Delivery and Diagnostic Modality for Breast Cancer: Recent Advances in Isolation and Application.

Breast cancer (BC) is the most common type of malignancy which covers almost one-fourth of all the cancers diagnosed in women. Conventionally, chemo-, hormonal-, immune-, surgery, and radiotherapy are the clinically available therapies for BC. However, toxicity and other related adverse effects are still the major challenges. A variety of nano platforms have been reported to overcome these limitations, among them, exosomes provide a versatile platform not only for the diagnosis but also as a delivery vehicle for drugs. Exosomes are biological nanovesicles made up of a lipidic bilayer and known for cell-to-cell communication. Exosomes have been reported to be present in almost all bodily fluids, viz., blood, milk, urine, saliva, pancreatic juice, bile, peritoneal, and cerebrospinal fluid. Such characteristics of exosomes have attracted immense interest in cancer diagnosis and therapy. They can deliver bioactive moieties such as protein, lipids, hydrophilic as well as hydrophobic drugs, various RNAs to both distant and nearby recipient cells as well as have specific biological markers. By considering the growing interest of the scientific community in this field, we comprehensively compiled the information about the biogenesis of exosomes, various isolation methods, the drug loading techniques, and their diverse applications in breast cancer diagnosis and therapy along with ongoing clinical trials which will assist future scientific endeavors in a more organized direction.

Nanotherapeutics in autophagy: a paradigm shift in cancer treatment.

Autophagy is a catabolic process in which an organism responds to its nutrient or metabolic emergencies. It involves the degradation of cytoplasmic proteins and organelles by forming double-membrane vesicles called "autophagosomes." They sequester cargoes, leading them to degradation in the lysosomes. Although autophagy acts as a protective mechanism for maintaining homeostasis through cellular recycling, it is ostensibly a cause of certain cancers, but a cure for others. In other words, insufficient autophagy, due to genetic or cellular dysfunctions, can lead to tumorigenesis. However, many autophagy modulators are developed for cancer therapy. Diverse nanoparticles have been documented to induce autophagy. Also, the highly stable nanoparticles show blockage to autophagic flux. In this review, we revealed a general mechanism by which autophagy can be induced or blocked via nanoparticles as well as several studies recently performed to prove the stated fact. In addition, we have also elucidated the paradoxical roles of autophagy in cancer and how their differential role at different stages of various cancers can affect its treatment outcomes. And finally, we summarize the breakthroughs in cancer disease treatments by using metallic, polymeric, and liposomal nanoparticles as potent autophagy modulators.

Targeted Delivery of Natural Bioactives and Lipid-nanocargos against Signaling Pathways Involved in Skin Cancer.

At present, skin cancer is considered a widespread malignancy in human beings. Among diverse population types, Caucasian populations are much more prone to this malignancy in comparison to darker skin populations due to the lack of skin pigmentation. Skin cancer is divided into malignant and non-melanoma skin cancer, which is further categorized as basal and squamous cell carcinoma. Exposure to ultraviolet radiation, chemical carcinogen (polycyclic aromatic hydrocarbons, arsenic, tar, etc.), and viruses (herpes virus, human papillomavirus, and human T-cell leukemia virus type-1) are major contributing factors to skin cancer. There are distinct pathways available through which skin cancer develops, such as the JAK-STAT pathway, Akt pathway, MAPKs signaling pathway, Wnt signaling pathway, to name a few. Currently, several targeted treatments are available, such as monoclonal antibodies, which have dramatically changed the line of treatment of this disease but possess major therapeutic limitations. Thus, many phytochemicals have been evaluated either alone or in combination with the existing synthetic drugs to overcome their limitations and have been found to play a promising role in the prevention and treatment. In this review, a complete overview of skin cancer, starting from the signaling pathways involved, newer developed drugs with their targets and limitations, along with the emerging role of natural products alone or in combination as potent anticancer agents and their molecular mechanism involved has been discussed. Apart from this, various nano-cargos have also been mentioned here, which can play a significant role in the management and treatment of different types of skin cancer.

Natural, Synthetic and their Combinatorial Nanocarriers Based Drug Delivery System in the Treatment Paradigm for Wound Healing Via Dermal Targeting.

A wound refers to the epithelial loss, accompanied by loss of muscle fibers collagen, nerves and bone instigated by surgery, trauma, frictions or by heat. Process of wound healing is a compounded activity of recovering the functional integrity of the damaged tissues. This process is mediated by various cytokines and growth factors usually liberated at the wound site. A plethora of herbal and synthetic drugs, as well as photodynamic therapy, is available to facilitate the process of wound healing. Generally, the systems used for the management of wounds tend to act through covering the ruptured site, reduce pain, inflammation, and prevent the invasion and growth of microorganisms. The available systems are, though, enough to meet these requirements, but the involvement of nanotechnology can ameliorate the performance of these protective coverings. In recent years, nano-based formulations have gained immense popularity among researchers for the wound healing process due to the enhanced benefits they offer over the conventional preparations. Hereupon, this review aims to cover the entire roadmap of wound healing, beginning from the molecular factors involved in the process, the various synthetic and herbal agents, and combination therapy available for the treatment and the current nano-based systems available for delivery through the topical route for wound healing.