The Effects of ROCK Inhibitor on Prevention of Dexamethasone-Induced Glaucoma Phenotype in Human Trabecular Meshwork Cells.

Purpose: This study investigated the effects of dexamethasone (Dex) on human trabecular meshwork (TM) cells, a model of glucocorticoid-induced glaucoma, and evaluated the impact of ripasudil (Rip) as a co-delivery or sequential dosing strategy. Methods: In vitro experiments were conducted to assess the effects of Dex and Rip on TM cells. Confocal microscopy was used to evaluate the impact of Dex and Rip on F-actin staining signals. Contractility of the TM cells upon Dex and Rip treatment mimicking co-delivery and sequential delivery was quantified using collagen gel contraction assay. Transepithelial electrical resistance (TEER) values and fluorescein isothiocyanate (FITC)-dextran permeability were also measured to assess the impact of Dex and Rip on TM cells. Results: Dex and Rip did not exhibit cytotoxicity at the maximum tested concentration (20 µM). Dex-treated TM cells exhibited higher F-actin staining signals compared to controls, which were reduced when co-treated with Rip. Rip inhibited Dex-induced collagen gel contraction activity in both co-delivery and sequential treatments. Dex resulted in increased TEER values as the dose increased, whereas TEER values were maintained when co-treated with Rip. Conclusions: Co-delivery of Rip has the potential to prevent glaucoma symptoms when patients are treated with Dex. This study highlights the importance of identifying strategies to reduce the side effects of prolonged use of glucocorticoids, such as Dex, in the treatment of various diseases. Translational Relevance: This study demonstrates the potential of co-delivering ripasudil with dexamethasone to mitigate glucocorticoid-induced ocular hypertension and a secondary glaucoma that resembles primary open-angle glaucoma, providing insights for the development of novel preventive strategies in clinical care.

Lipopolyplex-Mediated Co-Delivery of Doxorubicin and FAK siRNA to Enhance Therapeutic Efficiency of Treating Colorectal Cancer.

Tumor metastasis is a major concern in cancer therapy. In this context, focal adhesion kinase (FAK) gene overexpression, which mediates cancer cell migration and invasion, has been reported in several human tumors and is considered a potential therapeutic target. However, gene-based treatment has certain limitations, including a lack of stability and low transfection ability. In this study, a biocompatible lipopolyplex was synthesized to overcome the aforementioned limitations. First, polyplexes were prepared using poly(2-Hydroxypropyl methacrylamide-co-methylacrylate-hydrazone-pyridoxal) (P(HPMA-co-MA-hyd-VB6)) copolymers, which bore positive charges at low pH value owing to protonation of pyridoxal groups and facilitated electrostatic interactions with negatively charged FAK siRNA. These polyplexes were then encapsulated into methoxy polyethylene glycol (mPEG)-modified liposomes to form lipopolyplexes. Doxorubicin (DOX) was also loaded into lipopolyplexes for combination therapy with siRNA. Experimental results revealed that lipopolyplexes successfully released DOX at low pH to kill cancer cells and induced siRNA out of endosomes to inhibit the translation of FAK proteins. Furthermore, the efficient accumulation of lipopolyplexes in the tumors led to excellent cancer therapeutic efficacy. Overall, the synthesized lipopolyplex is a suitable nanocarrier for the co-delivery of chemotherapeutic agents and genes to treat cancers.

Application of Nanoparticles: Diagnosis, Therapeutics, and Delivery of Insulin/Anti-Diabetic Drugs to Enhance the Therapeutic Efficacy of Diabetes Mellitus.

Diabetes mellitus (DM) is a chronic metabolic disorder of carbohydrates, lipids, and proteins due to a deficiency of insulin secretion or failure to respond to insulin secreted from pancreatic cells, which leads to high blood glucose levels. DM is one of the top four noncommunicable diseases and causes of death worldwide. Even though great achievements were made in the management and treatment of DM, there are still certain limitations, mainly related to the early diagnosis, and lack of appropriate delivery of insulin and other anti-diabetic agents. Nanotechnology is an emerging field in the area of nanomedicine and NP based anti-diabetic agent delivery is reported to enhance efficacy by increasing bioavailability and target site accumulation. Moreover, theranostic NPs can be used as diagnostic tools for the early detection and prevention of diseases owing to their unique biological, physiochemical, and magnetic properties. NPs have been synthesized from a variety of organic and inorganic materials including polysaccharides, dendrimers, proteins, lipids, DNA, carbon nanotubes, quantum dots, and mesoporous materials within the nanoscale size. This review focuses on the role of NPs, derived from organic and inorganic materials, in the diagnosis and treatment of DM.

Synergistic Action of Immunotherapy and Nanotherapy against Cancer Patients Infected with SARS-CoV-2 and the Use of Artificial Intelligence.

Since 2019, the SARS-CoV-2 pandemic has caused a huge chaos throughout the world and the major threat has been possessed by the immune-compromised individuals involving the cancer patients; their weakened immune response makes them vulnerable and susceptible to the virus. The oncologists as well as their patients are facing many problems for their treatment sessions as they need to postpone their surgery, chemotherapy, or radiotherapy. The approach that could be adopted especially for the cancer patients is the amalgamation of immunotherapy and nanotherapy which can reduce the burden on the healthcare at this peak time of the infection. There is also a need to predict or analyze the data of cancer patients who are at a severe risk of being exposed to an infection in order to reduce the mortality rate. The use of artificial intelligence (AI) could be incorporated where the real time data will be available to the physicians according to the different patient's clinical characteristics and their past treatments. With this data, it will become easier for them to modify or replace the treatment to increase the efficacy against the infection. The combination of an immunotherapy and nanotherapy will be targeted to treat the cancer patients diagnosed with SARS-CoV-2 and the AI will act as icing on the cake to monitor, predict and analyze the data of the patients to improve the treatment regime for the most vulnerable patients.

Transferrin Modified GSH Sensitive Hyaluronic Acid Derivative Micelle to Deliver HSP90 Inhibitors to Enhance the Therapeutic Efficacy of Brain Cancers.

Herein, GSH-sensitive hyaluronic acid-poly(lactic-co-glycolic acid) (HA-SS-PLGA) was synthesized. Surface modification of PLGA with hyaluronic acid produced a highly stable micelle at physiological pH while a micelle was destabilized at a higher GSH level. Fluorescence microscopy results showed that rhodamine-encapsulated micelle was taken up by brain cancer cells, while competitive inhibition was observed in the presence of free HA and free transferrin. In vitro cytotoxicity results revealed that transferrin-targeted nanoformulated AUY922 (TF-NP-AUY922) shows higher cytotoxicity than either free AUY922 or non-targeted AUY922-loaded micelles (NP-AUY922). In comparison to the control groups, free AUY922, TF-NP-AUY922 or NP-AUY922 treatment revealed the upregulation of HSP70, while the expression of HSP90 client proteins was simultaneously depleted. In addition, the treatment group induced caspase-dependent PARP cleavage and the upregulation of p53 expression, which plays a key role in apoptosis of brain cancer cells. In vivo and ex vivo biodistribution studies showed that cypate-loaded micelle was taken up and accumulated in the tumor regions. Furthermore, in vivo therapeutic efficacy studies revealed that the AUY922-loaded micelle significantly suppressed tumor growth in comparison to the free AUY922, or control groups using tumor-bearing NOD-SCID mice. Moreover, biochemical index and histological analysis revealed synthesized micelle does not show any significant cytotoxicity to the selected major organs. Overall, a synthesized micelle is the best carrier for AUY922 to enhance the therapeutic efficiency of brain cancer.

Cancer Immunotherapy and Application of Nanoparticles in Cancers Immunotherapy as the Delivery of Immunotherapeutic Agents and as the Immunomodulators.

In the last few decades, cancer immunotherapy becomes an important tactic for cancer treatment. However, some immunotherapy shows certain limitations including poor therapeutic targeting and unwanted side effects that hinder its use in clinics. Recently, several researchers are exploring an alternative methodology to overcome the above limitations. One of the emerging tracks in this field area is nano-immunotherapy which has gone through rapid progress and revealed considerable potentials to solve limitations related to immunotherapy. Targeted and stimuli-sensitive biocompatible nanoparticles (NPs) can be synthesized to deliver immunotherapeutic agents in their native conformations to the site of interest to enhance their antitumor activity and to enhance the survival rate of cancer patients. In this review, we have discussed cancer immunotherapy and the application of NPs in cancer immunotherapy, as a carrier of immunotherapeutic agents and as a direct immunomodulator.

Combination Delivery of Alpha-Tocopheryl Succinate and Curcumin Using a GSH-Sensitive Micelle (PAH-SS-PLGA) to Treat Pancreatic Cancer.

Pancreatic cancer is one of the highest causes of mortality throughout the world; thus, it requires an effective treatment strategy. Some chemotherapeutic agents used in the clinics or under clinical trials are hydrophobic and have poor aqueous solubility; consequently, they also have minimal systemic bioavailability. Nanoparticle-based drug delivery tactics have the potential for overcoming these limitations and enhancing their therapeutic efficacy. Herein, a glutathione (GSH)-sensitive micelle (PAH-SS-PLGA) was synthesized for the combined delivery of alpha-tocopheryl succinate (TOS) and curcumin to improve its therapeutic efficacy. The chemical structures of PAH-SS-PLGA were analyzed using Proton Nuclear Magnetic Resonance (1H-NMR) and Fourier Transform Infrared (FTIR) spectroscopy, whereas the particle size, zeta potential, and surface morphology were observed using dynamic light scattering (DLS) and transmission electron microscopy (TEM). In vitro drug release results revealed that more TOS and curcumin were released in the presence of GSH (5 mM) than the physiological pH value. Fluorescence microscopy images revealed that nanoformulated curcumin/rhodamine was uptaken by PAN02 pancreatic cancer cells. In vitro cytotoxicity assays showed higher cytotoxicity for nanoformulated TOS and/or curcumin than free TOS and/or curcumin. In addition, higher cytotoxicity was observed for combination drugs than free drugs alone. Most interestingly, at all tested concentrations of nanoformulated drugs (PAH-SS-PLGA, TOS, and curcumin), the calculated combination index (CI) value was less than one, which shows that TOS and curcumin have a synergistic effect on cellular proliferation inhibition. Overall, synthesized co-polymers are the best carriers for combination drugs, TOS, and curcumin, because they enhance the therapeutic efficacy and improve pancreatic cancer treatments.

pH- and GSH-Sensitive Hyaluronic Acid-MP Conjugate Micelles for Intracellular Delivery of Doxorubicin to Colon Cancer Cells and Cancer Stem Cells.

A dual-sensitive polymeric drug conjugate (HA-SS-MP) was synthesized by conjugating hydrophobic 6-mercaptopurine (MP) to thiolated hyaluronic acid (HA) as the carrier and ligand to deliver doxorubicin (Dox) to parental colon cancer and colon cancer stem cells. Because of the amphiphilic nature of HA-SS-MP, it was self-assembled in the aqueous media, and Dox was physically encapsulated in the core of the micelles. The particle size and the zeta potential of the micelle were analyzed by dynamic light scattering (DLS), and the morphology of the micelle was investigated using transmission electron microscopy (TEM). Drug release study results revealed more drug release at pH 5.0 in the presence of GSH than that at the physiological pH value. The cytotoxicity of free Dox was slightly greater than that of Dox-loaded HA-SS-MP micelles. In vitro cytotoxicity of HA-SS-MP and Dox-loaded HA-SS-MP micelles was greater for cancer stem cells (HCT116-CSCs) than for parental HCT116 colon cancer cells and L929 normal fibroblast cells. The MTT and flow cytometry results confirmed that free HA competitively inhibited Dox-loaded HA-SS-MP uptake. Similarly, flow cytometry results revealed anti-CD44 antibody competitively inhibited cellular uptake of Rhodamine B isothiocyanate conjugated micelles, which confirms that the synthesized micelle is uptaken via CD44 receptor. Cell cycle analysis revealed that free drugs and Dox-loaded HA-SS-MP arrested parental HCT116 colon cancer cells at the S phase, while cell arrest was observed at the G0G1 phase in HCT116-CSCs. In addition, ex vivo biodistribution study showed that Dox-loaded HA-SS-MP micelles were accumulated more in the tumor region than in any other organ. Furthermore, the in vivo results revealed that Dox-loaded HA-SS-MP micelles exhibited more therapeutic efficacy than the free drugs in inhibiting tumor growth in BALB/C nude mice. Overall, the results suggested that the synthesized micelles could be promising as a stimuli carrier and ligand for delivering Dox to colon cancer cells and also to eradicate colon cancer stem cells.

Synthesis and Characterization of Dual-Sensitive Fluorescent Nanogels for Enhancing Drug Delivery and Tracking Intracellular Drug Delivery.

Here, dual-sensitive fluorescent branched alginate-polyethyleneimine copolymer (bAPSC) nanogels were synthesized from thiolated alginate and stearoyl-derivatized branched polyethyleneimine. The formation of bAPSC conjugates was confirmed through proton nuclear magnetic resonance and Fourier transform infrared spectroscopy, whereas dynamic light scattering was used to measure the particle size and ζ potential of the nanogels. The fluorescent properties of the nanogels were confirmed through fluorescent spectroscopy and microscopy. In addition to the excitation-dependent fluorescence behavior, the fluorescence emission intensity of bAPSC was altered by both pH and γ-irradiation. This intensity was higher at a lower pH than at a higher pH, and it slightly decreased after γ-irradiation. The drug loading and encapsulation efficiency of bAPSC were 25.9% and 11.2%, respectively. An in vitro drug release study revealed that the synthesized nanogels release their doxorubicin (Dox) contents in a time-dependent manner, and the drug release was higher after 96 h of incubation. Approximately 43.74% and 88.36% of Dox was released after 96 h of incubation at pH 5.5 in the absence and presence of glutathione (GSH), respectively. However, relatively lower drug release, approximately 21.6% and 16%, was observed in the presence and absence of GSH at pH 7.4, respectively. Fluorescence microscopy confirmed that Dox-loaded bAPSC nanogels were internalized by HeLa cells, and drug distribution was easily tracked using fluorescent materials without additional probing agents. Moreover, cellular cytotoxicity and hemolysis results revealed less cytotoxicity and hemocompatibility of the synthesized nanogels, confirming that they are the most favorable alternative drug carriers for drug delivery systems.

Synthesis and characterization of redox-sensitive heparin-ß-sitosterol micelles: Their application as carriers for the pharmaceutical agent, doxorubicin, and investigation of their antimetastatic activities in vitro.

Although there are several clinical attempts to treat tumors at the primary site, only few therapies can inhibit the spread of metastatic cancers. In this study, we synthesized redox-sensitive heparin-ß-sitosterol micelles that show antimetastatic activity. Proton nuclear magnetic resonance and Fourier transform infrared analyses confirmed the formation of bioreducible heparin-ß-sitosterol (bHSC) conjugates, whereas dynamic light scattering was used to measure the particle size and zeta potential. Both 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide and flow cytometry assays confirmed the low toxicity of the synthesized micelles. Doxorubicin (Dox) was encapsulated via the dialysis method, and its loading and encapsulation efficiencies were 16.49±1.2% and 58.47±1.87%, respectively. An in vitro release study showed that approximately 89% and 52% of Dox were released after 48h in the presence and absence of reduced glutathione, respectively. The hemocompatibility and antimetastatic effects of bHSC were evaluated using the hemolysis and scratch assays, respectively. F-Actin fluorescence microscopy showed that heparin- and bHSC-treated HeLa cells had poorly oriented stress fibers. In summary, the synthesized bHSC micelles are good candidates as drug delivery systems owing to their low toxicity, excellent hemocompatibility, and antimetastatic effects.

Encapsulation of Gadolinium Oxide Nanoparticle (Gd2O3) Contrasting Agents in PAMAM Dendrimer Templates for Enhanced Magnetic Resonance Imaging in Vivo.

There has been growing interest in the research of nanomaterials for biomedical applications in recent decades. Herein, a simple approach to synthesize the G4.5-Gd2O3-poly(ethylene glycol) (G4.5-Gd2O3-PEG) nanoparticles (NPs) that demonstrate potential as dual (T1 and T2) contrasting agents in magnetic resonance imaging (MRI) has been reported in this study. Compared to the clinically popular Gd-DTPA contrasting agents, G4.5-Gd2O3-PEG NPs exhibited a longer longitudinal relaxation time (T1) and better biocompatibility when incubated with macrophage cell line RAW264.7 in vitro. Furthermore, the longitudinal relaxivity (r1) of G4.5-Gd2O3-PEG NPs was 53.9 s-1 mM-1 at 7T, which is equivalent to 4.8 times greater than to the Gd-DTPA contrasting agents. An in vivo T1-weighted MRI results revealed that G4.5-Gd2O3-PEG NPs significantly enhanced signals in the intestines, kidney, liver, bladder, and spleen. In addition, the T2-weighted MRI results revealed darker contrast in the kidney, which proves that G4.5-Gd2O3-PEG NPs can be exploited as T1 and T2 contrasting agents. In summary, these findings suggest that the G4.5-Gd2O3-PEG NPs synthesized by an alternative approach can be used as dual MRI contrasting agents.

A pH-sensitive micelle composed of heparin, phospholipids, and histidine as the carrier of photosensitizers: Application to enhance photodynamic therapy of cancer.

In this study, we describe the synthesis of a stable, pH-sensitive micelle composed of heparin, 1, 2-distearoyl-sn-glycerol-3-phosphoethanolamine, and l-histidine (HDH) through 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide hydrochloride (EDC)/N-hydroxysuccinimide (NHS) chemistry. 1H-Nuclear magnetic resonance (NMR) and Fourier transform infrared spectroscopy (FTIR) analyses confirmed the formation of HDH copolymers and dynamic light scattering (DLS) measurements indicated a particle size of 111.57±12.36nm and zeta potential of -59.8±5.2mV for the nanoparticles. The drug-loading and encapsulation efficiency of the micelles were 14.52±1.2% and 65.47±1.87%, respectively. Drug release studies showed approximately 91% zinc phthalocyanine (ZnPc) release from micelles in acidic conditions (pH 5.0) in comparison with 63% in physiological conditions (pH 7.4) after 96h of incubation. Singlet oxygen (1O2) detection revealed that the micelles prevented ZnPc aggregation and enhanced 1O2 generation. Cellular uptake of ZnPc-loaded micelles (ZnPc-HDH) was observed using confocal microscopy. Phototoxicity experiments in HeLa cells showed that ZnPc-loaded micelles had higher toxicity than that shown by the same concentration of free ZnPc. Hence, pH-sensitive HDH micelles are a promising carrier for hydrophobic ZnPc and improving PDT efficacy.

A pH sensitive polymeric micelle for co-delivery of doxorubicin and α-TOS for colon cancer therapy.

Combination therapy through simultaneous delivery of two or more therapeutic agents using nanocarriers has emerged as an advanced tactic for cancer treatment. To ensure that two therapeutic agents can be co-delivered and rapidly release their cargo in tumor cells, a biocompatible pH-sensitive copolymer, methoxy poly(ethylene glycol)-b-poly(hydroxypropyl methacrylamide-g-α-tocopheryl succinate-g-histidine) (abbreviated as PTH), was designed and synthesized. The PTH copolymers spontaneously self-assembled into micellar-type nanoparticles in aqueous solutions and are used for co-delivery of therapeutic agents, doxorubicin (Dox) and α-TOS. During micellization, π-π stacking occurred between Dox/α-TOS and imidazole rings of PTH copolymers inducing a regular and tight arrangement of copolymers and drugs to form rod-like micelles, thus efficiently increasing the drug loading and encapsulation efficiency. The micelles enabled the rapid release of both Dox and α-TOS when the pH decreased from 7.4 to 4.5. The protein adsorption assay revealed that low amounts of IgG and BSA were adsorbed on the micelles. In vivo biodistribution demonstrated that the micelles could largely accumulate in the tumor tissues. Furthermore, drug-loaded micelles treated with HCT116 cancer cells exhibited higher cytotoxicity than normal cells, which confirmed that α-TOS exhibited a synergy effect with Dox towards cancer cells, while no recognizable side effects were observed during the treatment from organ function tests.

Polysaccharide based nanogels in the drug delivery system: Application as the carrier of pharmaceutical agents.

Polysaccharide-based nanoparticles have fascinated attention as a vesicle of different pharmaceutical agents due to their unique multi-functional groups in addition to their physicochemical properties, including biocompatibility and biodegradability. The existence of multi-functional groups on the polysaccharide backbone permits facile chemical or biochemical modification to synthesize polysaccharide based nanoparticles with miscellaneous structures. Polysaccharide-based nanogels have high water content, large surface area for multivalent bioconjugation, tunable size, and interior network for the incorporation of different pharmaceutical agents. These unique properties offer great potential for the utilization of polysaccharide-based nanogels in the drug delivery systems. Hence, this review describes chemistry of certain common polysaccharides, several methodologies used to synthesize polysaccharide nanoparticles and primarily focused on the polysaccharide (or polysaccharide derivative) based nanogels as the carrier of pharmaceutical agents.

IL-6 Antibody and RGD Peptide Conjugated Poly(amidoamine) Dendrimer for Targeted Drug Delivery of HeLa Cells.

In this study, PAMAM dendrimer (G4.5) was conjugated with two targeting moieties, IL-6 antibody and RGD peptide (G4.5-IL6 and G4.5-RGD conjugates). Doxorubicin anticancer drug was physically loaded onto G4.5-IL6 and G4.5-RGD with the encapsulation efficiency of 51.3 and 30.1% respectively. The cellular internalization and uptake efficiency of G4.5-IL6/DOX and G4.5-RGD/DOX complexes was observed and compared by confocal microscopy and flow cytometry using HeLa cells, respectively. The lower IC50 value of G4.5-IL6/DOX in comparison to G4.5-RGD/DOX is indication that higher drug loading and faster drug release rate corresponded with greater cytotoxicity. The cytotoxic effect was further verified by increment in late apoptotic/necrotic cells due to delivery of drug through receptor-mediated endocytosis. On the basis of these results, G4.5-IL6 is a better suited carrier for targeted drug delivery of DOX to cervical cancer cells.

Drug Carrier for Photodynamic Cancer Therapy.

Photodynamic therapy (PDT) is a non-invasive combinatorial therapeutic modality using light, photosensitizer (PS), and oxygen used for the treatment of cancer and other diseases. When PSs in cells are exposed to specific wavelengths of light, they are transformed from the singlet ground state (S0) to an excited singlet state (S1-Sn), followed by intersystem crossing to an excited triplet state (T1). The energy transferred from T1 to biological substrates and molecular oxygen, via type I and II reactions, generates reactive oxygen species, (¹O2, H2O2, O2*, HO*), which causes cellular damage that leads to tumor cell death through necrosis or apoptosis. The solubility, selectivity, and targeting of photosensitizers are important factors that must be considered in PDT. Nano-formulating PSs with organic and inorganic nanoparticles poses as potential strategy to satisfy the requirements of an ideal PDT system. In this review, we summarize several organic and inorganic PS carriers that have been studied to enhance the efficacy of photodynamic therapy against cancer.