Youssef W. Naguib

Biodistribution and in Vivo Activities of Tumor-Associated Macrophage-Targeting Nanoparticles Incorporated with Doxorubicin

Tumor-associated macrophages (TAMs) are increasingly considered a viable target for tumor imaging and therapy. Previously, we reported that innovative surface-functionalization of nanoparticles may help target them to TAMs. In this report, using poly(lactic-co-glycolic) acid (PLGA) nanoparticles incorporated with doxorubicin (DOX) (DOX-NPs), we studied the effect of surface-modification of the nanoparticles with mannose and/or acid-sensitive sheddable polyethylene glycol (PEG) on the biodistribution of DOX and the uptake of DOX by TAMs in tumor-bearing mice. We demonstrated that surface-modification of the DOX-NPs with both mannose and acid-sensitive sheddable PEG significantly increased the accumulation of DOX in tumors, enhanced the uptake of the DOX by TAMs, but decreased the distribution of DOX in mononuclear phagocyte system (MPS), such as liver. We also confirmed that the acid-sensitive sheddable PEGylated, mannose-modified DOX-nanoparticles (DOX-AS-M-NPs) targeted TAMs because depletion of TAMs in tumor-bearing mice significantly decreased the accumulation of DOX in tumor tissues. Furthermore, in a B16-F10 tumor-bearing mouse model, we showed that the DOX-AS-M-NPs were significantly more effective than free DOX in controlling tumor growth but had only minimum effect on the macrophage population in mouse liver and spleen. The AS-M-NPs are promising in targeting cytotoxic or macrophage-modulating agents into tumors to improve tumor therapy.

Solid lipid nanoparticle formulations of docetaxel prepared with high melting point triglycerides: in vitro and in vivo evaluation.

Docetaxel (DCX) is a second generation taxane. It is approved by the U.S. Food and Drug Administration for the treatment of various types of cancer, including breast, non-small cell lung, and head and neck cancers. However, side effects, including those related to Tween 80, an excipient in current DCX formulations, can be severe. In the present study, we developed a novel solid lipid nanoparticle (SLN) composition of DCX. Trimyristin was selected from a list of high melting point triglycerides as the core lipid component of the SLNs, based on the rate at which the DCX was released from the SLNs and the stability of the SLNs. The trimyristin-based, PEGylated DCX-incorporated SLNs (DCX-SLNs) showed significantly higher cytotoxicity against various human and murine cancer cells in culture, as compared to DCX solubilized in a Tween 80/ethanol solution. Moreover, in a mouse model with pre-established tumors, the new DCX-SLNs were significantly more effective than DCX solubilized in a Tween 80/ethanol solution in inhibiting tumor growth without toxicity, likely because the DCX-SLNs increased the concentration of DCX in tumor tissues, but decreased the levels of DCX in major organs such as liver, spleen, heart, lung, and kidney. DCX-incorporated SLNs prepared with one or more high-melting point triglycerides may represent an improved DCX formulation.

Tumor-Associated Macrophage-Mediated Targeted Therapy of Triple-Negative Breast Cancer

Triple-negative breast cancer (TNBC) is the most aggressive form of breast cancer. TNBC is often infiltrated with a large number of macrophages, which in turn promote tumor growth and metastasis. In this study, tumor-associated macrophages (TAMs) were exploited as a target to deliver doxorubicin (DOX), a chemotherapeutic agent, to TNBC using nanoparticles surface-functionalized by (i) acid-sensitive sheddable PEGylation and (ii) modifying with mannose (i.e., DOX-AS-M-PLGA-NPs). In mice with orthotopic M-Wnt triple-negative mammary tumors, a single intravenous injection of DOX-AS-M-PLGA-NPs significantly reduced macrophage population in tumors within 2 days, and the density of the macrophages recovered slowly. Repeated injections of DOX-AS-M-PLGA-NPs can help maintain the population of the macrophages at a lower level. In M-Wnt tumor-bearing mice that were pretreated with zoledronic acid to nonselectively deplete macrophages, the TAM-targeting DOX-AS-M-PLGA-NPs were not more effective than the DOX-AS-PLGA-NPs that were not surface-modified with mannose and thus do not target TAMs in controlling tumor growth. However, in M-Wnt tumor-bearing mice that were not pretreated with zoledronic acid, the TAM-targeting DOX-AS-M-PLGA-NPs were significantly more effective than the nontargeting DOX-AS-PLGA-NPs in controlling the tumor growth. The AS-M-PLGA-NPs or other nanoparticles surface-functionalized similarly, when loaded with a chemotherapeutic agent commonly used in adjuvant therapy of TNBC, may be developed into targeted therapy for TNBC.

The effect of microneedles on the skin permeability and antitumor activity of topical 5-fluorouracil

Topical 5-fluorouracil (5-FU) is approved for the treatment of superficial basal cell carcinoma and actinic keratosis. However, 5-FU suffers from poor skin permeation. Microneedles have been successfully applied to improve the skin permeability of small and large molecules, and even nanoparticles, by creating micron-sized pores in the stratum corneum layer of the skin. In this report, the feasibility of using microneedles to increase the skin permeability of 5-FU was tested. Using full thickness mouse skin mounted on Franz diffusion apparatus, it was shown that the flux of 5-FU through the skin was increased by up to 4.5-fold when the skin was pretreated with microneedles (500 μm in length, 50 μm in base diameter). In a mouse model with B16-F10 mouse melanoma cells implanted in the subcutaneous space, the antitumor activity of a commercially available 5-FU topical cream (5%) was significantly enhanced when the cream was applied on a skin area that was pretreated with microneedles, as compared to when the cream was simply applied on a skin area, underneath which the tumor cells were implanted, and without pretreatment of the skin with microneedles. Fluorouracil is not approved for melanoma therapy, but the clinical efficacy of topical 5-FU against tumors such as basal cell carcinoma may be improved by integrating microneedle technology into the therapy.

Acid-Sensitive Sheddable PEGylated PLGA Nanoparticles Increase the Delivery of TNF-α siRNA in Chronic Inflammation Sites

There has been growing interest in utilizing small interfering RNA (siRNA) specific to pro-inflammatory cytokines, such as tumor necrosis factor-α (TNF-α), in chronic inflammation therapy. However, delivery systems that can increase the distribution of the siRNA in chronic inflammation sites after intravenous administration are needed. Herein we report that innovative functionalization of the surface of siRNA-incorporated poly (lactic-co-glycolic) acid (PLGA) nanoparticles significantly increases the delivery of the siRNA in the chronic inflammation sites in a mouse model. The TNF-α siRNA incorporated PLGA nanoparticles were prepared by the standard double emulsion method, but using stearoyl-hydrazone-polyethylene glycol 2000, a unique acid-sensitive surface active agent, as the emulsifying agent, which renders (i) the nanoparticles PEGylated and (ii) the PEGylation sheddable in low pH environment such as that in chronic inflammation sites. In a mouse model of lipopolysaccharide-induced chronic inflammation, the acid-sensitive sheddable PEGylated PLGA nanoparticles showed significantly higher accumulation or distribution in chronic inflammation sites than PLGA nanoparticles prepared with an acid-insensitive emulsifying agent (i.e., stearoyl-amide-polyethylene glycol 2000) and significantly increased the distribution of the TNF-α siRNA incorporated into the nanoparticles in inflamed mouse foot.There has been growing interest in utilizing small interfering RNA (siRNA) specific to pro-inflammatory cytokines, such as tumor necrosis factor-α ( TNF-α), in chronic inflammation therapy. However, delivery systems that can increase the distribution of the siRNA in chronic inflammation sites after intravenous administration are needed. Herein we report that innovative functionalization of the surface of siRNA-incorporated poly (lactic-co-glycolic) acid (PLGA) nanoparticles significantly increases the delivery of the siRNA in the chronic inflammation sites in a mouse model. The TNF-α siRNA incorporated PLGA nanoparticles were prepared by the standard double emulsion method, but using stearoyl-hydrazone-polyethylene glycol 2000, a unique acid-sensitive surface active agent, as the emulsifying agent, which renders (i) the nanoparticles PEGylated and (ii) the PEGylation sheddable in low pH environment such as that in chronic inflammation sites. In a mouse model of lipopolysaccharide-induced chronic inflammation, the acid-sensitive sheddable PEGylated PLGA nanoparticles showed significantly higher accumulation or distribution in chronic inflammation sites than PLGA nanoparticles prepared with an acid-insensitive emulsifying agent (i.e., stearoyl-amide-polyethylene glycol 2000) and significantly increased the distribution of the TNF-α siRNA incorporated into the nanoparticles in inflamed mouse foot.

Applications of bacillus Calmette-Guerin and recombinant bacillus Calmette-Guerin in vaccine development and tumor immunotherapy.

Bacillus Calmette–Guerin (BCG) vaccines are attenuated live strains of Mycobacterium bovis and are among the most widely used vaccines in the world. BCG is proven to be effective in preventing severe infant meningitis and miliary tuberculosis. Intravesical instillation of BCG is also a standard treatment for non-muscle invasive bladder cancer. In the past few decades, recombinant BCG (rBCG) technology had been extensively applied to develop vaccine candidates for a variety of infectious diseases, including bacterial, viral, and parasite infections, and to improve the efficacy of BCG in bladder cancer therapy. This review is intended to show the vast applications of BCG and recombinant BCG (rBCG) in the prevention of infectious diseases and cancer immunotherapy, with a special emphasis on recent approaches and trends on both pre-clinical and clinical levels.

Nanomedicine: The Promise and Challenges in Cancer Chemotherapy

Cancer is a leading cause of death worldwide. Conventional chemotherapy has drawbacks ranging from limited effectiveness, chemoresistance, and treatment-related side-effects and damages to healthy tissues. The idea of drug targeting started to emerge as early as 1906. As outlined by Danhier et al., the challenge ahead is about how to find proper targets, to develop a drug that exploits the targets, and finally to design the proper delivery system (carrier) for the drug [1]. Cancer nanomedicine is the use of nano-sized entities (10–200 nm) to diagnose, prevent, or treat cancer. These entities can be loaded with chemotherapeutics, nucleic acids, radiosensitizers, diagnostic agents and/or probes. This chapter displays an updated review of several nano-carrier platforms, their common features, and applications, followed by a discussion of the major mechanisms by which nano-carriers are targeted to cancer cells. Finally it discusses barriers that hinder the applications of nanomedicine in cancer chemotherapy, and how the rational design of the nanomedicine may overcome these barriers.

In vivo distribution of zoledronic acid in a bisphosphonate-metal complex-based nanoparticle formulation synthesized by a reverse microemulsion method

Bisphosphonates are used to treat bone diseases such as osteoporosis and cancer-induced bone pain and fractures. It is thought that modifying the pharmacokinetics and biodistribution profiles of bisphosphonates (i.e. rapid renal clearance and extensive bone absorption) will not only reduce their side effects, but also expand their clinical applications to extraskeletal tissues. In the present work, using zoledronic acid (Zol) and calcium as model bisphosphonate and metal molecules, respectively, we prepared DOPA (an anionic lipid)-coated spherical Zol-Ca nanocomposites (Zol-Ca@DOPA) and developed Zol-nanoparticle formulations (i.e. Zol-Ca@bi-lipid NPs) based on the nanocomposites. The influence of the inputted weight ratio of Zol-Ca@DOPA to DSPE-PEG2k on the properties (e.g. size, size distribution, loading efficiency, encapsulation efficiency, zeta potential, and polydispersity) of Zol-Ca@bi-lipid NPs was investigated, and a type of Zol-Ca@bi-lipid NPs with size around 25nm was selected for further studies. In a mouse model, the Zol-Ca@bi-lipid NPs significantly reduced the bone distribution of Zol, increased the blood circulating time of Zol, and altered the distribution of Zol in major organs, as compared to free Zol. It is expected that similar nanoparticles prepared with bisphosphonate-metal complexes can be explored to expand the applications to bisphosphonates in extraskeletal tissues.

A method to improve the efficacy of topical eflornithine hydrochloride cream

Abstract Context: Facial hirsutism is a cosmetic concern for women and can lead to significant anxiety and lack of self-esteem. Eflornithine cream is indicated for the treatment of facial hirsutism. However, limited success rate and overall patients satisfaction, even with a long-term and high-frequency application, leave room for improvement. Objective: The objective of this study is to test the effect of microneedle treatment on the in vitro skin permeation and the in vivo efficacy of eflornithine cream in a mouse model. Materials and method: In vitro permeation study of eflornithine was performed using Franz diffusion cell. In vivo efficacy study was performed in a mouse model by monitoring the re-growth of hair in the lower dorsal skin of mice after the eflornithine cream was applied onto an area pretreated with microneedles. The skin and the hair follicles in the treated area were also examined histologically. Results and discussion: The hair growth inhibitory activity of eflornithine was significantly enhanced when the eflornithine cream was applied onto a mouse skin area pretreated with microneedles, most likely because the micropores created by microneedles allowed the permeation of eflornithine into the skin, as confirmed in an in vitro permeation study. Immunohistochemistry data revealed that cell proliferation in the skin and hair follicles was also significantly inhibited when the eflornithine cream was applied onto a skin area pretreated with microneedles. Conclusion: The integration of microneedle treatment into topical eflornithine therapy represents a potentially viable approach to increase eflornithines ability to inhibit hair growth.

Synthesis, Characterization, and In Vitro and In Vivo Evaluations of 4-(N)-Docosahexaenoyl 2′, 2′-Difluorodeoxycytidine with Potent and Broad-Spectrum Antitumor Activity

In this study, a new compound, 4-(N)-docosahexaenoyl 2′, 2′-difluorodeoxycytidine (DHA-dFdC), was synthesized and characterized. Its antitumor activity was evaluated in cell culture and in mouse models of pancreatic cancer. DHA-dFdC is a poorly soluble, pale yellow waxy solid, with a molecular mass of 573.3 Da and a melting point of about 96°C. The activation energy for the degradation of DHA-dFdC in an aqueous Tween 80–based solution is 12.86 kcal/mol, whereas its stability is significantly higher in the presence of vitamin E. NCI-60 DTP Human Tumor Cell Line Screening revealed that DHA-dFdC has potent and broad-spectrum antitumor activity, especially in leukemia, renal, and central nervous system cancer cell lines. In human and murine pancreatic cancer cell lines, the IC50 value of DHA-dFdC was up to 105-fold lower than that of dFdC. The elimination of DHA-dFdC in mouse plasma appeared to follow a biexponential model, with a terminal phase t1/2 of about 58 minutes. DHA-dFdC significantly extended the survival of genetically engineered mice that spontaneously develop pancreatic ductal adenocarcinoma. In nude mice with subcutaneously implanted human Panc-1 pancreatic tumors, the antitumor activity of DHA-dFdC was significantly stronger than the molar equivalent of dFdC alone, DHA alone, or the physical mixture of them (1:1, molar ratio). DHA-dFdC also significantly inhibited the growth of Panc-1 tumors orthotopically implanted in the pancreas of nude mice, whereas the molar equivalent dose of dFdC alone did not show any significant activity. DHA-dFdC is a promising compound for the potential treatment of cancers in organs such as the pancreas.