No abstract

Targeted delivery of drugs to the brain is challenging due to the restricted permeability across the blood brain barrier (BBB). Gliomas are devastating cancers and their positive treatment outcome using Temozolomide (TMZ) is limited due to its short plasma half-life, systemic toxicity and limited access through the blood-brain barrier (BBB). Nanoparticles made of Lactoferrin (Lf) protein, have been shown to enhance the pharmacological properties of drugs. Here, we report the specific ability of Lf nanoparticles to cross BBB and target over-expressed Lf receptors on glioma for enhanced TMZ delivery. TMZ-loaded Lf nanoparticles (TMZ-LfNPs) were prepared by our previously reported sol-oil method. While the Lf protein in the NP matrix aids in transcytosis across the BBB and preferential tumor cell uptake, the pH responsiveness leads to TMZ release exclusively in the tumor microenvironment. Delivery through LfNPs results in an enhanced and sustained intracellular concentration of TMZ in GL261 cells in vitro along with improving its in vivo pharmacokinetics and brain accumulation. TMZ-LfNPs treatment results in a significant reduction of tumor volume, higher tumor cell apoptosis and improved median survival in glioma bearing mice. These results demonstrate that LfNPs present an efficient TMZ delivery platform for an effective treatment of gliomas.

The physico-chemical properties of nanoparticles, as characterized under idealized laboratory conditions, have been suggested to differ significantly when studied under complex physiological environments. A major reason for this variation has been the adsorption of biomolecules (mainly proteins) on the nanoparticle surface, constituting the so-called "biomolecular corona". The formation of biomolecular corona on the nanoparticle surface has been reported to influence various nanoparticle properties viz. cellular targeting, cellular interaction, in vivo clearance, toxicity, etc. Understanding the interaction of nanoparticles with proteins upon administration in vivo thus becomes important for the development of effective nanotechnology-based platforms for biomedical applications. In this chapter, we describe the formation of protein corona on nanoparticles and the differences arising in its composition due to variations in nanoparticle properties. Also discussed is the influence of protein corona on various nanoparticle activities.

Ovarian cancer (OvCa) is the leading cause of gynecological cancer-related deaths in the United States, with five-year survival rates of 15–20% for stage III cancers and 5% for stage IV cancers. The standard of care for advanced OvCa involves surgical debulking of disseminated disease in the peritoneum followed by chemotherapy. Despite advances in treatment efficacy, the prognosis for advanced stage OvCa patients remains poor and the emergence of chemoresistant disease localized to the peritoneum is the primary cause of death. Therefore, a complementary modality that is agnostic to typical chemo- and radio-resistance mechanisms is urgently needed. Photodynamic therapy (PDT), a photochemistry-based process, is an ideal complement to standard treatments for residual disease. The confinement of the disease in the peritoneal cavity makes it amenable for regionally localized treatment with PDT. PDT involves photochemical generation of cytotoxic reactive molecular species (RMS) by non-toxic photosensitizers (PSs) following exposure to non-harmful visible light, leading to localized cell death. However, due to the complex topology of sensitive organs in the peritoneum, diffuse intra-abdominal PDT induces dose-limiting toxicities due to non-selective accumulation of PSs in both healthy and diseased tissue. In an effort to achieve selective damage to tumorous nodules, targeted PS formulations have shown promise to make PDT a feasible treatment modality in this setting. This targeted strategy involves chemical conjugation of PSs to antibodies, referred to as photoimmunoconjugates (PICs), to target OvCa specific molecular markers leading to enhanced therapeutic outcomes while reducing off-target toxicity. In light of promising results of pilot clinical studies and recent preclinical advances, this review provides the rationale and methodologies for PIC-based PDT, or photo-immunotherapy (PIT), in the context of OvCa management.

The process of moving hydrophobic amino acids into the core of a protein by desolvation is important in protein folding. However, a rapid and forced desolvation can lead to precipitation of proteins. Desolvation of proteins under controlled conditions generates nanoparticles – homogeneous aggregates with a narrow size distribution. The protein nanoparticles, under physiological conditions, undergo surface erosion due to the action of proteases, releasing the entrapped drug/gene. The packing density of protein nanoparticles significantly influences the release kinetics. We have investigated the desolvation process of gelatin, exploring the role of pH and desolvating agent in nanoparticle synthesis. Our results show that the desolvation process, initiated by the addition of acetone, follows distinct pathways for gelatin incubated at different pH values and results in the generation of nanoparticles with varying matrix densities. The nanoparticles synthesized with varying matrix densities show variations in drug loading and protease-dependent extra- and intracellular drug release. These results will be useful in fine-tuning the synthesis of nanoparticles with desirable drug release profiles.

Precision imaging, utilizing molecular targeted agents, is an important tool in cancer diagnostics and guiding therapies. While there are limitations associated with single mode imaging probes, multimodal molecular imaging probes enabling target visualization through complementary imaging technologies provides an attractive alternative. However, there are several challenges associated with designing molecular probes carrying contrast agents for complementary multimodal imaging. Here, we propose a dual function antibody conjugate (DFAC) comprising an FDA approved photosensitizer Benzoporphyrin derivative (BPD) and a naphthalocyanine‐based photoacoustic dye (SiNc(OH)) for multimodal infrared (IR) imaging. While fluorescence imaging, through BPD, provides sensitivity, complementing it with photoacoustic imaging, through SiNc(OH), provides a depth‐resolved spatial resolution much beyond the optical diffusion limits of fluorescence measurements. Through a series of in vitro experiments, we demonstrate the development and utilization of DFACs for multimodal imaging and photodynamic treatment of squamous cell carcinoma (A431) cell line. The proposed DFACs have potential use in precision imaging applications such as guiding tumor resection surgeries and photodynamic treatment of residual microscopic disease thereby minimizing local recurrence. The data demonstrated in this study merits further investigation for its preclinical and clinical translation.

Pancreatic ductal adenocarcinoma (PDAC) is an aggressive disease characterized by increased levels of desmoplasia that contribute to reduced drug delivery and poor treatment outcomes. In PDAC, the stromal content can account for up to 90% of the total tumor volume. The complex interplay between stromal components, including pancreatic cancer-associated fibroblasts (PCAFs), and PDAC cells in the tumor microenvironment has a significant impact on the prognoses and thus needs to be recapitulated in vitro when evaluating various treatment strategies. This study is a systematic evaluation of photodynamic therapy (PDT) in 3D heterotypic coculture models of PDAC with varying ratios of patient-derived PCAFs that simulate heterogeneous PDAC tumors with increasing stromal content. The efficacy of antibody-targeted PDT (photoimmunotherapy; PIT) using cetuximab (a clinically approved anti-EGFR antibody) photoimmunoconjugates (PICs) of a benzoporphyrin derivative (BPD) is contrasted with that of liposomal BPD (Visudyne), which is currently in clinical trials for PDT of PDAC. We demonstrate that both Visudyne-PDT and PIT were effective in heterotypic PDAC 3D spheroids with a low stromal content. However, as the stromal content increases above 50% in the 3D spheroids, the efficacy of Visudyne-PDT is reduced by up to 10-fold, while PIT retains its efficacy. PIT was found to be 10-, 19-, and 14-fold more phototoxic in spheroids with 50, 75, and 90% PCAFs, respectively, as compared to Visudyne-PDT. This marked difference in efficacy is attributed to the ability of PICs to penetrate and distribute homogeneously within spheroids with a higher stromal content and the mechanistically different modes of action of the two formulations. This study thus demonstrates how the stromal content in PDAC spheroids directly impacts their responsiveness to PDT and proposes PIT to be a highly suited treatment option for desmoplastic tumors with particularly high degrees of stromal content.

Keratitis is a major cause of avoidable visual impairment. About 30% of patients with fungal keratitis eventually become permanently blind in the developing world. Proteases, secreted by the pathogen and the host, damage the cornea before the infection is resolved. Treating keratitis is a challenge because both infection and inflammation need to be addressed. An additional challenge is to maintain a therapeutic dose at the corneal surface as blinking and tear film wash away the drugs, administered as eye drops. We have developed a nanoparticle-based drug delivery system that enhances the drug residence time by anchoring to the cornea, down-regulates inflammation and releases the antifungal drug: all in a condition-responsive manner. The expression of Toll-Like Receptors (TLR4) on the corneal epithelial cells increases in response to infection. We have conjugated anti-TLR4 antibodies on the surface of ketoconazole-encapsulated gelatin nanoparticles. The anti-TLR4 antibody not only facilitates binding of nanoparticles to the cornea, enhancing their residence time, but also reduces the levels of inflammatory cytokines. Host and fungal proteases degrade the gelatin nanoparticle, an alternative substrate for proteases, thereby reducing corneal damage and releasing the encapsulated drug, ketoconazole, proportional to the severity of infection. After testing the efficacy of the system with human corneal epithelial cells, we have extended our studies to a rat model of keratitis. The results show a significantly increased corneal retention, suppressed inflammation and resolution of infection in the infected eyes. We believe that this will be an excellent approach to manage keratitis as well as other topical ocular infections.