Injectable in-situ gel depot system for targeted delivery of biologics to the retina.

In current clinical settings, frequent intravitreal (IVT) injections of anti-vascular endothelial growth factors are used due to their short in-vivo half-life and rapid clearance from the back of the eye. The IVT injections are associated with pain, risk of infection, retinal detachment, and financial burden. Biologics molecules can undergo physical, chemical, and enzymatic degradation during formulation development and in the biological environment. Moreover, the complex ocular structures also act as a rate-limiting barrier for these biologics. Thus, delivering stable and clinically relevant biologics concentration to the back of the eye is still a challenge. Compare to other drug delivery platforms, injectable in-situ gelling depot systems (IISGDs) have emerged as an effective system for biologics delivery. In this review, we have discussed various biologics used in ocular therapeutics and their associated challenges. Different routes of delivery and associated tissue barriers are also discussed. Different types of IISGDs developed to date for biologics delivery to the back of the eye were also covered. To conclude, various critical parameters related to the formulation development process and injectable depot systems that need careful consideration and further investigations were highlighted.

Potential of subconjunctival aflibercept in treating choroidal neovascularization.

The study aimed to evaluate the intraocular pharmacokinetics and efficacy of aflibercept after subconjunctival injection in animal models for treating choroidal neovascularization (CNV) associated with Age-Related Macular Degeneration (AMD). New Zealand albino rabbits received aflibercept (2000 µg/50 µl) in one eye, and the other eye was used as control. At 7, 14, 21 and 28 days, the animals were sacrificed to dissect the ocular tissues, and serum was collected at 1hr, 3 h, 1, 7, 14, 21 and 28 days. The concentration of aflibercept in various ocular tissues and serum were measured using the immunoassay technique. The concentration maximum (Cmax) at the Retinal Pigment Epithelium (RPE)-choroid complex and retina in treated eyes was 261.55 and 33.83 ng/gm, respectively. The area under the curve (AUC0-last) for RPE-Choroid and retina were 2094.02 and 290.33 days. ng/gm respectively. The time maximum (Tmax) for the ocular tissues was reached on day 7. In the vitreous humour, a lower level of aflibercept was retrieved. The Cmax (1766.84 ng/mL) in the serum was reached on day 1, followed by a decline in the concentration till the end of the study period. In treated eyes, the levels of aflibercept in most of the ocular tissues were maintained for at least 21 days above the invitro IC50 concentration. The results of the efficacy study show that subconjunctival aflibercept could reach the therapeutic target to inhibit CNV. The subconjunctival aflibercept could be a less invasive route for treating CNV with AMD.

Modulating release of ranibizumab and aflibercept from thiolated chitosan-based hydrogels for potential treatment of ocular neovascularization.

BACKGROUND: This paper describes the synthesis of thiolated chitosan-based hydrogels with varying degrees of crosslinking that has been utilized to modulate release kinetics of two clinically relevant FDA-approved anti-VEGF protein drugs, ranibizumab and aflibercept. These hydrogels have been fabricated into disc shaped structures for potential use as patches on ocular surface. METHODS: Protein conformational changes and aggregation after loading and release was evaluated by circular dichroism (CD), steady-state tryptophan fluorescence spectroscopy, electrophoresis and size-exclusion chromatography (SEC). Finally, the capacity of both released proteins to bind to VEGF was tested by ELISA and surface plasmon resonance (SPR) technology. RESULTS: The study demonstrates the versatility of thiolated chitosan-based hydrogels for delivering proteins. The effect of various parameters of the hydrogel on protein release kinetics and mechanism of protein release was studied using the Korsmeyer-Peppas release model. Furthermore, we have studied the stability of released proteins in detail while comparing it with non-entrapped proteins under physiological conditions to understand the effect of formulation conditions on protein stability. CONCLUSIONS: The disc-shaped thiolated chitosan-based hydrogels provide a potentially useful platform to deliver ranibizumab and aflibercept for the treatments of ocular diseases such as wet AMD, DME and corneal neovascularization.

Crizotinib Fails to Enhance the Effect of Radiation in Head and Neck Squamous Cell Carcinoma Xenografts.

AIM: Mesenchymal-epithelial transition factor (MET), a receptor tyrosine kinase, is expressed in head and neck squamous cell carcinomas (HNSCC) and is involved in tumor progression and associated with poor prognosis. MET can be inhibited by crizotinib, a potent ATP-competitive kinase inhibitor. We examined the effects of combining crizotinib and radiation in a pre-clinical HNSCC model. MATERIALS AND METHODS: Nine HNSCC cell lines were screened for MET expression, copy-number amplification and mutational status. The in vitro effects of crizotinib and radiation were assessed with clonogenic survival assays. MET signaling proteins were assessed with western blot and receptor tyrosine kinase array. Tumor growth-delay experiments with UT-SCC-14 and UT-SCC-15 oral tongue xenografts were used to assess in vivo tumor radiosensitivity. RESULTS: All nine HNSCC cell lines showed a varying degree of MET protein and RNA expression. Increased MET copy number was not present. MET was expressed after irradiation both in vitro and in vivo. Crizotinib alone inhibited phosphorylation of MET and inhibited cell growth in vitro but did not inhibit phosphorylation of downstream signaling proteins: MAPK, AKT or c-SRC. When combined with radiation in vitro, crizotinib demonstrated radiation enhancement in only one cell line. Crizotinib did not enhance the effect of radiation in either UT-SCC-14 or UT-SCC-15 tumors grown as xenografts. CONCLUSION: MET is overexpressed in HNSCC cell lines, however, crizotinib failed to enhance the radiation response and failed to inhibit MET downstream signaling proteins in this HNSCC model.

Association of Angiotensin-converting enzyme insertion(I)/deletion (D) genotype in Alzheimer's disease patients of north Indian population.

Several lines of evidence support for the role of angiotensin-converting enzyme (ACE) in Alzheimer's disease (AD) patients. Most human genetic studies have focussed on ACE insertion (I)/deletion (D) polymorphism and have yielded conflicting results. We have evaluated the association of ACE polymorphism with serum ACE activity in 95 AD patients and 110 healthy controls from north Indian population. In Alzheimer's patients a higher frequency of D allele was detected (I/D ratio 0.53:0.47) compared with the control group (I/D ratio 0.54:0.45), the difference being not statistically significant (p > .05). AD patients were found to be more homozygous for the D allele (26.3%) compared with controls (20.8%). The observed genotype distribution was in agreement with Hardy-Weinberg equilibrium. We observed that the D/D genotype is more in patients with a higher serum ACE activity. The D allele and the D/D genotype in AD patients may influence increased risk of cognitive impairment.