Kishor M. Wasan

Efficacy of Postnatal In Vivo Nonsense Suppression Therapy in a Pax6 Mouse Model of Aniridia

Nonsense mutations leading to premature stop codons are common occurring in approximately 12% of all human genetic diseases. Thus, pharmacological nonsense mutation suppression strategies would be beneficial to a large number of patients if the drugs could be targeted to the affected tissues at the appropriate time. Here, we used nonsense suppression to manipulate Pax6 dosage at different developmental times in the eye of the small eye (Pax6Sey/+; G194X) mouse model of aniridia. Efficacy was assessed by functional assays for visual capacity, including electroretinography and optokinetic tracking (OKT), in addition to histological and biochemical studies. Malformation defects in the Pax6Sey/+ postnatal eye responded to topically delivered nonsense suppression in a dose- and time-dependent manner. Elevated levels of Mmp9, a direct downstream target of Pax6 in the cornea, were observed with the different treatment regimens. The lens capsule was particularly sensitive to Pax6 dosage, revealing a potential new role for Pax6 in lens capsule maintenance and development. The remarkable capacity of malformed ocular tissue to respond postnatally to Pax6 dosage in vivo demonstrates that the use of nonsense suppression could be a valuable therapeutic approach for blinding diseases caused by nonsense mutations.

Review and analysis of FDA approved drugs using lipid-based formulations

Abstract Lipid-based drug delivery systems (LBDDS) are one of the most studied bioavailability enhancement technologies and are utilized in a number of U.S. Food and Drug Administration (FDA) approved drugs. While researchers have used several general rules of thumb to predict which compounds are likely to benefit from LBDDS, formulation of lipid systems is primarily an empiric endeavor. One of the challenges is that these rules of thumb focus in different areas and are used independently of each other. The Developability Classification System attempts to link physicochemical characteristics with possible formulation strategies. Although it provides a starting point, the formulator still has to empirically develop the formulation. This article provides a review and quantitative analysis of the molecular properties of these approved drugs formulated as lipid systems and starts to build an approach that provides more directed guidance on which type of lipid system is likely to be the best for a particular drug molecule.

Lipid-lowering Activity of Natural and Semi-Synthetic Sterols and Stanols.

Consumption of plant sterols/ stanols has long been demonstrated to reduce plasma cholesterol levels. The objective of this review is to demonstrate the lipid-lowering activity and anti-atherogenic effects of natural and semi-synthetic plant sterols/ stanols based on evidence from cell-culture studies, animal studies and clinical trials. Additionally, this review highlights certain molecular mechanisms by which plant sterols/ stanols lower plasma cholesterol levels with a special emphasis on factors that affect the cholesterol-lowering activity of plant sterols/stanols. The crystalline nature and the poor oil solubility of these natural products could be important factors that limit their cholesterol-lowering efficiency. Several attempts have been made to improve the cholesterol-lowering activity by enhancing the bioavailability of crystalline sterols and stanols. Approaches involved reduction of the crystal size and/or esterification with fatty acids from vegetable or fish oils. However, the most promising approach in this context is the chemical modification of plant sterols /stanols into water soluble disodium ascorbyl phytostanyl phosphates analogue by esterification with ascorbic acid. This novel semi-synthetic stanol derivative has improved efficacy over natural plant sterols/ stanols and can provide additional benefits by combining the cholesterol-lowering properties of plant stanols with the antioxidant potential of ascorbic acid. This article is open to POST-PUBLICATION REVIEW. Registered readers (see For Readers) may comment by clicking on ABSTRACT on the issues contents page.

Physician centric healthcare: is it time for a paradigm shift?:

Kishor M Wasan, Lois Berry and Jawahar Kalra College of Pharmacy and Nutrition, University of Saskatchewan, Saskatoon, Saskatchewan S7N 5A2, Canada Office of the Vice Provost Health, University of Saskatchewan, Saskatoon, Saskatchewan S7N 5A2, Canada College of Nursing, University of Saskatchewan, Saskatoon, Saskatchewan S7N 5A2, Canada College of Medicine, University of Saskatchewan, Saskatoon, Saskatchewan S7N 5A2, Canada Corresponding author: Kishor M Wasan. Email: Kishor.Wasan@usask.ca

Bone-seeking agents for the treatment of bone disorders

The targeting and delivery of therapeutic and diagnostic agents to bone tissue presents both a challenge and opportunity. Osteoporosis, Paget’s disease, cancer, and bone metastases are all skeletal diseases whose treatment would benefit from new targeted therapeutic strategies. Osteoporosis, in particular, is a very prevalent disease, affecting over one in three women and one in five men in Canada alone with the cost to the healthcare system estimated at over

Scavenger receptor class BI (SR-BI) mediates uptake of CPX-351 into K562 leukemia cells

2.3xa0billion in 2010. Bone tissue is often considered a rigid structure when in reality there is a process of continuous remodeling that takes place via complex endocrine-regulated cell signaling pathways in addition to the signaling pathways unique to bone tissue. It is these specific boneremodeling processes that provide unique targeting opportunities but also present a number of challenges.

Development and in vitro characterization of chitosan-coated polymeric nanoparticles for oral delivery and sustained release of the immunosuppressant drug mycophenolate mofetil

Abstract Purpose: CPX-351 is a liposomal formulation of cytarabine and daunorubicin encapsulated at a 5:1 molar ratio, for the treatment of acute myeloid leukemia. The Scavenger Receptor class B type I (SR-BI) plays an important role in mediating the uptake of high-density lipoproteins. The purpose of this study is to assess the role of the cell surface lipoprotein receptor SR-BI in the uptake of CPX-351 liposomes (Jazz Pharmaceuticals) into K562 leukemia cells. Methods: K562 cells were pre-treated with 10u2009nM siRNA for 48u2009h and then treated with varying amount of CPX-351 for 24, 48 and 72u2009h. Cells were then collected and analyzed at 480/590u2009nm on a CytoFLEX Multicolour flow instrument to determine cellular uptake of daunorubicin. Experimental data were analyzed using two-way ANOVA with Bonferroni multiple comparisons. Significance was set at pu2009<u2009.05. Results: K562 cells pre-treated with SR-BI siRNA for 48u2009h had a reduced SRB1 cell surface concentration (74–85%). Addition of CPX-351 at 10–50u2009nM followed by measurement of cellular daunorubicin at 48, 48 or 72u2009h showed a significantly lower percentage of daunorubicin positive population compared with control K562 cells (pu2009<u2009.05). There was significantly less daunorubicin taken up in the SR-BI knock-down cells across all drug concentrations and at all three time points, although there were no concentration-related trends. Conclusions: These preliminary studies suggest that SR-BI may be one potential mechanism by which CPX-351 is taken up into K562 cells.

World Health Organization and Essential Medicines

Abstract Objective: To develop an oral sustained release formulation of mycophenolate mofetil (MMF) for once-daily dosing, using chitosan-coated polylactic acid (PLA) or poly(lactic-co-glycolic) acid (PLGA) nanoparticles. The role of polymer molecular weight (MW) and drug to polymer ratio in encapsulation efficiency (EE) and release from the nanoparticles was explored in vitro. Methods: Nanoparticles were prepared by a single emulsion solvent evaporation method where MMF was encapsulated with PLGA or PLA at various polymer MW and drug: polymer ratios. Subsequently, chitosan was added to create coated cationic particles, also at several chitosan MW grades and drug: polymer ratios. All the formulations were evaluated for mean diameter and polydispersity, EE as well as in vitro drug release. Differential scanning calorimetry (DSC), surface morphology, and in vitro mucin binding of the nanoparticles were performed for further characterization. Results: Two lead formulations comprise MMF: high MW, PLA: medium MW chitosan 1:7:7 (w/w/w), and MMF: high MW, PLGA: high MW chitosan 1:7:7 (w/w/w), which had high EE (94.34% and 75.44%, respectively) and sustained drug release over 12u2009h with a minimal burst phase. DSC experiments revealed an amorphous form of MMF in the nanoparticle formulations. The surface morphology of the MMF NP showed spherical nanoparticles with minimal visible porosity. The potential for mucoadhesiveness was assessed by changes in zeta potential after incubation of the nanoparticles in mucin. Conclusion: Two chitosan-coated nanoparticles formulations of MMF had high EE and a desirable sustained drug release profile in the effort to design a once-daily dosage form for MMF.

Abstract 1800: Inhibition of scavenger receptor class B type I suppresses androgen pathway activity and induces cytotoxic stress in C4-2 castration resistant prostate cancer cells

In June 2017, the World Health Organization released 20th Model List of Essential Medicines for adults and sixth Model List of Essential Medicines for children. In our commentary, we describe the changes to the Essential Medicine list, and identify deficits in excluding medicines for management of diseases with a high burden. In using tracer conditions such as cardiovascular and thromboembolic disease, mental health, and diseases of the musculoskeletal system, we highlight the absence of several medicines, which are incorporated into major clinical practice guidelines. We recommend that the World Health Organization review its process with respect to identifying disease conditions as well as evidence-based therapies.

A check-up on Canada's health system

Proceedings: AACR 107th Annual Meeting 2016; April 16-20, 2016; New Orleans, LAnnExisting therapies for castration-resistant prostate cancer (CRPC) extend life and provide clinical benefit; however, patients continue to develop therapeutic resistance. Androgen pathway activity persists in CRPC even under maximal androgen blockade conditions. A proposed mechanism for continued androgen pathway signaling is de novo intratumoral synthesis of androgen receptor agonists from the precursor cholesterol. The high density lipoprotein(HDL)-cholesterol receptor, scavenger receptor class B type I (SR-BI), is upregulated in CRPC models in vitro and in vivo. Here, we test the hypothesis that SR-BI is a source of cholesterol for de novo steroidogenesis in CRPC cells using the castration-resistant LNCaP-derived cell line, C4-2. Cells were transfected with either non-targeted (NC) or stealth RNAi duplexes (Thermo Fisher) targeting SR-BI to silence protein expression (SR-B1 KD). Prostate specific antigen (PSA) levels in media from SR-B1 KD samples were reduced to 39% of that seen in control cell media (20.9 ± 1.4 ng/mL/μg protein SRBI-KD vs. 34.5 ± 2.4 ng/mL/μg protein NC; n = 4, p < 0.05). Intracellular testosterone concentrations measured by LC-MS showed a 2-fold reduction in SRBI-KD samples (0.20 ng/mL/mg) compared to NC (0.41 ng/mL/mg; n = 1). Additionally, SR-BI KD cells exhibited reduced cell viability as measured by MTS assay and induction of G1/S cell cycle arrest as assessed by propidium iodide staining cell cycle analysis by flow cytometry (70.9 ± 7.9% G0-G1 phase fraction SRBI-KD vs. 58.3 ± 4.1% NC; n = 4, p < 0.05). SR-BI KD cells exhibited evidence of increased cell stress relative to NC cells. SR-BI-KD cells showed elevated induction of autophagy as assessed by measuring changes in LC3-I:LC3-II ratio by immunoblotting and formation of autophagosomes by immunofluorescence, alongside induction of senescence as assessed by measuring senescence associated beta-galactosidase (SA-β-gal) activity using flow cytometry analysis and a fluorogenic substrate. Further, treating C4-2 cells with an established SR-B1 inhibitor, BLT-1, lead to decrease PSA secretion, and co-treatment with the common cholesterol synthesis inhibitor, simvastatin, synergistically decreased PSA secretion from C4-2 cells. These results suggest that under androgen-deprived conditions, SR-BI targeting can decrease androgen pathway signalling and induce cellular stress, leading to G1/S arrest and decreased proliferation in the absence of an apoptotic induction. Lastly, combined inhibition of cholesterol uptake (BLT-1) and synthesis (Simvastatin) lead to a synergistic decrease in PSA secretion indicating the possibility that a multi-faceted cholesterol blockade approach to CRPC treatment may improve response to maximal hormone blockade by disrupting intratumoral steroid production.nnCitation Format: Jacob A. Gordon, Ankur Midha, Mitali Pandey, Kishor Wasan, Michael E. Cox. Inhibition of scavenger receptor class B type I suppresses androgen pathway activity and induces cytotoxic stress in C4-2 castration resistant prostate cancer cells. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 1800.