Targeting GNE Myopathy: A Dual Prodrug Approach for the Delivery of N-Acetylmannosamine 6-Phosphate

Abstract

ProTides comprise an important class of prodrugs currently marketed and developed as antiviral and anticancer therapies. The ProTide technology employs a phosphate masking groups capable of providing more favorable drug-like properties and an intracellular activation mechanism for enzyme-mediated release of a nucleoside monophosphate. Herein we describe the application of phosphoramidate chemistry to 1,3,4-O-acetylated N-acetylmannosamine (Ac3ManNAc) to deliver ManNAc-6-phosphate (ManNAc-6-P), a critical intermediate in sialic acid biosynthesis. Sialic acid deficiency is a hallmark of GNE myopathy, a rare congenital disorder of glycosylation (CDG) caused by mutations in GNE that limit the production of ManNAc-6-P. Synthetic methods were developed to provide a library of Ac3ManNAc-6phosphoramidates that were evaluated in a series of studies for their potential as a treatment for GNE myopathy. Prodrug 12b showed rapid activation in a carboxylesterase (CPY) enzymatic assay and favorable ADME properties, while also being more effective than ManNAc at increasing sialic acid levels in GNE-deficient cell lines than ManNAc. These 2 results provide a potential platform to address substrate deficiencies in GNE myopathy and other CDGs.

Synthesis of 2‐Acetamido‐1,3,4‐Tri‐O‐Acetyl‐2‐Deoxy‐D‐Mannopyranose ‐6‐Phosphate Prodrugs as Potential Therapeutic Agents

“…This causes severe digestion problems in the patient, which has led in several cases to their abandoning of the trial or the treatment. By delivering a more lipophilic prodrug (the acetyl groups contribute to this lipophilicity) the passive diffusion will allow the administration of a significantly lower dosage of the compound, with a drastic reduction of the side effects (Morozzi et al., 2019). Secondly, the direct delivery of the ManNac‐6P will bypass, completely, the defective enzyme.…”

“…In this protocol, we report the preparation of ManNAc ‐6‐phosphate prodrugs as a potential therapeutic agent for GNE myopathy. The rationale is that when the prodrug is activated by enzymatic removal of phosphate masking groups and acetyl groups from the sugar hydroxyl groups, it will release the product of the defective GNE enzyme (providing a source of ManNac‐6P) thereby restoring the level of sialic acid and rescuing the GNE phenotype (Morozzi et al., 2019). This strategy is advantageous over the administration of simple ManNac as a nutritional supplement (clinical trial NCT04231266) because of the following: due to their high polarity, carbohydrate molecules do not passively cross the gut membrane and require specific transporter proteins to reach the bloodstream.…”

Ceritinib is a novel triple negative breast cancer therapeutic agent

“…Using the molecular-based sub-classifications, FDA-approved tumor actionable drugs may be repurposed to treat chemo-resistant subtype-specific TNBC tumors. Bicalutamide and enzalutamide are FDA-approved AR antagonists for prostate cancer treatment [ 15 ]. AR + TNBCs respond to these AR antagonists’ treatment in vitro and in vivo [ 11 , 16 ].…”

A mild and concise synthesis of aryloxy phosphoramidate prodrug of alcohols via transesterification reaction

“…4 The nature of different components of the phosphoramidate core is essential for the prodrug's potency, especially for the case of non-nucleoside drug candidates in which other amino acid motifs are more effective than the commonly used l -alanine. 5 As a result, SAR studies of amino acid ester and aryl moieties would be necessary to identify the optimal combination of these masking groups when applying ProTide technology to a new therapeutic chemical entity. Consequently, an efficient method capable of rapidly assembling aryloxy phosphoramidate prodrug library from parent drug would be very attractive to the discovery of new ProTide prodrugs.…”

“…While previous studies suggested that l -alanine was optimal for antiviral and anticancer ProTide prodrugs, some new SAR data obtained in the studies of non-nucleoside drug candidates indicated that other amino acids, in some cases, might be more effective than l -alanine. 5 Therefore, we decided to explore the scope of N -diphenylphosphoryl amino acid esters (Table 2). Under standard conditions, a variety of l -alanine esters were tolerated, affording desired aryloxy phosphoramidates prodrugs in good yields (products 4–6 ).…”