Dynamic Regulatory Assessment: Unpacking the Process Reveals Readiness to Pilot for Europe-An EFPIA View.

The medicines regulatory network of the European Economic Area comprises 30 countries, their National Competent Authorities (NCA), and the European Medicines Agency (EMA). The NCAs and EMA are involved at different stages of the medicine life cycle; not all are engaged in a particular medicine's development discussions. As a result, knowledge management (ie, acquisition and transfer between medicine developer and the NCAs) is fragmented and inefficient. Dynamic regulatory assessment (DRA), a regulatory science concept developed by the European Federation of Pharmaceutical Industries and Associations (EFPIA), could drive increased connectedness supporting more continuous knowledge building. DRA works via iterative release and assessment of discrete data packets (DDPs) at mutually agreed milestones during development, culminating in more efficient development and faster authorization. This commentary seeks to build on an earlier article by unpacking the DRA concept, with a particular focus on DDPs. Its aim is to show how DDPs can support efficient and predictable release of data to encourage development and assessment of promising medicines, and it makes the case for piloting the DRA concept with European regulators now.

Lipases of germinating jojoba seeds efficiently hydrolyze triacylglycerols and wax esters and display wax ester-synthesizing activity.

BACKGROUND: Simmondsia chinensis (jojoba) is the only plant known to store wax esters instead of triacylglycerols in its seeds. Wax esters are composed of very-long-chain monounsaturated fatty acids and fatty alcohols and constitute up to 60% of the jojoba seed weight. During jojoba germination, the first step of wax ester mobilization is catalyzed by lipases. To date, none of the jojoba lipase-encoding genes have been cloned and characterized. In this study, we monitored mobilization of storage reserves during germination of jojoba seeds and performed detailed characterization of the jojoba lipases using microsomal fractions isolated from germinating seeds. RESULTS: During 26 days of germination, we observed a 60-70% decrease in wax ester content in the seeds, which was accompanied by the reduction of oleosin amounts and increase in glucose content. The activity of jojoba lipases in the seed microsomal fractions increased in the first 50 days of germination. The enzymes showed higher activity towards triacylglycerols than towards wax esters. The maximum lipase activity was observed at 60 °C and pH around 7 for triacylglycerols and 6.5-8 for wax esters. The enzyme efficiently hydrolyzed various wax esters containing saturated and unsaturated acyl and alcohol moieties. We also demonstrated that jojoba lipases possess wax ester-synthesizing activity when free fatty alcohols and different acyl donors, including triacylglycerols and free fatty acids, are used as substrates. For esterification reactions, the enzyme utilized both saturated and unsaturated fatty alcohols, with the preference towards long chain and very long chain compounds. CONCLUSIONS: In in vitro assays, jojoba lipases catalyzed hydrolysis of triacylglycerols and different wax esters in a broad range of temperatures. In addition, the enzymes had the ability to synthesize wax esters in the backward reaction. Our data suggest that jojoba lipases may be more similar to other plant lipases than previously assumed.

Detailed characterization of the substrate specificity of mouse wax synthase.

Wax synthases are membrane-associated enzymes catalysing the esterification reaction between fatty acyl-CoA and a long chain fatty alcohol. In living organisms, wax esters function as storage materials or provide protection against harmful environmental influences. In industry, they are used as ingredients for the production of lubricants, pharmaceuticals, and cosmetics. Currently the biological sources of wax esters are limited to jojoba oil. In order to establish a large-scale production of desired wax esters in transgenic high-yielding oilseed plants, enzymes involved in wax esters synthesis from different biological resources should be characterized in detail taking into consideration their substrate specificity. Therefore, this study aims at determining the substrate specificity of one of such enzymes -- the mouse wax synthase. The gene encoding this enzyme was expressed heterologously in Saccharomyces cerevisiae. In the in vitro assays (using microsomal fraction from transgenic yeast), we evaluated the preferences of mouse wax synthase towards a set of combinations of 11 acyl-CoAs with 17 fatty alcohols. The highest activity was observed for 14:0-CoA, 12:0-CoA, and 16:0-CoA in combination with medium chain alcohols (up to 5.2, 3.4, and 3.3 nmol wax esters/min/mg microsomal protein, respectively). Unsaturated alcohols longer than 18°C were better utilized by the enzyme in comparison to the saturated ones. Combinations of all tested alcohols with 20:0-CoA, 22:1-CoA, or Ric-CoA were poorly utilized by the enzyme, and conjugated acyl-CoAs were not utilized at all. Apart from the wax synthase activity, mouse wax synthase also exhibited a very low acyl-CoA:diacylglycerol acyltransferase activity. However, it displayed neither acyl-CoA:monoacylglycerol acyltransferase, nor acyl-CoA:sterol acyltransferase activity.