Methods for determining the efficacy profile of a drug candidate

Abstract

This patent application relates to a method for determining the in vitro efficacy profile of a drug candidate using standardized cell cultures of uniformly distributed differentiated neural cells (NCs) from at least two primate species, wherein the differentiated NC cultures are qualified for high throughput screening based on a dissociation and reseeding step performed on the differentiated NCs. The method includes differentiating human and/or non-human primate neuronal precursor cells (NPCs) to neuronal cells (NCs) followed by dissociating the differentiated NCs from its support and reseeding the differentiated NCs in a high-throughput cell culture format resulting in robust cultures suitable for high-throughput drug screening assays, in particular to screen antisense oligonucleotides.

1 . A method for determining the in vitro efficacy profile of a drug candidate using standardized cell cultures of uniformly distributed differentiated neural cells (NCs) from at least two primate species, wherein the differentiated NC cultures are qualified for high throughput screening, the method comprising the steps of:
 a) dissociating the differentiated NCs from its support after about 20 days to about 45 days of differentiation and reseeding the differentiated NCs in a high-throughput cell culture format;   b) incubating the reseeded NCs in a differentiation medium;   c) contacting the reseeded NCs with the drug candidate; and   d) determining the in vitro efficacy profile of the drug candidate.   
     
     
         2 . The method according to  claim 1 , wherein the primate species are selected from the group consisting of human ( Homo sapiens ), Cynomolgus monkey ( Macaca fascicularis ) and Rhesus monkey ( Macaca mulatta ). 
     
     
         3 . The method according to of any one of  claim 1  or  2 , wherein the differentiated NCs are derived from induced pluripotent stem cells (iPSCs). 
     
     
         4 . The method according to  claim 1 , wherein the differentiated NCs are uniformly distributed over the cell culture area as assessed by cell nucleus staining. 
     
     
         5 . The method according to  claim 1 , wherein step d) additionally comprises monitoring the cell cultures for signs of toxicity. 
     
     
         6 . The method according to  claim 1 , wherein step d) comprises monitoring the cell cultures for a phenotypic change indicative of the efficacy of the drug candidate. 
     
     
         7 . The method according to  claim 1 , wherein the determined in vitro efficacy profile of the drug candidate is used for inter-species comparison of the efficacy profile of a drug candidate, wherein the cell cultures are produced individually from cells of at least two primate species, wherein essentially the same conditions are applied to the cultures for all primate species and wherein the efficacy profile is determined and compared for all primate species. 
     
     
         8 . A method for selecting a drug candidate for further development comprising the steps of:
 (i) determining the in vitro efficacy profile of the drug candidate for a first and a second species according to the method of any one of  claims 1  to  7 ; and   (ii) selecting the drug candidate for further development if the efficacy profile of the drug candidate is favourable.   
     
     
         9 . The method according to  claim 8 , wherein the first species is cynomolgus monkey ( Macaca fascicularis ) and the second species is human ( Homo sapiens ). 
     
     
         10 . The method according to  claim 8 , wherein the drug candidate comprises a nucleic acid molecule or targets a specific nucleic acid sequence. 
     
     
         11 . The method according to  claim 8 , wherein the drug candidate comprises at least one nucleic acid molecule such as a RNAi agent or an antisense oligonucleotide. 
     
     
         12 . The method according to  claim 8 , wherein the further development comprises determining the in vivo efficacy and/or toxicity profile of the drug candidate. 
     
     
         13 . A method for determining the potential in vivo efficacy of a drug candidate wherein the in vitro efficacy profile of a drug candidate is determined according to any one of  claims 1  to  7  and wherein the in vitro efficacy profile is indicative for in vivo efficacy. 
     
     
         14 . The method according to  claim 13 , wherein the in vivo efficacy profile is determined in cynomolgus monkey ( Macaca fascicularis ). 
     
     
         15 . The method according to  claim 14 , wherein the determined in vitro efficacy profile and/or in vivo efficacy profile of the drug candidate is indicative for in vivo efficacy in human ( Homo sapiens ). 
     
     
         16 . (canceled)