title: HuCAL PLATINUM, a synthetic Fab library optimized for sequence diversity and superior performance in mammalian expression systems.
abstract: This article describes the design of HuCAL (human combinatorial antibody library) PLATINUM, an optimized, second-generation, synthetic human Fab antibody library with six trinucleotide-randomized complementarity-determining regions (CDRs). Major improvements regarding the optimized antibody library sequence space were implemented. Sequence space optimization is considered a multistep process that includes the analysis of unproductive antibody sequences in order to, for example, avoid motifs such as potential N-glycosylation sites, which are undesirable in antibody production. Gene optimization has been used to improve expression of the antibody master genes in the library context. As a result, full-length IgGs derived from the library show both significant improvements in expression levels and less undesirable glycosylation sites when compared to the previous HuCAL GOLD library. Additionally, in-depth analysis of sequences from public databases revealed that diversity of CDR-H3 is a function of loop length. Based upon this analysis, the relatively uniform diversification strategy used in the CDR-H3s of the previous HuCAL libraries was changed to a length-dependent design, which replicates the natural amino acid distribution of CDR-H3 in the human repertoire. In a side-by-side comparison of HuCAL GOLD and HuCAL PLATINUM, the new library concept led to isolation of about fourfold more unique sequences and to a higher number of high-affinity antibodies. In the majority of HuCAL PLATINUM projects, 100-300 antibodies each having different CDR-H3s are obtained against each antigen. This increased diversity pool has been shown to significantly benefit functional antibody profiling and screening for superior biophysical properties.
authors.affiliation: MorphoSys AG, Lena-Christ-Strasse 48, 82152 Martinsried, Germany. Josef.Prassler@morphosys.com
authors.forename: Josef
authors.initials: J
authors.lastname: Prassler
authors.forename: Stefanie
authors.initials: S
authors.lastname: Thiel
authors.forename: Catrin
authors.initials: C
authors.lastname: Pracht
authors.forename: Andrea
authors.initials: A
authors.lastname: Polzer
authors.forename: Solveig
authors.initials: S
authors.lastname: Peters
authors.forename: Marion
authors.initials: M
authors.lastname: Bauer
authors.forename: Stephanie
authors.initials: S
authors.lastname: Nörenberg
authors.forename: Yvonne
authors.initials: Y
authors.lastname: Stark
authors.forename: Johanna
authors.initials: J
authors.lastname: Kölln
authors.forename: Andreas
authors.initials: A
authors.lastname: Popp
authors.forename: Stefanie
authors.initials: S
authors.lastname: Urlinger
authors.forename: Markus
authors.initials: M
authors.lastname: Enzelberger
mesh_terms: D015870:Gene Expression; D015723:Gene Library; D014644:Genetic Variation; D005822:Genetic Vectors; D006031:Glycosylation; D006801:Humans; D007140:Immunoglobulin Fab Fragments / Q000235:genetics*
chemical_list: D007140:Immunoglobulin Fab Fragments