• Lp(a) gene encoding apo(a), bound to apoB-100 by noncovalently and covalently way to assembly of the Lp(a) particles. Lp(a) particles are highly polymorphic particles that are structurally similar to but larger than low-density lipoprotein (LDL). Apo(a) have multiple repeated copies of sequences homologous to plasminogen's kringle 4 domain (KIV), followed by a kringle 5-like (KV) domain and a protease-like domain. Therein, KIV2 multiple copies determine apo(a) isoform size, which is inversely correlated with plasma levels. The Lp(a) concentration is heterogeneous and, to a major extent, controlled by genetics. Other factors such as ethnicity and race, medical and environmental conditions also play roles in Lp(a) regulation. Lp(a) has been associated with increased risks of atherosclerosis, thrombosis, and aortic valve calcification1,2.
• Biocytogen developed an LPA humanized mice, the full coding sequences of humanLpagene that driven by the ALB promoter was inserted into theGt(ROSA)26Sorgene locus site. Then, confirmed that human LPA protein was detected in the serum of heterozygous B-hLPA mice. Meanwhile, Olpasiran analog, a double-stranded small interfering RNA (siRNA) targeting lipoprotein(a), reduced plasma Apo(a) concentrations in heterozygous B-hLPA mice.
• PCSK9 is expressed in liver, intestine and kidney tissues and escorts specific receptors for lysosomal degradation. It plays a role in cholesterol and fatty acid metabolism. PCSK9 binds to the receptor for LDL, if PCSK9 is blocked, more LDLRs are recycled and are present on the surface of cells to remove LDL-particles from the extracellular fluid.
• Biocytogen developed an PCSK9 humanized mice. The genome of mouse Pcsk9 gene including 5’UTR and 3’UTR was replaced by human PCSK9 genome including 5’UTR and 3’UTR in B-hPCSK9 mice plus.
• B-hLPA/hPCSK9 plus micewere derived from mating B-hLPA mice (112723) and hPCSK9 mice plus (112751). For validation data of this mouse model, you can refer to the validation data from the related gene humanized mouse models.
• There is only human Apo(a) and human PCSK9, but not Lp(a) complex in B-hLPA/hPCSK9 plus.

B-hLPA/hPCSK9 plus micewere derived from mating B-hLPA mice (112723) and hPCSK9 mice plus (112751). For validation data of this mouse model, you can refer to the validation data from the related gene humanized mouse models.

Gene targeting strategy for B-hLPA mice.The full coding sequences of humanLpagene that was driven byAlbpromoter was inserted into the Gt(ROSA)26Sor gene locus site. And there’s noLPAgene in the mouse genome sequence.

Gene targeting strategy for B-hPCSK9 mice plus. The genome of mousePcsk9gene including 5’UTR and 3’UTR was replaced by humanPCSK9genome including 5’UTR and 3’UTR in B-hPCSK9 mice plus.

Strain specific PCSK9 expression analysis in homozygous humanized B-hLPA/hPCSK9 plus mice by ELISA. Serum was collected from wild-type C57BL/6 mice (+/+) (6 weeks old, male, n=3) and homozygous B-hLPA/hPCSK9 plus mice (H/H, H/H) (6 weeks old, male, n=3; female, n=3). Protein expression level of PCSK9 was analyzed by ELISA (Human PCSK9 ELISA Kit, Proteintech, KE00278). Human PCSK9 was exclusively detectable in homozygous B-hLPA/hPCSK9 plus mice. Values are expressed as mean ± SEM.

Strain specific Apo(a) protein analysis in homozygous humanized B-hLPA/hPCSK9 plus mice by ELISA. Serum was collected from wild-type C57BL/6 mice (+/+) (6 weeks old, male, n=3) and homozygous B-hLPA/hPCSK9 plus mice (H/H, H/H) (6 weeks old, male, n=3; female, n=3). Protein expression level of Apo(a) was analyzed by ELISA (Human Lipoprotein A ELISA Kit, Abcam, ab212165). Human Apo(a) was exclusively detectable in homozygous B-hLPA/hPCSK9 plus mice. Values are expressed as mean ± SEM.

The inhibitory efficiency of the nucleic acid drugs against human PCSK9 and LPA in B-hLPA/hPCSK9 plus mice. B-hLPA/hPCSK9 plus mice (n=4) were treated with PCSK9-targeted nucleic acid drug and LPA-targeted nucleic acid drug (from a client). The serum was collected to detect the expression levels of human PCSK9 and Apo(a) by ELISA. (A) The expression levels of human PCSK9 protein in serum. (B) The expression levels of human Apo(a) protein in serum. The human PCSK9 levels and Apo(a) levels were significantly reduced after treatment, demonstrating that B-hLPA/hPCSK9 plus mice provide a powerful preclinical model forin vivoevaluation of human PCSK9 and human LPA-targeted nucleic acid drugs. (Data from a client)