PCSK9: A hepatic secretory protein that governs circulating LDL-C concentrations by modulating LDL receptor (LDL-R) turnover on hepatocyte surface

• Gene Information: Proprotein convertase subtilisin/kexin type 9 (PCSK9) belongs to the serine protease family.
• Protein Expression: It is mainly synthesized by the liver, with detectable expression in extrahepatic tissues such as the intestines, myocardium, and neurons.
• Signaling Pathway: Low-density lipoprotein receptor (LDLR) functions to remove LDL-C from the bloodstream. PCSK9 cleaves the intracellular segment of cell-surface LDLR, driving its lysosomal degradation. Loss of functional LDLR further raises circulating LDL-C levels.
• Therapeutic Method: PCSK9 inhibitors exert therapeutic effects by hindering PCSK9-mediated degradation of LDL receptors. This intervention preserves abundant functional LDL receptors on hepatocyte surfaces, which in turn enhances the hepatic clearance efficiency of circulating LDL cholesterol.

LPA

• The full coding sequences (CDS) of the human LPA gene, which is driven by the Alb promoter, were inserted into the Gt(ROSA)26Sor gene locus site. And there’s no LPA gene in the mouse genome sequence.

APOB

• The exons 1-29 of the mouse Apob gene that encode the whole molecule (ATG to STOP codon), including 5’UTR and 3’UTR, were replaced by human counterparts in B-hAPOB mice. The human APOB expression is driven by the human APOB promoter, while the mouse Apob gene transcription and translation will be disrupted.

PCSK9

• The genome of the mouse Pcsk9 gene, including 5’UTR and 3’UTR, was replaced by human counterparts in B-hPCSK9 mice plus. The human PCSK9 expression is driven by the human PCSK9 promoter, while the mouse Pcsk9 gene transcription and translation will be disrupted.

B-hLPA/hAPOB/hPCSK9 plus mice were derived from mating B-hLPA mice (112723), B-hAPOB mice (112951), 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.

• Human Apo(a), APOB, and PCSK9 was exclusively detectable in homozygous B-hLPA/hAPOB/hPSCK9 plus mice.

Protein expression analysis in homozygous humanized B-hLPA/hAPOB/hPCSK9 plus mice by ELISA. Serum was collected from homozygous B-hLPA/hAPOB/hPCSK9 plus mice (H/H, H/H, H/H) (6-week-old, male and female, n=3 per sex). Protein expression level of Apo(a), APOB, and PCSK9 were analyzed by ELISA (Apo(a): Abcam, ab212165; APOB: Abcam, ab108807; PCSK9: Proteintech, KE00278). Human Apo(a), APOB, and PCSK9 were detectable in homozygous B-hLPA/hAPOB/hPCSK9 plus mice. Values are expressed as mean ± SEM.

Biochemical test of B-hLPA/hAPOB/hPCSK9 plus mice. Values are expressed as mean ± SD.

The inhibitory efficiency of the nucleic acid drugs against human LPA and PCSK9 in B-hLPA/hAPOB/hPCSK9 plus mice. B-hLPA/hAPOB/hPCSK9 plus mice were randomly divided into four groups (6-week-old, n=2-3 per group). Olpasiran-analog (provided by a client), Inclisiran, and the combination treatment were administered to the mice individually. Serum were collected to measure the Apo(a) (Abcam, ab212165) and PCSK9 (R&D, DPC900) by ELISA.

•  LPA and PCSK9 siRNA reduce the expression of Lp(a) and PCSK9 after a single dose treatment.

The inhibitory efficiency of the nucleic acid drugs against human LPA and PCSK9 in B-hLPA/hAPOB/hPCSK9 plus mice. B-hLPA/hAPOB/hPCSK9 plus mice were randomly divided into four groups (6-week-old, n=2-3 per group). Olpasiran-analog (provided by a client), Inclisiran, and the combination treatment were administered to the mice individually. Serum were collected to measure the Apo(a) (Abcam, ab212165) and PCSK9 (R&D, DPC900). (A) % Mean change of human PCSK9 relative to baseline. (B) % Mean change of human Apo(a) relative to baseline after administration. Values are expressed as mean ± SEM.

In vivo efficacy of WD-induced B-hLPA/hAPOB/PCSK9 plus mice. B-hLPA/hAPOB/hPCSK9 plus mice were fed with Western Diet (XT079B, 40% energy from fat and 0.15% cholesterol) for 5 weeks at first, and B-hLPA/hAPOB/hPCSK9 plus mice were divided into four groups according to the LDL-C. Olpasiran-analog (synthesized according to patents), Inclisiran, and the combination treatment were administered to the mice individually.

• LPA and PCSK9 siRNA reduce the LDL-C after a single dose treatment.

In vivo efficacy of WD-induced B-hLPA/hAPOB/PCSK9 plus mice. B-hLPA/hAPOB/hPCSK9 plus mice were fed with Western Diet (XT079B, 40% energy from fat and 0.15% cholesterol) for 5 weeks at first, and B-hLPA/hAPOB/hPCSK9 plus mice were divided into four groups according to the LDL-C. Olpasiran-analog (synthesized according to patents), Inclisiran, and the combination treatment were administered to the mice individually. (A) LDL-C change after WD induction. (B) LDL-C change after treatment. Values are expressed as mean ± SEM. WD: Western Diet.