B6-hPCSK9(CDS) Mice

Catalog Number: I001222

Strain Name: C57BL/6NCya-Pcsk9em2(hPCSK9)/Cya

Genetic Background: C57BL/6NCya

 

Strain Description

Proprotein convertase subtilisin/kexin 9 (PCSK9) is a serine protease primarily produced in the liver but expressed in other tissues, including the intestine, heart, and neurons. The N-terminal domain of the PCSK9 protein is responsible for protein localization and stability, while the C-terminal domain is responsible for protein enzymatic activity [1]. The Low-density lipoprotein receptor (LDLR) is a receptor that is responsible for clearing low-density lipoprotein cholesterol (LDL-C) from the blood. PCSK9 cleaves the intracellular domain of LDLR on the cell surface, causing it to detach from the cell membrane and be transported to the lysosome for degradation, promoting LDLR degradation, and increasing plasma LDL-C. Overexpression or gain-of-function mutations of the PCSK9 gene can lead to LDL-C accumulation by reducing LDLR levels. This can cause hypercholesterolemia, which increases the risk of cardiovascular diseases, such as atherosclerosis and coronary heart disease, and neurodegenerative diseases, such as Alzheimer's disease [2]. PCSK9 has become an important target for the development of lipid-lowering drugs. Several PCSK9-targeted antibodies or small nucleic acid drugs have been approved for marketing worldwide, including evolocumab from Amgen, alirocumab from Sanofi and Regeneron, and inclisiran from Novartis. These drugs primarily work by inhibiting PCSK9 activity or preventing PCSK9 protein from binding to LDLR, lowering LDL-C levels in the blood to treat hypercholesterolemia [3-4]. In addition, PCSK9 can promote tumor growth and development by regulating cell proliferation, migration, and invasion. It can also regulate the expression of inflammatory factors that contribute to inflammation. Therefore, targeting the expression of PCSK9 has been investigated in tumor immunotherapy and autoimmune disease therapy [5-6].

B6-hPCSK9(CDS) mice are generated by integrating the human PCSK9 protein coding sequence (CDS) and 3' untranslated region (UTR) into the mouse Pcsk9 gene locus using gene editing techniques, resulting in a model expressing human PCSK9 protein while disrupting the expression of the mouse endogenous Pcsk9 gene. These mice can be used in the study of various metabolic diseases, neurodegenerative diseases, tumor development, and autoimmune diseases, as well as for the development, screening, and preclinical pharmacological evaluation of PCSK9-targeted drugs. Compared to the similar B6-hPCSK9 mice (PCSK9 genomic humanized model, Catalog Number: I001179), B6-hPCSK9(CDS) mice have higher levels of human PCSK9 protein expression in serum, while LDLR expression is closer to the normal endogenous levels in mice.

Strain Strategy


Figure 1. Gene editing strategy of B6-hPCSK9(CDS) mice. The partial sequence of exon 1 of the mouse Pcsk9 gene was replaced with the "Kozak-Human PCSK9 CDS Human PCSK9 3'UTR-WPRE BGH pA" gene expression element.

Applications

  • Development and screening of PCSK9-targeted therapies;
  • Preclinical pharmacological and efficacy evaluation of PCSK9-targeted therapies;
  • Research on metabolic diseases such as hypercholesterolemia, atherosclerosis, and coronary heart disease;
  • Research on neurodegenerative diseases such as stroke and Alzheimer's disease.

 

Validation Data

1. Human PCSK9 gene and mouse Pcsk9 gene expression


Figure 2. Expression of human PCSK9 gene and mouse Pcsk9 gene in liver and colon tissues of 6-week-old homozygous B6-hPCSK9(CDS) mice and wild-type (WT) mice. RT-qPCR results show significant expression of human PCSK9 gene in the liver and colon of homozygous B6-hPCSK9(CDS) mice, with no expression of mouse Pcsk9 gene. Wild-type mice express only the mouse Pcsk9 gene and not the human PCSK9 gene. Additionally, the expression of human PCSK9 gene in the liver of B6-hPCSK9(CDS) mice is higher than in the colon, consistent with the expression pattern of endogenous mouse Pcsk9 (Bars represent mean ± SEM, n≥3). (ND: Not detected)

2. Human PCSK9 protein expression

Figure 3. Expression of human PCSK9 protein in the serum of wild-type (WT) mice and homozygous B6-hPCSK9(CDS) mice (6 weeks old). ELISA using human-specific antibodies shows significant expression of human PCSK9 protein in the serum of B6-hPCSK9(CDS) mice, with no expression detected in wild-type mice serum (Bars represent mean ± SEM, n=4).

3. Ldlr gene expression

Figure 4. Expression of mouse Ldlr gene in liver tissues of 6-week-old homozygous B6-hPCSK9(CDS) mice and wild-type (WT) mice. RT-qPCR results show that the expression level of mouse Ldlr gene in the liver of B6-hPCSK9(CDS) mice is comparable to that of wild-type mice (Bars represent mean ± SEM, n≥3).

4. LDLR protein expression

Figure 5. Western Blot analysis of LDLR protein expression in the liver of 6-week-old homozygous B6-hPCSK9(CDS) mice, B6-hPCSK9 mice (PCSK9 genomic humanized model, Catalog Number: I001179), and wild-type (WT) mice. Results show that the LDLR protein expression level in the liver of B6-hPCSK9(CDS) mice is comparable to that of wild-type mice, while the expression level in B6-hPCSK9 mice (Catalog Number: I001179) is significantly higher than in B6-hPCSK9(CDS) mice (Catalog Number: I001222) and wild-type mice.

 

References
[1]Melendez QM, Krishnaji ST, Wooten CJ, Lopez D. Hypercholesterolemia: The role of PCSK9. Arch Biochem Biophys. 2017 Jul 1;625-626:39-53.
[2]Seidah NG, Awan Z, Chrétien M, Mbikay M. PCSK9: a key modulator of cardiovascular health. Circ Res. 2014 Mar 14;114(6):1022-36.
[3]Pasta A, Cremonini AL, Pisciotta L, Buscaglia A, Porto I, Barra F, Ferrero S, Brunelli C, Rosa GM. PCSK9 inhibitors for treating hypercholesterolemia. Expert Opin Pharmacother. 2020 Feb;21(3):353-363.
[4]Sabatine MS. PCSK9 inhibitors: clinical evidence and implementation. Nat Rev Cardiol. 2019 Mar;16(3):155-165.
[5]Ding Z, Pothineni NVK, Goel A, Lüscher TF, Mehta JL. PCSK9 and inflammation: role of shear stress, pro-inflammatory cytokines, and LOX-1. Cardiovasc Res. 2020 Apr 1;116(5):908-915.
[6]Liu X, Bao X, Hu M, Chang H, Jiao M, Cheng J, Xie L, Huang Q, Li F, Li CY. Inhibition of PCSK9 potentiates immune checkpoint therapy for cancer. Nature. 2020 Dec;588(7839):693-698.