• Amyloid precursor protein (APP) is a 100-140 kDa transmembrane glycoprotein that plays a key role in the pathogenesis of Alzheimer's disease (AD). In the disease state, β-secretase and γ-secretase aberrantly cleave APP, resulting in the release of amyloid β (Aβ) peptides Aβ40 and Aβ42, which are neurotoxic fragments capable of oligomerizing, aggregating, and subsequently forming plaques.
• Gene editing strategy: The partial 5’UTR and CDS that encodes the full-length human APP protein, followed by human 3’UTR-STOP is inserted right after mouse App 5’UTR to replace the exon 1 of mouse App gene. The APP protein expression will be driven by endogenous mouse App promoter, while mouse App gene transcription and translation will be disrupted.
• mRNA expression analysis: Mouse App mRNA was detectable in wild-type C57BL/6JNifdc mice. Human APP mRNA was detectable in homozygous B-hAPP mice but not in wild-type mice.
• Protein expression analysis: Human APP was only detectable in the cortex, hippocampus, cerebellum and spinal cord from homozygous B-hAPP mice but not in wild-type mice.
• In vivo efficacy: The human APP mRNA was significantly reduced in the hippocampus, cortex and spinal cord after administration of ALN-APP analog, demonstrating that B-hAPP mice provide a powerful preclinical model for in vivo evaluation of human APP targeted nucleic acid drugs.
• Application: This product is used for pharmacodynamics and safety evaluation of Alzheimer's disease (AD).

Gene targeting strategy for B-hAPP mice. The partial 5’UTR and CDS that encodes the full-length human APP protein, followed by human 3’UTR-STOP is inserted right after mouse App 5’UTR to replace the exon 1 of mouse App gene. The APP protein expression will be driven by endogenous mouse App promoter, while mouse App gene transcription and translation will be disrupted.

Strain specific analysis of APP mRNA expression in wild-type C57BL/6JNifdc mice and homozygous B-hAPP mice by RT-PCR. Cortex RNA was isolated from wild-type C57BL/6JNifdc mice (+/+) and homozygous B-hAPP mice (H/H), then cDNA libraries were synthesized by reverse transcription, followed by PCR with mouse or human APP primers. Mouse App mRNA was detectable in wild-type mice. Human APP mRNA was detectable in homozygous B-hAPP mice, but not in wild-type mice.

Western blot analysis of APP protein expression in homozygous B-hAPP mice. Various tissue lysates were collected from wild-type C57BL/6JNifdc mice (+/+) and homozygous B-hAPP mice (H/H), and then analyzed by western blot with species-specific anti-APP antibody (abcam, ab133588). 40 μg total proteins were loaded for western blotting analysis. Human APP was detectable only in cortex, hippocampus, cerebellum and spinal cord from homozygous B-hAPP mice but not in wild-type mice.

The inhibitory efficiency of the siRNA drugs against human APP in B-hAPP mice. B-hAPP mice were randomly divided into two groups (n=4/group, 10-week-old, male). The ALN-APP analog (in-house) and vehicle were administered to the mice individually by intra-cerebroventricular injection (ICV). The mice were sacrificed on day 14 and day 28, respectively. Then the cortex, hippocampus and spinal cord tissue were collected to detect the human APP mRNA by qRT-PCR. (A) The schematic diagram of experimental processing. (B) The expression of human APP mRNA in cortex, hippocampus and spinal cord. The human APP mRNA in the treatment group (G2) was significantly reduced compared to the control group (G1), demonstrating that B-hAPP mice provide a powerful preclinical model for in vivo evaluation of human APP targeted nucleic acid drugs. Values are expressed as mean ± SEM. Significance was determined by unpaired t test. *P < 0.05, **P < 0.01, ***P < 0.001.