Phase 3 clinical trials in the process of new drug development are often heard even if they do not have a deep understanding. Of course, there is still a lot of preparation before entering a Phase 3 clinical trial, such as experimental data obtained in animal models. At this time, it is crucial for researchers to develop an animal model that understands the suitability of the study drug in the preclinical stage of the new drug.
The initial model of hepatitis B was developed, humanized NSG-PiZ mice, and the cost was less than one-tenth of the commercially available
First, the research background
At HEP DART 2021, researchers from the Department of Vaccines and Infectious Diseases at the Fred Hutchinson Cancer Research Center in Seattle, Washington, USA, brought advances in the development of a preclinical animal model. According to the researchers, a gene editing method for hepatitis B virus (HBV) treatment was previously studied, which uses adeno-associated virus (AAV) vector-mediated SaCas9 to deliver HBV cccDNA in mice targeting humanized FRGs in the liver.
However, in the past, the scientific community has found that if further research is needed, it will be limited by the cost of humanized FRG mice. In this study, the researchers demonstrated innovative ideas for HBVDNA gene editing, developing a new humanized NSG-PiZ mouse model.
2. Research methods
The researchers built a mouse model of humanized NSG-PiZ of the liver in the lab and assessed its permissibility for HBV replication. NSG-PiZ mice are easily humanized and only need to be pretreated by injection of liver poison before intrasplenic human liver cell transplantation, without the need for mass rearing. Notably, the human cost of NSG-PiZ mice is less than 1/10 of that of commercially available humanized uPA-SCID and FRG mice, making it affordable for academic researchers.
The researchers transplanted human liver cells to NSG-PiZ mice and obtained implant levels comparable to uPA-SCID and FRG mice, typically reaching hAlbumin levels of 1-10 mg/mL within ten weeks. Importantly, humanized NSG-PiZ mice are fully tolerated for HBV infection. NSG-PiZ mice attacked with genotype D clinical HBV isolates have been tracked for 24 weeks, and infectious viruses can be passed sequentially through mice. Viremia levels correlate with hAlbumin levels and peak above 109 IU/mL.
HBSAg secretion in HBV+ mice peaks at 10 weeks and expression of HBsAg in hCK18+ human liver cells indicates that species-specific replication occurs in the humanized liver. At autopsy, hRPP30 and mRRP30 and liver-associated total HBV DNA and cccDNA are used to quantify liver chimerism by ddPCR. A set of experiments was then conducted to demonstrate the suitability of the NSG-PiZ model for studies of HBV treatment.
First, the effect of RTi entecavir (ETV) on the HBV replication +/- capsid assembly inhibitor ciclopiroxone (CPX) was studied to determine efficacy and determine whether combination therapy provided additional antiviral effects. Viral load-PiZ mice are tracked in viral NSGs for six weeks, including four weeks of ETV or ETV/CPX therapy, followed by two weeks of no-drug therapy to determine the effects of single or combination drug therapy. Mice treated with ETV reduced viral load by approximately 3 log-stages over 4 weeks and rebounded after ETV discontinuation. This level of inhibition is comparable to that observed in HBV+ uPA-SCID or FRG mice.
The combination of ETV/CPX had no additional effect on mice treated with ETV alone. Finally, it was determined whether the AAV vector could efficiently transduce HBV+ human hepatocytes in NSG-PiZ mice. HBV+ NSG-PiZ mice were administered GFP-expressing hepatophilic AAV3B or AAV. LK03 vector and analyze gene transfer in chimeric liver after 28 days. Both vectors transduced human and mouse hepatocytes, as well as HBV+ human hepatocytes.
3. Conclusions of the study
In summary, the researchers concluded that the humanized NSG-PiZ mouse model of the liver supports HBV replication and can be used to monitor the efficacy of antiviral small molecule therapies. They also seem well suited to study AAV-mediated antiviral therapies against HBV cccDNA.
Xiaofan Health Conclusion: Let's briefly comb through this humanized NSG-PiZ mouse model presented by Fred Hutchinson researchers at HEP DART 2021, which supports global research on the antiviral efficacy of hepatitis B. Researchers had used gene editing technology to target the cccDNA of hepatic humanized FRG mice in animal experiments in the early days, because the development cost of humanized FRG mice limited further research.
To make academic researchers economically affordable, Researcher Fred Hutchinson developed the NSG-PiZ mouse model, which costs only one-tenth the cost of the humanized uPA-SCID and FRG mouse models on the market. The significance of this animal model is that it can be used to evaluate the effectiveness of new drugs for chronic hepatitis B in the early small molecule stage, and it is also applicable to new antiviral therapies with the potential to target cccDNA.