• DLL3 is a highly tumor-selective cell surface target predominantly expressed in neural or neuroendocrine tumors, including small cell lung cancer (SCLC) and pleomorphic gliomas, but absent in normal lung tissues or adjacent non-cancerous tissues. DLL3 modulates Notch signaling by interacting with Notch receptors: it binds to DLL1 and the Notch1 receptor, thereby modulating and promoting Notch1 receptor degradation to suppress Notch signaling activation and upregulation. DLL3 expression is directly regulated by the transcription factor ASCL1 (a neuroendocrine lineage transcription factor), which is critical for neuroendocrine cell development and also acts as an oncogenic driver.
• In B-NDG hDLL3 mice, mouse Dll3 exons 2-7 (encoding the extracellular domain) were replaced with human orthologs. Human DLL3 mRNA was exclusively detected in homozygous B-NDG hDLL3 mice, not in wild-type B-NDG mice. DLL3 protein was detected in the brain, lung, kidney, liver, and testes of both B-NDG and homozygous B-NDG hDLL3 mice, likely due to antibody cross-reactivity between mouse and human DLL3.
• Application: This mouse model is suitable for pharmacodynamic and safety evaluations of human DLL3-targeted therapeutics.

Gene targeting strategy for B-NDG hDLL3 mice. The exons 2-7 of mouse Dll3 gene that encode extracellular domain were replaced by human counterparts in B-NDG hDLL3 mice. The genomic region of mouse Dll3 gene that encodes the transmembrane domain and cytoplasmic portion is retained. The promoter, 5’UTR and 3’UTR region of the mouse gene were also retained. The chimeric DLL3 expression was driven by endogenous mouse Dll3 promoter, while mouse Dll3 gene transcription and translation will be disrupted.

Strain specific DLL3 expression analysis in homozygous B-NDG hDLL3 mice by western blot. Various tissue lysates were collected from B-NDG mice (+/+) and homozygous B-NDG hDLL3 mice (H/H), and analyzed by western blot with anti-DLL3 antibody (CST, 2483s). m/hDLL3 showed high expression in the testis and kidneys, while lower protein expression was detected in the brain, lung, and liver of B-NDG mice and homozygous B-NDG hDLL3 mice.

Species specific analysis of DLL3 gene expression in B-NDG mice and homozygous humanized B-NDG hDLL3 mice by RT-PCR. Brain was collected from B-NDG mice (+/+) and homozygous B-NDG hDLL3 mice (H/H). Mouse Dll3 mRNA was detectable only in B-NDG mice. Human DLL3 mRNA was detectable only in homozygous B-NDG hDLL3 mice, but not in B-NDG mice.