4-1BB is a member of the TNF-receptor superfamily. This receptor contributes to the clonal expansion, survival, and development of T cells. It can also induce proliferation in peripheral monocytes, enhance T cell apoptosis induced by TCR/CD3 triggered activation, and regulate CD28 co-stimulation to promote Th1 cell responses. The expression of this receptor is induced by lymphocyte activation. TRAF adaptor proteins have been shown to bind to this receptor and transduce the signals leading to activation of NF-kappaB. 4-1BB-L is a cytokine that belongs to the tumor necrosis factor (TNF) ligand family. This transmembrane cytokine is a bidirectional signal transducer that acts as a ligand for TNFRSF9/4-1BB, which is a costimulatory receptor molecule in T lymphocytes. This cytokine and its receptor are involved in the antigen presentation process and in the generation of cytotoxic T cells. The receptor TNFRSF9/4-1BB is absent from resting T lymphocytes but rapidly expressed upon antigenic stimulation. The ligand encoded by this gene, TNFSF9/4-1BBL, has been shown to reactivate anergic T lymphocytes in addition to promoting T lymphocyte proliferation. This cytokine has also been shown to be required for the optimal CD8 responses in CD8 T cells. This cytokine is expressed in carcinoma cell lines, and is thought to be involved in T cell-tumor cell interaction.
Hepatotoxicity evaluation-urelumab(in house)
Day 21, collect serum and detect ALT and AST. The results as followed:
a) In B-h4-1BB mice, compared with the control group, ALT was significantly increased by 20mg/kg Urelumab treatment, but there was no significant change in AST.
b) In B-h4-1BB/h4-1BBL mice, compared with the control group, AST was significantly increased by 20mg/kg Urelumab treatment.
c) Both in B-h4-1BB mice and B-h4-1BB/h4-1BBL mice, there was no significant change in ALT and AST in 1mg/kg dose group.
d) 20mg/kg Urelumab(in house) have no significant effect in ALT between the two mice. But, AST was significantly increased in B-h4-1BB/h4-1BBL mice.
Hepatotoxicity evaluation-Urelumab(in house)
Summary table of liver lesions and severity
Evaluation Criteria：Hepatic perivascular cell infiltration or chronic inflammation. sight（+）：Lesion site accounts for about 5% of the total area mild（++）：Lesion site accounts for about 5~20% of the total area moderate（+++）：Lesion site accounts for about 20~40% of the total area severe（++++）：Lesion site accounts for about 40% of the total area Normal：NVL
In B-h4-1BB Mice (G1-G3), no obvious abnormal changes were observed in the liver when the urelumab dose was 1mg/kg (G2). At a dose of 20 mg/kg (G3) changes were observed in 5/5 of the animals, manifested as perivascular cell infiltration or chronic inflammation in the liver, with mild lesions. In B-h4-1BB /4-1BBL mice (G4-G6), 1 mg/kg group(G5) 3/5 of mice show pathological changes(slight 2/5, mild 1/5), but in 20 mg/kg dose (G6), all of the experimental animals could see moderate changes in liver(5/5) . Overall, the degree and incidence of liver lesions in the 20 mg/kg group (G6) were significantly higher than that in the 1 mg/kg group (G5). The above results suggested that urelumab at 20 mg/kg was more likely to perivascular cell infiltration or chronic inflammation in the liver than that at 1 mg/kg. B-h4-1BB/h4-1BBL mice were more sensitive to urelumab toxicity effects than B-h4-1BBL mice. B-h4-1BB/h4-1BBL mice was a good preclinical toxicity evaluation model.
In vivo efficacy of anti-4-1BB antibodies
Antitumor activity of anti-human 4-1BB antibodies in B-h4-1BB/h4-1BBL mice. (A) Anti-human 4-1BB antibodies urelumab (in house) inhibited MC38 tumor growth in B-h4-1BB/h4-1BBL mice. Murine colon cancer MC38 cells were subcutaneously implanted into homozygous B-h4-1BB/h4-1BBL mice (female, 6-7 week-old, n=5). (B) Body weight changes during treatment. As shown in panel A, anti-human 4-1BB antibodies were efficacious in controlling tumor growth in B-h4-1BB/h4-1BBL mice, demonstrating that the B-h4-1BB/h4-1BBL mice provide a powerful preclinical model for in vivo evaluation of anti-human 4-1BB antibodies. Values are expressed as mean ± SEM.