Fig. 2. In vivo efficacy of IMM0306.

A The cell inoculation method was used to create human leukemia cell Daudi CB17-SCID mice xenograft subcutaneous tumor models: tumor cells were collected in logarithmic growth stages and subcutaneously inoculated into SCID mice with 2 × 106/0.1 ml/mouse cell suspension in 1xPBS with Matrigel at a 1:1 ratio at a cell concentration of 2 × 10⁷/ml. When the tumor volumes reached 100–200 mm³, the animals were randomly divided into different groups with the tumor difference between each group being less than 10% of the mean value. Different agents were administered to the different groups of mice. The tumor volumes and the body weight of the animals were measured three times a week, and the clinical symptoms were observed and recorded once a day. The results revealed that IMM0306 exerts excellent cancer killing efficacy by activating both macrophages and NK cells via blockade of CD47-SIRPα interaction and FcɣR engagement. After administration of IMM0306 at 1.5 mg/kg for 3 weeks, 100% of the SCID mice in the Daudi xenograft model achieved complete remission (CR). B CB17-SCID mice xenograft orthotropic Raji tumor model was established by inoculating 0.2 ml cell suspension at the concentration of 5 × 10⁶/mouse by caudal vein. Three days after inoculation, animals were randomly divided into different groups with the body weight difference between each group being less than 10% of the mean value. Starting on day 0, the drugs were administered according to the weight of the animals. During the administration, if the weight of individual animals decreases by more than 15% compared to day 0 (BWL ≧15%), the drug will be stopped until the weight of animals recovers (BWL ≦ 15%). During the experiment, tumor volumes and the body weight of the animals were measured three times a week, and the clinical symptoms were observed and recorded once a day, and the TGI values were calculated. The results showed that IMM0306 is more potent than rituximab and the combo of two single agents in tumor-inhibition, with a TGI of 92.7% with IMM0306 versus 27.6% with rituximab, respectively. C Raji-luc inoculated CB17-SCID mice in situ to evaluate the therapeutic effect of IMM0306, rituximab, lenalidomide, and their combinations. CB17 SCID mice were inoculated with Raji-luc human lymphoma cells via tail vein injection. On the 14th day after inoculation, the mice were intraperitoneally injected with the fluorescent substrate D-Luciferin. About 15 min later, the mice were anesthetized with 3–4% isoflurane, and the fluorescence intensity of the mice was detected with a live imaging system (PerkinElmer, Lumina XRMS). During the period, the weight of the mice was monitored, and the clinical symptoms were recorded. The mice were imaged in vivo on days 7, 12 and 17 after grouping to detect the tumor fluorescence intensity. The data on the 12th day of in vivo imaging showed that the tumor fluorescence intensity of mice in the solvent group (non-drug treatment group) was significantly increased after tumor cells were inoculated, and the tumor fluorescence intensity of mice in the drug treatment group was weaker than that in the solvent group, showing a certain therapeutic effect. The tumor fluorescence intensity of IMM0306 combined with lenalidomide was the lowest, which was significantly better than that of any single drug group of IMM0306 and lenalidomide, which was consistent with the weight and clinical symptom data of mice. The data showed that the three drugs all showed a certain degree of anti-tumor effect, but the therapeutic effect of IMM0306 combined with lenalidomide was significantly better than that of any single drug, and better than that of rituximab combined with lenalidomide.