Fig. 3.

Improved efficacy of metronomic administration of Doxil combined with immunotherapy. (A) Experimental protocol of the in vivo studies in 4T1 and MCA205 tumors. When tumors reached an average volume of 150 mm³ for 4T1 and 200 mm³ for MCA205, shear wave elastography (SWE) and dynamic contrast-enhanced ultrasound (DCEUS) were performed prior to initiation of Doxil treatment. Three dose schedules were employed for Doxil: i) 1 mg/kg daily for six days, ii) 2 mg/kg every other day and iii) 6 mg/kg once a week. After completion of the first cycle of treatment (Day 18), SWE and DCEUS were repeated. During the second cycle of treatment an immune checkpoint inhibitor (ICI), aPDL1 (10 mg/kg), was added to the treatment protocol and was administered on days 18, 21 and 24. SWE and DCEUS were repeated prior tumor excision (Day 25). Tumor volume data for (B) 4T1 and (C) MCA205 tumors. Low and more frequent doses of Doxil (1 mg/kg and 2 mg/kg) were able to decrease tumor growth rate for the first treatment cycle and improve the efficacy of nano-immunotherapy in the second treatment cycle. Statistical analyses were performed by comparing the Doxil 6 mg/kg group with the control and the aPDL1 groups * and the Doxil 1 mg/kg and Doxil 2 mg/kg with all other treatment groups **, p ≤ 0.05, determined by t-test. Data are presented as mean ± SEM (n = 6 mice per group).