Fig. 5. Combination treatment with neratinib and dinaciclib leads to anti-tumor activity in vivo.

A Approximately, 1.5 × 10⁶ (1.5 million) cells derived from a HER2-positive breast cancer PDX model (WHIM 22) were injected orthotopically into each NSG mouse (both sides) and monitored for subsequent growth. After tumors reached a size of ~150 mm³, mice were treated with 40 mg/kg neratinib 5 days a week (Monday–Friday), 40 mg/kg dinaciclib twice a week, or their combination for 16 days. Tumor measurements were performed every day by calipers, and the percentage (%) of changes in volume for each tumor is shown by a waterfall plot (control = 4 tumors, neratinib = 4 tumors, dinaciclib = 4 tumors, combination = 4 tumors). For statistical analysis one-way Anova test was performed for comparisons between neratinib, dinaciclib, and combination cohorts. Dunnett’s test was used as post hoc. Differences were considered statistically different if p < 0.05. A p value < 0.05 is indicated by *, p < 0.01 by **, p < 0.001 by ***, and p < 0.0001 by ****. B Weights of the WHIM 22 PDX model-bearing mice of the single agents and the combination cohorts. The number of mice was: control = 2 mice, neratinib = 2 mice, dinaciclib = 2 mice, and combination = 3 mice. p Values were calculated using the two-tailed Student’s t test. C Same as A using the WHIM 8, HER2-positive breast cancer PDX model (18 days of treatment, control = 5 tumors, neratinib = 5 tumors, dinaciclib = 4 tumors, combination = 3 tumors). D Same as B using the WHIM 8 PDX model. The number of mice was: control = 5 mice, neratinib = 5 mice, dinaciclib = 2 mice, and combination = 3 mice. E Tumors were harvested from WHIM 8 PDX tumor-bearing mice approximately 2 h after the last drug administration and tumor lysates were subjected to western blot analyses and probed for the indicated proteins.