Fig. 5: NCT-58 inhibits tumor growth in trastuzumab-resistant JIMT-1 xenografts.

A–C Effect of NCT-58 on tumor growth in vivo. JIMT-1 cells (3 × 10⁶) were injected into the mammary fat pads of BALB/c nude mice. Mice were administered intraperitoneally with NCT-58 (30 mg/kg·BW, every other day) or solvent control for 47 days (n = 6/each group). Tumor volumes were measured with a caliper at the intervals indicated. A, B NCT-58 administration resulted in significant decreases in tumor growth (A, ***p < 0.001, n = 6) and tumor weight (B, *p < 0.05, n = 6). C Changes in body weight of the xenografted mice after exposure to NCT-58 or control vehicle (NS; not significant, n = 6). D Representative histological analysis of lung, liver, and kidney sections stained with hematoxylin and eosin (H&E) and analyzed by microscope slide scanner. E Effects of NCT-58 on serum biochemical parameters of liver and kidney function. Blood biochemical analysis indicated there was no significant change in serum ALT, AST, BUN (NS; not significant). ALT, alanine aminotransferase; AST, aspartate aminotransferase; BUN, blood urea nitrogen. F Influence of NCT-58 on Ki-67 expression. Tumor tissue sections were immunostained for Ki-67 (red) and DAPI (blue). The graph shows the percentage of Ki-67-positive cells (***p < 0.001, n = 6). G NCT-58-induced apoptosis was determined by TUNEL assay. Extent of apoptosis expressed as the percentage of total TUNEL-positive cells (***p < 0.001, n = 6). H Effect of NCT-58 on tumor angiogenesis, as determined by a microvessel density (MVD) assay. Tumor tissues were immunostained with a specific endothelial marker CD31 (red) and DAPI (blue). The number of CD31-positive microvessels in the intratumoral and peritumoral areas were quantified, respectively (**p < 0.01; ***p < 0.001, n = 6). I, J Immunohistochemical analysis for the intracellular domain (ICD)-HER2 (green, I) and full-length HER2 (green, J) in vivo. Quantitative graphs of signal intensities shown in the right panels (***p < 0.001, n = 6). The results are presented as mean ± SD and data were analyzed by unpaired Student’s t-test and two-way ANOVA followed by Bonferroni’s post hoc test.