Table 1.
Human cytokine antibody array kit (RayBiotech, USA) contending 20 different growth factors/cytokines were used for analysis of protein expression profile in tumor tissue and cells lysate.
Figure 1.
HET0016 treatment reduced tumor growth in breast cancer nude mice.
A) HET0016 treatment decreased tumor volume in both the early and delayed treatment groups. Significant difference was observed between control and treatment groups as early as 7 days of treatment. When compared the tumor growth rates (B), animal that received delayed treatment showed rapid growth at late stage of treatment. C) Representative samples of mammary tumors developed by MDA-MB-231 cells implantation on the right flank of mice. **p<0.001, ***p<0.0001.
Figure 2.
SPECT analysis of in vivo accumulation of Tc-99m-HYNIC-VEGF-c.
One hour after injection, SPECT images were obtained using dedicated animal scanner. A) Semi-quantitative analysis of total radioactivity in the tumors normalized to corresponding control tumors showing the intensity of radioactivity in the vehicle and HET0016 treated groups. B) Images of vehicle treated mice showed increased accumulation of Tc-99m-HYNIC-VEGF-c compared to that HET0016 treatment groups (early and delayed). Intersection of lines indicates the tumor. Error bars: ± standard error, **p = 0.0131.
Figure 3.
Immunohistochemistry stained with vWF in vehicle and HET0016 treated tumors groups.
A) Quantitative estimation of Mean Vascular Density. B) Top images were taken from tumor of 21 days and bottom images were taken from tumor of 28 days. The arrows shows micro-vessels. Error bars: ± standard error, **p<0.001. Scale bar = 100 µm.
Figure 4.
Immunohistochemistry stained with Ki-67 in vehicle and HET0016 treated tumors groups.
A) Quantitative estimation of cell proliferation showing number of cells per area (m2). B) Top images were taken from tumor of 21 days and bottom images were taken from tumor of 28 days. The arrows shows cellular nucleus stained. Error bars: ± standard error, **p<0.001. Scale bar = 100 µm.
Figure 5.
Quantitative estimation of growth factors/cytokines in protein extracted from (A) 21 and (B) 28 days of breast tumor using a custom designed membrane based protein array.
Error bars: ± standard error, *p<0.05, **p<0.001, ***p<0.0001.
Figure 6.
Quantitative estimation of growth factors in protein from cell lysate and supernatant using a custom designed membrane based protein array kit.
Cell lysed (A and B); supernatant (C and D). Collected after 4 h of treatment 10 µm of HET0016 (A and C) and 24 h (B and D). Error bars: ± standard error. *p<0.05.
Figure 7.
Determination of (A) HIF-1 and (B) MMP-2.
No significant differences were observed among the treatment groups.