Figure 7

An antibody targeting cancer-specific periostin variants inhibited mesenchymal cell expansion and tumor invasion after chemotherapy and halted tumor progression. (A) Diagram of splicing variants of human and mouse POSTN. Arrows indicate the location of designed primers. (B) Quantitative PCR analysis of POSTN splicing variants in tumor with or without PTX treatment. The region over exon 20 and 21 (hPNex20-21) was also amplified. Relative values against human total POSTN expression in vehicle-treated tumor are presented. Relative values are presented as mean ± standard error, *P < 0.02, Student’s t test. (C) Immunofluorescence staining of periostin variants containing exon 21 in tumor upon PTX treatment and periostin knockdown. Scale bar, 100 µm. (D) Tumor growth curve in MCF10DCIS xenografts treated with PTX and/or PN21-Ab. Upper arrows indicate times of PTX administration, and lower arrows indicate times of PN21-Ab administration. Tumor volumes were measured on the indicated days and are presented as mean ± standard error (n = 10). **P < 0.01, Student’s t test. (E) Flow cytometry analysis of tumors at Day 24 using CD44 and CD24 antibodies. Samples were prepared from a mixture of 10 tumors. Ratio of CD44^high/CD24^low to CD24^high tumor cells is presented on the plots. (F) The ratio of CD44^high/Ki67⁺ cancer cells against total Ki67⁺ cells was quantified in tumors collected at Day 24 (n = 15). Bars represent mean, and error bars represent standard deviation. ****P < 0.0001, Mann-Whitney’s test. (G) Immunofluorescence staining of CD44 and Ki67 in tumors collected at Day 24. Scale bar, 100 µm. (H) Immunofluorescence staining of laminin-5 and periostin in tumors collected at Day 24. Scale bar, 100 µm.