Figure 4. Reduced LCP1 expression in the early stages of metastasis decreases lung metastasis formation and is correlated with lower tumor grade and preferable intrinsic tumor subtype.

(A) Photographs of primary breast tumors of scrambled, Lcp1, Abce1 KD and Abce1 KD + Lcp1 KD 4T1 cells that were removed on day 14 post-injection. Ruler pictured in metric units. (B) Tumor weights were measured as they were harvested from the mice. Abce1 KD + Lcp1 KD tumors weighed significantly less than tumors of other treatment groups. (C) Western blot analysis of protein extracted from breast tumors derived from scrambled, Lcp1 KD, Abce1 KD, and Abce1 KD + Lcp1 KD 4T1 cells revealed reduced protein expression of Abce1 in Abce1 KD and dual Abce1+Lcp1 KD groups compared to the scrambled group, while the Lcp1 protein expression level was reduced in the Lcp1 KD and the dual Abce1+ Lcp1 KD groups compared to the scrambled group. (D) Lung microCT scans performed on day 28 (orthotopic) and day 21 (IV) post-4T1 injection and (E) quantification of LMets in microCT show significantly fewer metastatic growths (indicated by red arrows) in mice that orthotopically received in mice that orthotopically received Abce1 KD, Lcp1 KD or Abce1 KD + Lcp1 KD cells compared to the scrambled control. Injection of Abce1 KD+Lcp1 KD cells resulted in the most prominent reduction of orthotopic LMets compared to knockdown of Abce1 or Lcp1 alone. IV injection of Abce1 KD, Lcp1 KD or Abce1 KD + Lcp1 KD cells did not result in fewer lung foci compared to scrambled. Scale bars represent 1 mm. (F) Kaplan–Meier survival analysis demonstrated increased overall survival of mice that received Abce1 KD + Lcp1 KD compared to the scrambled control. Data are presented as mean ± SEM. ^*p < 0.05.