Figure 3. Lunasin reduced self-renewal capacity and oncosphere formation of CICs.

ALDH^high populations of melanoma cells were plated in low adherent culture for 21 d and treated with vehicle or Lunasin twice weekly. Representative images taken at 7 d (A) and 21 d (B) illustrate the ability of Lunasin to inhibit sphere formation of A375 ALDH^high melanoma cells. Lunasin treatment decreased sphere formation by 81% and 55% in A375 ALDH^high and SKMEL-28 ALDH^high cell lines, respectively (C). ALDH^high melanoma cells showed increased sensitivity to Lunasin versus their parental counterparts when treated in soft agar. Lunasin-treated ALDH^high cells derived from A375 and SKMEL-28 lines exhibited a decreased ability to form colonies in soft agar. Representative images for A375 and SKMEL-28 colonies treated with vehicle (D, F) or Lunasin (E, G) illustrate the morphological differences between treatment groups. Both the number and size of colonies formed by Lunasin-treated cells were decreased in size and density in ALDH^high fractions of A375 (H) and SKMEL-28 (I) cell lines. Significance (p < 0.05) was determined from three independent experiments and assessed by student's t-test. White scale bars shown on images represent 100 μm. All representative images were taken at 40× magnification. Error bars displayed on graphs represent mean ± S.D.