Figure 8.

Osteosarcoma tumors treated with ATG4B antagonist NSC185058 fail to grow. (A) Saos-2 cells were incubated in nutrient-rich (closed circles) or amino acid-deprived (open triangles) medium in the presence of NSC185058 (3 to 200 μM). After 48 h, cell viability was quantified by MTT-based assays. The values represent the mean ± SEM (n = 6). The statistical differences between comparable concentrations of NSC185058 are indicated. ***P < 0.001 (B) Immunodeficient nu/nu female mice were injected subcutaneously with 6 × 10⁶ Saos-2 (GFP-LC3B) cells. At 7 d, the mice were divided into 2 groups of 5 mice each and injected IP on Monday, Wednesday, and Friday with either peanut oil vehicle or NSC185058 (100 mg/kg body weight) in peanut oil. This is a dosage that we have shown is sufficient to suppress liver autophagy in the mouse. The values represent the mean ± SEM (n = 4) of tumor volumes (cm³). (C) A second set of immunodeficient nu/nu female mice was injected subcutaneously with 6 × 10⁶ Saos-2 (GFP-LC3B) cells. Palpable tumors were detected at 12 d, and the mice were divided into 2 groups and injected IP on Monday, Wednesday, and Friday with either peanut oil vehicle (n = 8) or NSC185058 (100 mg/kg body weight) dissolved in peanut oil (n = 9). At 45 d, a crossover was done. The vehicle treated group was subjected to NSC185058 (open diamonds), while the NSC185058 treated group was switched to vehicle alone (open circles). NSC185058 significantly inhibited tumor growth, but the changes in tumor size after crossover failed to reach statistical significance. The values represent the mean ± SEM (n = 8 to 10) of tumor volumes (cm³ × 10⁻¹). The statistical differences between comparable time points are indicated. *P < 0.05; **P < 0.01; ***P < 0.001