Potentiation of the antitumor effect of Merocyanine 540-mediated photodynamic therapy by amifostine and amphotericin B

Abstract

Leukemia and lymphoma cells are much more sensitive to Merocyanine 540 (MC540)-mediated photodynamic therapy (PDT) than normal pluripotent hematopoietic stem cells and normal colony forming unit-granulocyte/macrophage progenitors (CFU-GM). By contrast, most solid tumor cells are only moderately sensitive to MC540-PDT. The limited activity against solid tumor cells has detracted from MC540's appeal as a broad-spectrum purging agent. We report here that noncytotoxic concentrations of amifostine (Ethyol, Ethiofos, WR-2721) and amphotericin B used either alone or in combination potentiate the MC540-sensitized photoinactivation of leukemia cells, wild-type small cell lung cancer cells and cisplatin-resistant small cell lung cancer cells. Amphotericin B also enhances the MC540-sensitized photoinactivation of normal CFU-GM, whereas amifostine protects CFU-GM against the cytotoxic action of MC540-PDT. The yield of CD34-positive normal hematopoietic stem and progenitor cells is only minimally diminished by pretreatment with amifostine, amphotericin B or combinations of amifostine plus amphotericin B. Purging protocols that combine MC540-PDT with amifostine or with amifostine plus amphotericin B could offer a simple and effective approach to the purging of autologous stem cell grafts that are contaminated with solid tumor cells or the purging of stem cell grafts from heavily pretreated leukemia patients that contain reduced numbers of normal stem and progenitor cells and, therefore, can ill afford additional losses caused by purging.

Figure 1.

Potentiating effects of amifostine, amphotericin B, and amifostine combined with amphotericin B on the the MC540-sensitized photoinactivation of wild-type H69 human small cell lung cancer cells (a), cisplatin-resistant H69/CDDP human small cell lung cancer cells (b), and normal human granulocyte/macrophage progenitors (c). Data represent mean colony counts of 4 culture dishes ± standard errors. Most error bars are smaller than the data symbols. While the survival of hCFU-GM was better in the presence of amifostine at all four fluences tested, only 2 data points (0 kJ/m², 146 kJ/m²) showed differences that were statistically significant with P-values of 0.008 and 0.02, respectively (unpaired t-test.) Untreated H69 cells, H69/CDDP cells, and human bone marrow cells (surviving fraction 1.0) formed 111.75, 172.25, and 163 colonies per plate, respectively.

Figure 2.

Potentiating effects of amifostine, amphotericin B, and amifostine combined with amphotericin B on the MC540-sensitized photoinactivation of murine L1210 leukemia cells (a) and normal murine granulocyte/macrophage progenitors (b). Data represent mean colony counts of 4 culture dishes ± standard errors. Most error bars are smaller than the data symbols. Cultures of untreated L1210 and normal murine bone marrow cells (surviving fraction 1.0) formed 197 and 83.5 colonies per plate, respectively.

Figure 3.

Effects of amifostine, amphotericin B, and amifostine combined with amphotericin B on the binding of MC540 to L1210 leukemia cells. Analysis was done by flow cytometer (a) or by fluorescence spectrophotometer after extraction with detergent (b). Data represent means of 2 determinations ± standard error.