Polycationic calixarene PTX013, a potent cytotoxic agent against tumors and drug resistant cancer

Abstract

Previously, we reported on the anti-tumor activities of two designed calix[4]arene-based topomimetics (PTX008 and PTX009) of the amphipathic, angiostatic peptide Anginex. Here, we chemically modified the hydrophobic and hydrophilic faces of PTX008 and PTX009, and discovered new calixarene compounds that are more potent, cytotoxic anti-tumor agents. One of them, PTX013, is particularly effective at inhibiting the growth of several human cancer cell lines, as well as drug resistant cancer cells. Mechanistically, PTX013 induces cell cycle arrest in sub-G1 and G0/G1 phases of e.g. SQ20B cells, a radio-resistant human head and neck carcinoma model. In the syngeneic B16F10 melanoma tumor mouse model, PTX013 (0.5 mg/Kg) inhibits tumor growth by about 50-fold better than parent PTX008. A preliminary pharmacodynamics study strongly suggests that PTX013 exhibits good in vivo exposure and a relatively long half-life. Overall, this research contributes to the discovery of novel therapeutics as potentially useful agents against cancer in the clinic.

Fig. 1

Structural representations of calix[4]arene diguanidine compound PTX009 and tetra-amine compounds PTX008, PTX012, PTX013, PTX014, and PTX015

Fig. 2

Cytotoxicity effects of calix[4]arenes. Cell viability of HUVEC, fibroblasts, and human ovarian carcinoma (MA148) and lung carcinoma (A549) cells, as well as murine endothelial (2H11), fibrosarcoma (FSAII), mammary carcinoma (SCK), and melanoma (B16F10) cells, was measured in the absence or presence of various concentrations of calixarenes. Cell viability and survival was assessed by colorimetric analysis as described in the “Experimental”

Fig. 3

Cytotoxicity effects of PTX013 on human cancer cell lines and corresponding resistant counterparts. a Human head and neck carcinoma (SQ20B), colon adenocarcinoma (Colo-205), breast adenocarcinoma (MCF-7), and colon adenocarcinoma (DLD-1) and their respective resistant variations (please see “Material and Methods” for details) were exposed to various concentrations of PTX013 for 72 h. A dose dependent inhibition of cancer cell survival was observed (with an overall approximate IC₅₀ ~3 µM), as assessed by colorimetric analysis (“Material and Methods”). b Cell cycle analysis was performed in SQ20B cells using PTX013 at 3 μM. Exposure to PTX013 increased the number of cells in sub-G1 and GO/G1 phases of the cell cycle and was associated with decreased numbers of cells in S and G2/M phases

Fig. 4

Tumor growth inhibition by calixarenes in the B16F10 melanoma model. a B16F10 tumor growth inhibition by calixarenes PTX008, PTX009, PTX013, and PTX015 (10 mg/kg BID IP for 7 days). b Body weights of the mice during PTX008, PTX009, PTX013, and PTX015 (10 mg/kg BID IP for 7 days) treatment. Data points represent means±SEM (n=10 each group)

Fig. 5

Dose-dependent B16F10 tumor growth inhibition by PTX013 treatment. a Dose-dependent B16F10 tumor growth inhibition by 1.5, 5, and 10 mg/kg (BID IP for 7 days) treatment of PTX013. b Body weights of the mice during PTX013 treatment (1.5, 5, and 10 mg/kg BID IP for 7 days). c Dose-dependent B16F10 tumor growth inhibition by 0.1, 0.2, and 0.5 mg/kg (BID IP for 7 days) PTX013 treatment. d Body weights of the mice during PTX013 treatment (0.1, 0.2, and 0.5 mg/kg BID IP for 7 days). Data points represent means±SEM (n= 10–20 each group)

Fig. 6

Tumor growth inhibition by PTX013 in the B16F10 melanoma model using different treatment schedules. a The effect of different scheduling of PTX013 while maintaining the same accumulative total treatment dose. All groups received a total of 20 mg/kg PTX013 at the end of the treatment span of 10 days. However, the groups were divided to receive either 10 injections of 2 mg/kg every day (q1dx10), 4 injections of 5 mg/kg every 3 days (q3dx4), or 2 injections of 10 mg/kg every 5 days (q5dx2). b Body weights of the mice during different treatment schedules of PTX013. Data points represent means ±SEM (n=10 each group)