Nanoformulation of Talazoparib Increases Maximum Tolerated Doses in Combination With Temozolomide for Treatment of Ewing Sarcoma

Abstract

The Pediatric Preclinical Testing Program previously identified the PARP inhibitor talazoparib (TLZ) as a means to potentiate temozolomide (TMZ) activity for the treatment of Ewing sarcoma. However, the combination of TLZ and TMZ has been toxic in both preclinical and clinical testing, necessitating TMZ dose reduction to ~15% of the single agent maximum tolerated dose. We have synthesized a nanoparticle formulation of talazoparib (NanoTLZ) to be administered intravenously in an effort to modulate the toxicity profile of this combination treatment. Results in Ewing sarcoma xenograft models are presented to demonstrate the utility of this delivery method both alone and in combination with TMZ. NanoTLZ reduced gross toxicity and had a higher maximum tolerated dose than oral TLZ. The dose of TMZ did not have to be reduced when combined with NanoTLZ as was required when combined with oral TLZ. This indicated the NanoTLZ delivery system may be advantageous in decreasing the systemic toxicity associated with the combination of oral TLZ and TMZ.

Keywords: Ewing sarcoma; combination therapy; nanoparticle; talazoparib; temozolomide.

Figure 1

NanoTLZ is as efficacious at free TLZ in vitro. ES-6, ES-7, and EW-8 cell lines were treated with TLZ or NanoTLZ for 72 h and viability was assessed by Alamar Blue (n = 3/line). IC₅₀ plots for TLZ and NanoTLZ in ES-6, ES-7, and EW-8 cell lines (A). Statistical significance of TLZ and NanoTLZ IC₅₀ values were assessed for each cell line by Student's t-tests with α = 0.05 for significance; *p < 0.05 vs. NanoTLZ; **p < 0.01 vs. NanoTLZ.TC-71 cells were treated with the IC₁₀ of TLZ or NanoTLZ and dose response to TMZ was measured using Alamar Blue. Potentiation of TMZ activity in TC-71 cells combined with TLZ or NanoTLZ (B). Statistical significance of TMZ, TLZ, and NanoTLZ combination IC₅₀ values were assessed by one way ANOVA followed by Tukey's test for multiple comparisons; ***p < 0.001 vs. NanoTLZ; ****p < 0.0001 vs. TMZ; ^#p < 0.001 vs. TMZ + NanoTLZ. Mice harboring KT-10 xenografts were treated with 0.165 mg/kg TLZ BID x5 PO or 0.33 mg/kg NanoTLZ SID, IV, on days 1, 3, and 5. NanoTLZ and TLZ demonstrate inhibitory effect on total PARP1 protein levels in KT-10 tumor xenograft (n = 3/group) (C).

Figure 2

NanoTLZ preferentially accumulates within tumors. The size distribution of NanoTLZ as measured by DLS does not change with the addition of Cy5 dye (A). Live animal fluorescent imaging demonstrated NanoTLZ-Cy5 accumulates in the tumor 24 h after injection (n = 3) (B).

Figure 3

KT-10 xenografts exhibit a dose dependent response to TLZ. Animals bearing KT-10 xenografts were treated with either 0.1625 mg/kg (A), 0.33 mg/kg oral TLZ BID x 5 (B), or 1.0 mg/kg NanoTLZ (IV) SID x 5 (C) and relative tumor volume was plotted for each animal (n = 8–10/group). Percent change in weight during and after treatment (D).

Figure 4

NanoTLZ and TMZ are tolerable at higher doses than previously established with TLZ and TMZ. Tumor free mice (n = 3/group) were treated with 0.5 mg/kg NanoTLZ SID x5 in combination with increasing doses of TMZ and body weight was measured daily for 21 days (A). An alternative schedule assessed weight change during treatment with 50 mg/kg TMZ for 5 days and 1.0 mg/kg NanoTLZ on days 1, 3, and 5 (B).

Figure 5

TC-71 xenografts are responsive to the combination of NanoTLZ + TMZ. Mice bearing TC-71 xenografts (n = 10/group) were treated with 1 mg/kg vehicle or NanoTLZ (IV) on days 1, 3, and 5, 50 mg/kg TMZ (PO) on days 1–5, or the combination of the two and tumor volume was monitored twice weekly (A). Median relative tumor volume during 8 weeks of treatment (B). Change in body weight during and up to 30 days after treatment (C). Kaplan-Meier survival of TC-71 xenografts for 12 weeks after treatment initiation (D). Statistical significance assessed via the log-rank test followed by the Bonferroni correction for multiple comparisons, **p < 0.01 vs. NanoTLZ+TMZ.