Paclitaxel in tyrosine-derived nanospheres as a potential anti-cancer agent: in vivo evaluation of toxicity and efficacy in comparison with paclitaxel in Cremophor

Abstract

Paclitaxel (PTX) has gained widespread clinical use yet its administration is associated with significant toxicity. In the present study, the toxicity and anti-tumor efficacy of tyrosine-derived nanospheres (NSP) for the delivery of PTX was compared to a clinical formulation of PTX in PBS-diluted Cremophor® EL (PTX-CrEL-D). Maximum tolerated dose was determined using a concentration series of PTX in NSP and CrEL-D, with toxicity assessed by measuring changes in body weight. Healthy mice administered PTX-NSP continued to gain weight normally while treatment with PTX-CrEL-D resulted in significant weight loss that failed to recover following treatment. Even at the dose of 50mg/kg, PTX-NSP showed better tolerance than 25mg/kg of PTX-CrEL-D. Xenograft studies of breast cancer revealed that the anti-tumor efficacy of PTX-NSP was equal to that of PTX-CrEL-D in tumors originating from both MDA-MB-435 and ZR-75-1 cancer lines. Larger volume of distribution and longer half-life were measured for PTX-NSP administration compared to those reported in the literature for a CrEL formulation. This trend suggests the potential for improved therapeutic index of PTX when administered via NSP. The findings reported here confirm that the NSP formulation is an efficient method for PTX administration with significant increase in maximum tolerated dose, offering possible clinical implications in the treatment of breast tumors.

Fig. 1

Viability of MDA-MB-435 breast cancer cells exposed to 0.06 and 0.6 ng/mL of PTX delivered via NSP (2 and 6 ng/mL), CrEL-D (8 and 24 ng/mL), and drug free formulations of NSP (2 and 6 ng/mL) and CrEL-D (8 and 24 ng/mL). Cell viability was measured after 72 h by AlamarBlue^® cell proliferation indicator assay. Cells incubated with PBS and media were used as negative control. The results are expressed as mean value with its standard error indicated (mean ± SE) of 6 measurements of 3 independent experiments; p < 0.05 (*), 0.01 (**), and 0.001 (***) were considered to be statistically significant.

Fig. 2. In vivo

toxicity of NSP and CrEL-D as a function of treatment schedule in healthy NCr nu/nu mice (N = 6 for each treatment). Treatments (0.2 mL, IV) were administered on days 1, 5, 9, and 13 (arrows, q5dx4). Concentration of NSP and CrEL in each treatment was 13 and 52 mg/0.2 mL, respectively. Each data represents the mean value with standard error (mean ± SE); p < 0.05 (*), 0.01 (**), and 0.001 (***) were considered to be statistically significant in the body weight change on the selected times comparing each group to pre-treatment weight.

Fig. 3

Determination of toxicity/MTD for PTX delivered via NSP and CrEL-D. Mice (N = 6 per group) were injected (0.2 mL) on days 1, 5, 9, and 13 with (A) 5, (B) 10, (C) 20, (D) 25, (E) 30 mg/kg of PTX in each vehicle: ( ) PTX-NSP; ( ) PTX-CrEL-D. (F) 40 ( ) and 50 ( ) mg/kg of PTX in NSP. Concentration of NSP and CrEL-D in each treatment was 13 and 52 mg/0.2 mL, respectively. Lethal toxicity (5-10% mortality) was observed in all PTX-CrEL-D treatments, regardless of PTX dose, by the time of the third treatment. The statistical data is expressed as the mean value ± S.E. (standard error); p < 0.05 (*), 0.01 (**), and 0.001 (***) were considered to be statistically significant in the body weight change compared to the initial pre-treatment weight.

Fig. 4

Anti-tumor activity in NCR nu/nu mice bearing subcutaneous (A) MDA-MB-435 estrogen-independent (ER−) and (B) ZR-75-1 estrogen dependent (ER+) breast cancer xenografts. Mice (N = 6 per group) were treated (0.2 mL) on days 1, 5, 9, and 13 with 20 mg/kg of PTX in NSP (13 mg) or CrEL-D (52 mg) in both studies. The control groups received PBS (MDA-MB-435 study, N = 6) and NSP and CrEL-D (ZR-75-1 study, N = 6) using the same administration schedule. Lethal toxicity (10–15% mortality) was observed in all PTX–CrEL-D treatments by the time of the third treatment. The data is presented as a mean value with standard error (mean ± SE) of tumor size in each tested group with no statistical difference seen between both treatments.

Fig. 5

Viability of MDA-MB-435 and ZR-75-1 breast cancer cells exposed for 72 h to PTX–NSP and corresponding amounts of NSP alone. Cell viability was measured by AlamarBlue^® cell proliferation indicator assay. Cells incubated with PBS and media were used as negative control and treatments data was normalized to these values. The results are expressed as mean value with its standard error (mean ± SD) indicated for 5 measurements of 3 independent experiments.

Scheme 1

Chemical structure of PEG5K-b-oligo(desaminotyrosyl-tyrosine octyl ester suberate)-b-PEG5K triblock copolymer, DTO–SA/5K. The oligo B-block is distinguished by both their alkyl pendent chain “octyl” linked to the (desaminotyrosyl-tyrosine alkyl ester) unit and the SA “suberic acid” to form the oligomer of (DTO–SA). mPEG A-blocks are abbreviated as 5K, indicating the molecular weight of the PEG components.