Single-drug concentration-response curves are assessed. Seeding is done at a density of 2,000 cells/well for PC-3 and LNCaP, in 96-well plates. Cells are treated with each single drug and their combination for 72 h at different drug concentrations. Docetaxel is used at concentrations of 0.1-1,000 nM. GLU is used at concentrations of 0.1-300 μm. Cytotoxicity is assessed at the end of drug exposure using SRB assay. Following 72 h exposure the cells are fixed with 10% trichloroacetic acid (150 μL) for 1 h at 4°C. Then, cells are stained for 10 min at room temperature with 0.4% SRB dissolved in 1% acetic acid. The plates are then air dried for 24 h and the dye is made soluble with 150 μL Tris (10 mM, PH 7.4) for 5 min on a shaker at 1,600 rpm. Absorbance is then measured at 545 nM using microplate reader. Results are expressed as the relative percentage of absorbance compared to control^[1].

MCE has not independently confirmed the accuracy of these methods. They are for reference only.

Mice^[3]
Five-week-old male Balb/c mice are used. Docetaxel (0, 10, 20, 30, 40, 60, and 80 mg/kg per week) is given once a week for 3 weeks for mice. Because more than 30 mg/kg per week of Docetaxel causes body weight loss in mice, 20 mg/kg per week of Docetaxel is judged to be the maximum nontoxic dose. Docetaxel (20 mg/kg per week) is given to mice once a week for 3 weeks at one of the following different points (2, 10, 14, or 22 HALO). Seventy-two hours after the final dosing of the agent, the intestinal mucosa of the small intestine (proximal 8 cm) is removed, fixed in 20 N Mildform solution (containing 8% formaldehyde in a buffered solution), and embedded in paraffin blocks, and sections of 5 μm are put on glass slides. Apoptosis is detected using the terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling (TUNEL) method, using the Apop Tag Peroxidase In Situ Apoptosis Detection Kit.
Rats^[4]
Male Sprague-Dawley rats with body weight between 230-250 g and age between 6-7 weeks are used. About 25 SD rats are divided into five groups receiving Docetaxel (7 mg/kg, i.v.), PIP (35 mg/kg, p.o.) and their combined administration (DOX+PIP) as well as PIP (3.5 mg/kg, p.o.) and PIP (3.5 mg/kg, i.v.). A day before the drug administrations, the rats are anesthetized. Right jugular vein is cannulated with a polyethylene tubing (0.5 mm ID, 1 mm) for blood collection.

MCE has not independently confirmed the accuracy of these methods. They are for reference only.

• [1]. Attia RT, et al. The chemomodulatory effects of glufosfamide on docetaxel cytotoxicity in prostate cancer cells. PeerJ. 2016 Jun 29;4:e2168.  [Content Brief]

  [2]. Che CL, et al. DNA microarray reveals different pathways responding to NSC 125973 and docetaxel in non-small cell lung cancer cell line. Int J Clin Exp Pathol. 2013 Jul 15;6(8):1538-48.  [Content Brief]

  [3]. Obi-Ioka Y, et al. Involvement of Wee1 in the circadian rhythm dependent intestinal damage induced by docetaxel. J Pharmacol Exp Ther. 2013 Oct;347(1):242-8.  [Content Brief]

  [4]. Li C, et al. Non-linear pharmacokinetics of piperine and its herb-drug interactions with docetaxel in Sprague-Dawley rats. J Pharm Biomed Anal. 2016 Sep 5;128:286-93.  [Content Brief]