Elsevier

Synergy

Volume 3, Issue 3, September 2016, Pages 15-30
Synergy

Full length paper
Drug combination in vivo using combination index method: Taxotere and T607 against colon carcinoma HCT-116 xenograft tumor in nude mice

https://doi.org/10.1016/j.synres.2016.06.001Get rights and content

Abstract

The median-effect equation (MEE) of the mass-action law and the combination index (CI) theorem have been used for quantitative determination of synergism (CI <1), antagonism (CI >1) and additive effect (CI = 1) in animals in vivo. Experimental design, the theoretical algorithm and the CompuSyn software simulation have been used to illustrate step-by-step for the combination of two anti-cancer agents, Taxotere and T607 compound, with similar mode of actions of targeting microtubule polymerization, but with distinct chemical structures. These two compounds acted synergistically against human colon carcinoma HCT-116 xenograft tumor in athymic nude mice. In all, only 78 nude mice have been used. The synergy is especially significant (p < 0.01–0.05) following Q3Dx4, x3 i.v. treatments, at higher doses and at later stages of treatment. The MEE and the CI theorem of Chou-Talalay quantitatively determined synergism or antagonism at different doses and different effect levels as indicated by the Fa-CI plot and by isobolograms in CompuSyn simulation and automated graphics. The practical logistics on pre-experimental planning, scheme/design/layout, and precautions in terms of dose number, dose range, dose density, drug combination ratio, conservation of laboratory animals as well as regulatory and cost-effective considerations have been presented. The mass-action law based CI algorithm has been proven to be simple to use, economy to practice, even for in vivo experimentations. Most significantly, the mass-action law based algorithm provides quantitative indexed conclusions.

Introduction

Drug combination has been widely used in treating various diseases including cancer, acquired immune deficiency syndrome (AIDS), and other diseases involving traditional Chinese medicines [1], [2]. Based on Google, PubMed and Web of Science, the term “drug combination” and “synergistic effect” are listed in >200,000 published scientific papers (Table 1). The large majority of them were carried out in vitro in cellular, cell-free, or molecular levels. The rest were carried out in vivo in animas or in clinical trials. Unfortunately, most of the animal studies in vivo and the phase I clinical trials have been carried out with experimental designs that are impossible to determine synergism or antagonism quantitatively [1], [3].

Among the twelve methods for synergism/antagonism assessments in bio-medical literature during the past eight decades (Table 2), the combination index (CI) method [4], [5] based on the physico-chemical principle of the median-effect equation (MEE) [6] of the mass-action law, with theoretical basis, experimental design, algorithm and computerized simulation, has been most widely used. The CI method quantitatively determines/simulates synergism or antagonism at all doses and all effects levels with small number of data points [1], [7], [8], [9], [10], [11], [12], [13]. So far, the CI method introduced by Chou and Talalay in 1984 [4] has been cited over 4681 times [Google Scholar Citations − Ting-Chao Chou] in over 711 different bio-medical journals (Thomson Reuters Web of Knowledge. http://www.researcherid.com/rid/B-4111-2009).

The drug combinations in vivo using the CI method have been presented in scientific meetings [14], [15] and briefly discussed in the review/commentary articles [8], [10], but have not been discussed in details with stepwise analysis in experimental design, actual data analysis and tactical and practical precautions in a full length paper. Recently, a step-by-step illustrations of drug combination in vitro for anti-tumor fludelone and panaxytriol in MX-1 cells have been published [9]. The present paper is prepared to provide progressively virtual illustration of anti-tumor drug combinations using the combination index method with Taxotere and T607 compound in human colon carcinoma HCT-116 xenograft tumor bearing athymic nude mice. It is intended to serve as a classic model illustration of quantitative analysis of drug interaction dynamics in complex biological systems.

Section snippets

Materials

Taxotere (Docetaxel, TXT) is a synthetic taxoil anticancer compound targeting microtubule polymerization. The T-900607 (T607) is a derivative of 2-fluoro-1-methoxy-4-pentafluorophenylsulfonamidobezene which covalently modifies β-tubulin residue Cys-239 as described previously [16]. T607, distinct from Taxotere, is efficacious in inhibiting the growth of both sensitive and multidrug resistant cancer cells. The Immunodeficient/athymic nude mice, nu/nu (female, 20–23 gm bodyweight), were purchased

Results

The dose-effect relationships for the therapeutic results of T607 (15, 25, 30, and 35 mg/kg, i.v. Q3Dx4, x3, n = 6 mice per dose for 4 doses), and vehicle control (6 mice), against HCT-116 xenograft tumor, total n = 30, beginning Day 14 to Day 41 are shown in Fig. 1. The full detailed numerical recording of tumor size changes is given in Appendix I in Supplementary information. The optimal/maximal therapeutic effect from Day 29 to Day 35 are summarized in Table 6. The average tumor volumes were

Grant support

This study was supported in parts by Aventis Pharmaceuticals, Bridgewater, NJ 08807; Tularik, Inc., South San Francisco, CA 94080; and SKI Institutional Core Fund, Memorial Sloan-Kettering Cancer Center, New York, NY 10065.

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