Abstract 47: A clinically curative combination therapy acts by low cross-resistance between drugs, not by synergistic interactions between drugs
Authors: Palmer Adam C. , Sorger Peter K. 
Published: August 2018

Abstract
Introduction: The development of optimal combination therapies is one of the central challenges of research to improve the treatment of cancers. The discovery of drug synergy is a common goal of preclinical development of combination therapies, but it is unclear whether this pharmacologic synergy, as experimentally measured in the potency of a dose response, is a property of clinically effective combination therapies. Furthermore, the historical motivation for combination therapy was not synergistic interactions per se but the use of different mechanisms of drug action to overcome subclonal cancer heterogeneity. Here we tested which of these interaction mechanisms are properties of a clinically exemplary combination, the chemo-immunotherapy regimen R-CHOP, which cures most cases of diffuse large B-cell lymphoma (DLBCL).
Methods: Pharmacodynamic interactions (synergy, additivity, antagonism) between the drugs in RCHOP were measured by Isobologram Analysis in a DLBCL cell culture system that facilitates in vitro activity of R-CHOP. Subclonal cross-resistance between drugs was measured using high-complexity DNA barcoding (ClonTracer) to label a million DLBCL subclones. Labelled subclones were expanded to billions and split into numerous replicate cultures. Each culture received different single drug treatment (each drug tested in triplicate cultures), and deep sequencing of DNA barcodes before and after treatment measured each subclone's resistance or sensitivity to each drug in the combination. Cross-referencing resistant subclones between drugs quantified distributions of drug-drug cross-resistance.
Results: Isobologram analysis of RCHOP revealed many antagonistic and at best additive interactions between drugs, despite its curative clinical effects. DNA barcoding experiments revealed that every drug in R-CHOP is resisted by thousands of subclones; hundreds of subclones can resist pairs of different drugs; tens of subclones can resist three different drugs; but not one subclone of a million has resistance to each drug in this combination.
Conclusion: The profound “clinical synergy” of RCHOP—cure—derives from overcoming within-tumor heterogeneity by combining different drugs with low cross-resistance, with no need for pharmacologic synergy, such as is measured by bulk cell inhibition assays. These experiments show that synergy of the pharmacologic variety is not essential to the clinical efficacy of a drug combination, and demonstrate the utility of lineage tracing by DNA barcoding as a method to identify drug combinations that can collectively overcome drug resistance. We propose that these high-throughput measurements of subclonal drug resistance and cross-resistance should be a complement to the measurement of drug interactions such as synergy in the preclinical development of combination therapies.
Citation Format: Adam C. Palmer, Peter K. Sorger. A clinically curative combination therapy acts by low cross-resistance between drugs, not by synergistic interactions between drugs [abstract]. In: Proceedings of the Second AACR Conference on Hematologic Malignancies: Translating Discoveries to Novel Therapies; May 6-9, 2017; Boston, MA. Philadelphia (PA): AACR; Clin Cancer Res 2017;23(24_Suppl):Abstract nr 47.