The Combi-Targeting Concept:  Chemical Dissection of the Dual Targeting Properties of a Series of “Combi-Triazenes”

Abstract

The combi-targeting concept postulates that a molecule termed a "combi-molecule" designed to interact with an oncoreceptor on its own and allowed to further degrade to another more stable inhibitor of the latter receptor + a DNA-damaging species should be more potent than the individual combination of the same inhibitor with a DNA-damaging agent in cells expressing the targeted receptor. Recently, using the epidermal growth factor receptor (EGFR) as a target, we demonstrated the feasibility of combi-molecules with dual EGFR/DNA-targeting properties and with the ability to degrade to another potent inhibitor of EGFR. However, despite a clear demonstration of their superior potency when compared with classical combinations in EGFR-expressing cells, the true contribution of each fragment of the combi-molecules to their overall antiproliferative activity remained elusive. Here, we report a structure-function approach whereby a series of quinazoline-based "combi-triazenes" were altered to either abrogate the affinity of the EGFR-targeting quinazoline head or to suppress the DNA-damaging property of the triazene tail. The results showed that (a) inactivation of the quinazoline head by appending an N-methylaniline group to its 4-position reduced EGFR tyrosine kinase (TK) inhibitory activity by ca. 200-fold and decreased the ability of the combi-molecule to block serum-induced growth stimulation in c-erbB2 transfected NIH3T3 cells by ca. 10-fold, (b) abrogation of the alkylating activity or the DNA-damaging potential of the triazene tail by forming 3,3-dimethyltriazenes did not suppress EGFR TK inhibitory affinity but decreased the antiproliferative activity in basal growth assays, and (c) the antiproliferative activities of the monoalkyltriazenes that possessed binary EGFR TK inhibitory and alkylating activities were superior to those of their monotargeted counterparts. The results in toto suggest that each component of the dual targeting property of combi-triazenes plays a critical role in their overall antiproliferative activity.

Subcellular Distribution of a Fluorescence-Labeled Combi-Molecule Designed to Block Epidermal Growth Factor Receptor Tyrosine Kinase and Damage DNA with a Green Fluorescent Species

“…The growth of refractory tumors is driven by multiple signaling disorders that often cannot be blocked by the use of a single drug. The combi-molecule approach is the first that seeks to create molecules capable of blocking at least two divergent targets in the cells by allowing the intact molecules to block one target on their own and to degrade into other species directed at the same or different targets (18,20,22,23,26,40,41). To gain insight into the subcellular distribution and degradation of combimolecules, we designed a new prototype termed AL237 to (a) be fluorescent on its own and (b) degrade under physiologic conditions to FD105 (an EGFR inhibitor) that fluoresces in the blue and a DNA alkylating fragment that fluoresces in the green (see Fig.…”

Multiple mechanisms of action of ZR2002 in human breast cancer cells: A novel combi‐molecule designed to block signaling mediated by the ERB family of oncogenes and to damage genomic DNA

“…Within the same line of idea, we demonstrated recently that combi-molecules with binary EGFR targeting/DNA damaging properties and with the ability to be hydrolyzed to another EGFR inhibitor, induced sustained antiproliferative activity in cells overexpressing EGFR. [17][18][19][20] We report on a new type of combi-molecule that does FIGURE 7 -Reversibility of antiproliferative effect of ZR2002 (a), PD168393 (b) and ZR01 (c) in MDA-MB-453 cells. Cells were exposed to each drug for 2 hr, after which they were allowed to recover for 120 hr in drug free medium, or continuously for 120 hr.…”

“…9,14,20 The synthesis of ZR2002 will be reported elsewhere. In all assays, drug was dissolved in DMSO and subsequently diluted in RPMI-1640 containing 10% FBS (Wisent Inc., St.-Bruno, Canada) or in DMEM containing 10% FBS immediately before the treatment of cell cultures.…”

The Combi-Targeting Concept: Selective Targeting of the Epidermal Growth Factor Receptor- and Her2-Expressing Cancer Cells by the Complex Combi-Molecule RB24

“…Here, we describe a similar strategy based on the use of a small-molecule approach to target EGFR or Her2 TK-mediated signaling pathways while damaging DNA. This strategy, termed "combi-targeting," seeks to design a single molecule targeted to the TK function of Her2 or EGFR and programmed to be a latent DNA-damaging agent (Matheson et al, 2001(Matheson et al, , 2003(Matheson et al, , 2004aBrahimi et al, 2002Brahimi et al, , 2004Banerjee et al, 2003Banerjee et al, , 2004Rachid et al, 2003;Qiu et al, 2004).…”