The use of genetically-defined human disease models, such as X-MAN™ cell-lines, to define the optimum ‘on-target’ activity and patient populations for specific drug candidates, is now becoming fully embedded in the pharmaceutical pipeline.

Academic and industrial researchers are now increasingly turning to isogenic disease models in screens to find the next-generation of drug targets; or even novel drug candidates (see drug re-profiling section). Depending on how they are uncovered they could have substantial benefits over current targeted therapies that rely principally on ‘gene-addiction’.

The principle areas of need for future cancer therapies are:

1. For agents to be less susceptible to acquired drug resistance.  Signaling pathway agents such as Gefitinib are particularly prone to rapid resistance generation via secondary mutations in the target (e.g., T790M mutations); downstream or parallel pathway activation (e.g., loss of PTEN, or up-regulation of IGF1R); or even real-time feedback mechanisms (e.g., positive feedback within the PI3K pathway upon downstream target inhibition).

2. The ability to address undruggable or hard to address target genotypes that account for a large fraction of cancer types and drug resistance mechanisms (e.g., important oncogenes such as K-Ras, cMyc and β-catenin); and tumour supressors whose activity, or even presence, is absent in tumours (e.g., p53, APC and p16).

3. Agents that cause irreversible effects on tumour cells carrying specific disease driving mutations, so that ‘spikey’ dosing cycles in real patients are sufficient to provide tangible and long-term benefits.  Targeted agents tend to only inhibit tumour growth as long as the compound is present; which further exacerbates the generation of acquired resistance. 

These synthetic lethal processes are not exploiting ‘gene-addiction’ mechanisms, where such pathways are also used by normal cells to some degree; and therefore have the potential ability to be used more aggressively.  Moreover, acquired resistance may also be harder to generate given that the loss of DNA-repair genes, for example, cannot easily be re-acquired or compensated for by the tumour.

The prototypical case study for synthetic lethality is PARP-inhibitors (which inhibit PARPs function to mediate single-stranded DNA break repair) on cancer cells carrying mutations in BRCA proteins; which are consequently already severely compromised in repairing DNA-lesions via homologous recombination. 

Clinical studies of such PARP inhibitors in BRCA-null patients are showing remarkable tumour-regression effects and tolerability (Fong, P et al, NEJM, 2009).  Moreover, acquired resistance seems less opportune given that a ‘read-through’ reversion mutation had to be obtained in order to generate resistance in a BRCA2 truncation mutant cell-line treated with continuous low doses of PARP-inhibitor (Ashworth et al, Cancer Research, 2008).     

Recapitulating the patient synthetic lethality data; PARP inhibitors and DNA-damaging agents show remarkable potency on X-MAN cells carrying mutations in BRCA2 (see figure below) compared to their isogenic normal cells.  These cells therefore form ideal reagents to optimise second-generation PARP inhibitors, or even search for new targets that exploit defects in DNA-repair.

The reader should also note that many other DNA-repair X-MAN cell lines have been added to our product catalogue recently view »

Due to their simplicity of correlation to target patient genotypes and compatibility with HTS-platforms, X-MAN tumour vs. normal cell-line pairs form the ideal system for synthetic lethal target and drug-screens; wherein large siRNA, shRNA or compound libraries can be rapidly profiled to uncover all the genetic weak-points in cancer cells harbouring specific disease driving mutations. By screening compound libraries, one can also find novel drug candidates in a fraction of the time compared to standard biochemical-based approaches.

An example of this strategy is given below, showing a drug-candidate isolated from a screen of 30,000 compounds that displays selective toxicity for cancer cells harbouring a mutant form the signalling protein ‘K-Ras’; which has so far eluded conventional discovery pipelines.

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Two candidates were shown to have a significant therapeutic window and one was safe in active doses in-vivo (below) and was investigated further by a big Pharma company.

Horizon Offering

Horizon is now proud to offer both drug screening (through our HDS service arm) and the latest generation shRNA-pooled library screens through a collaboration with the word-renowned Ashworth laboratory.

Clients can also use X-MAN cell lines in a ‘systems biology’ approach to discover synthetic lethal networks and targets; and we welcome leaders in the field to contact us for collaborative efforts in this direction.

The particular power of understanding cell-biology to rapidly translate novel candidate treatments in to the clinic, was highlighted by our collaborative partner and SAB member, Professor Ben Park (Johns Hopkins University); who firstly demonstrated that GSK3b phosphorylation is elevated in cells carrying mutations in PI3K; and then counter to conventional wisdom, went on to show that known inhibitors of GSK3b developed originally developed for other therapeutic indications (e.g., Lithium Chloride) have selective effects on mutant PI3K-cells (see below). 

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Additional data-sets are available for each application and can be shared under confidential disclosure agreement. Please contact us for more information.

References

• Torrance et al, Nature Biotechnology, 2001
• Gustin et al, PNAS, 2009
• Featured publications
• All publications
• Fong et al, NEJM, 2009
• Ashworth, Cancer Research, 2008

Downloads

• Horizon Technology Presentation