Validation of bypass mechanisms of resistance to ALK TKIs in ALCL
Supervisor: Šárka POSPÍŠILOVÁ
ALK inhibitors have been slow to reach the clinic for children with ALK-expressing ALCL but experience of treating adults tells us that resistance is a distinct possibility and rapid relapses on cessation of therapy with ALK TKIs have been reported for those children that have received ALK TKIs.
Whilst resistance can develop due to mutations in the target protein, bypass tracks are also prevalent representing alternative therapeutic options. We have previously conducted CRISPRa screens and have identified a plethora of potential bypass resistance tracks.
These will be compared to the transcriptional profiles of
- Cell lines made resistant to ALK TKI by culture in increasing concentrations of the drugs;
- Bone fide relapse tumour material from patients;
- PDX resistant to ALK inhibitor, in order to select the most relevant pathways for further investigation.
Initially, the ability of the identified proteins to reduce cell sensitivity to ALK TKI will be investigated by specific induction of sgRNA in dCas9 expressing cell lines. Rescue experiments with co-transfection of shRNA to inhibit the identified protein together with CRISPRa overexpression will also be carried out. The effect the expression/repression of these proteins has on the ALK TKI IC50 will be determined. We will then identify pathways affected by expression of the resistance-inducing alteration by RNAseq and seek out drugs that inhibit their activity working with the Milner Therapeutics Institute. The ability of these drugs to inhibit the growth of ALK TKI-resistant cell lines and PDX will be assessed as will their potential synergy with ALK TKI for the treatment of inhibitor naïve disease.
Ultimately, bypass tracks that drive resistance to ALK TKI in ALCL will have been explored and identified. We expect to have validated at least 5 pathways and to have developed therapeutic protocols to counteract their effects. In addition, using PDX, dosing mechanisms would have been explored and validated to prevent resistance to ALK TKI from developing, for example, metronomic versus continuous dosing.
Year 1: THT Bio: Development of organoid model systems working alongside scientists at the company who have significant expertise in this technology (4 months)
Year 2: Masters Chancellors and Scholars of the university of Cambridge: Access to PDX for drug studies uniquely available in the host institute who will also provide training in animal studies (4 months)