In Vitro and In Vivo Profile of a Selective, Allosteric Akt2 Inhibitor Martin O'Rourke1, Caroline Boyd1, Estelle McLean1, Natalie Page1, Shane Rountree1, Steven Shepherd1, Mary McFarland1 Colin O’Dowd1, Graham Trevitt1, Iain James1, Timothy Harrison1,2, Ultan McDermott3 1Almac Discovery, Craigavon, United Kingdom; 2Centre for Cancer Research and Cell Biology, Queens University, Belfast, United Kingdom 3 Wellcome Trust Sanger Institute, Hinxton, Cambs, CB10 1SA
Biochemical efficacy of ADC-0008830, an Akt2 selective Pathway biomarker Overview inhibitor engagement in vitro Figure 1: Dose dependent inhibition of a) Akt1 b) Akt2 and c) Akt3 in an FP Figure 2: Inhibition of pathway • Akt protein family members (Akt biochemical assay biomarkers by ELISA 1-3) are involved in a wide variety a b c pGSK3β pPRAS 40 of biological processes including Cell line pAkt (nM) cell proliferation, apoptosis, and (nM) (nM)
tumorigenesis. PC3 60 100 15
(IC50 7,600nM) (IC 4,200nM) (IC 27nM) • We have identified a novel, 50 50 LnCaP 500 6000 3600 allosteric Akt 2 selective inhibitor, A2780 160 740 840 which exhibits single agent A549 130 129 150 efficacy in a panel of cell-lines. Panc-1 9 234 Not tested • In collaboration with the Sanger • ADC-0008830 is a potent inhibitor of the Akt2 isoform Institute, we have identified genes • ADC-0008830 inhibits key • No activity observed against PH domain null Akt isoforms confirming of sensitivity and resistance for components of the Akt pathway an allosteric mode of action subtype selective Akt inhibitors in multiple cell lines • Excellent selectivity against a kinase counterscreen panel (data not shown)
Dose and time dependent inhibition of Cellular profiling of ADC-0008939, a prototypical isoform selective Akt2 pathway biomarkers (Panc-1) inhibitor, in the Sanger cell line panel Figure 3: Dose and time dependent inhibition of Akt Figure 4: Pharmacogenomic analysis in 432 cell lines of ADC-0008939, a prototypical isoform selective pathway in Panc-1 cell line Akt inhibitor (IC50 Akt1 300nM; Akt2 7nM; Akt3 1230nM). Data was generated by the Sanger Institute. Panel a) Volcano plot showing genes of sensitivity and b) tissues of sensitivity. DMSO -5 -6 -7 -8 -9 pAkt a b
pGSK3β
Total Akt
Total GSK3β
0 1 4 24 48 72hrs 8830 DMSO 8830 DMSO 8830 DMSO 8830 DMSO 8830 DMSO 8830 DMSO
pAkt
Total Akt
pGSK3β
Total GSK3β • Genes of sensitivity: PIK3CA, CTNNB1, IDH1, AKT2, SETD2 ADC-0008830 dose dependently inhibits pAKT and pGSK3β • Breast cell lines showed the highest level of tissue sensitisation in Panc-1 cell line up to 72 hrs (1μM compound treatment) • Cell lines with RAS mutations were resistant to Akt2 inhibition
Pharmacokinetic / Pharmacodynamic properties of ADC-0008830 in Conclusions a Panc-1 xenograft Study Figure 5: a) Exposure of 30mg/kg ADC-0008830 in plasma and tumour, b) dose and time dependent pAkt ADC-0008830 is a novel, potent, allosteric knockdown in tumours by western blotting, and c) immunohistochemistry Akt2 inhibitor a b C C 1 1 1 4 4 4 8 8 8 24 24 24hrs pAkt Vehicle Potent anti-proliferative activity observed Total Akt in breast cancer cell lines
pAkt Genes of sensitivity identified using a 10mg/kg Total Akt prototypical Akt2 inhibitor by pharmacogenomic profiling (Sanger) pAkt 30mg/kg Total Akt ADC-0008830 has an excellent oral pharmacokinetic profile pAkt 100mg/kg Total Akt In a xenograft study, robust pathway c engagement observed at all doses
• ADC-0008830 has excellent exposure in mice ADC-0008830 growth delay xenograft over a 24hr period in both plasma and tumour studies are in progress • Significant inhibition of biomarker observed by Western and IHC at all doses up to 24hrs Further analogues with pharmaceutical • Complete inhibition of pAkt observed at all time profile suitable for pre-clinical points at 30 and 100 mg/kg doses development have been identified
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