In vitro activity: S63845 is a potent, selective and high affinity small molecule inhibitor of MCL1 (myeloid cell leukemia 1) with Ki value < 1.2 nM. S63845 potently kills MCL1-dependent cancer cells, including multiple myeloma, leukaemia and lymphoma cells, by activating the BAX/BAK-dependent mitochondrial apoptotic pathway. In vivo, S63845 shows potent anti-tumour activity with an acceptable safety margin as a single agent in several cancers. Moreover, MCL1 inhibition, either alone or in combination with other anti-cancer drugs, proved effective against several solid cancer-derived cell lines. These results point towards MCL1 as a target for the treatment of a wide range of tumours. It is the first high affinity MCL-1 inhibitor with potent in vivo activity, it acts by binding to the BH3-binding groove of MCL-1 with a KD value of 0.19 nM for human MCL-1. S63845 is also highly selective against other BCL family proteins such as BCL-2 (Ki>10,000nM) and BCL-XL (Ki>10,000nM). S63845 showed the clear-on-target activity killing MCL-1-dependent cancer cells, including multiple myeloma, leukemia and lymphoma cells. It exhibited potent in vivo antitumor activity with an acceptable safety profile as monotherapy in the treatment of several cancer types.

Kinase Assay: 10 mM HEPES pH 7.4, 175 mM NaCl, 25 μM EDTA, 1 mM TCEP, 0.01% P20 and 1% DMSO is used as a running buffer. The ligand surface is generated using double His-tagged proteins. Serial dilutions of the compound in buffer are injected over the protein surface. All sample measurements are performed at a flow rate of 30 μL per min (injection time 120 s, dissociation time 360 s). The sensor surface is regenerated by consecutive injections of 0.35 M EDTA pH 8.0 with 0.1 mg/mL trypsin, 0.5 M imidazole and 45% DMSO (60 s, 15 μL per min).

Cell Assay: S63845 is a highly selective and potent MCL1 inhibitor. S63845 bound human MCL1 with KD value of 0.19 nM. S63845 was approximately 1,000-fold more potent in killing MCL1-dependent H929 multiple myeloma cells than MCL1 inhibitor A-1210477. S63845 also induced caspase-dependent phosphatidyl-serine exposure, PARP cleavage and cytochrome c release from mitochondria. In HeLa cells, S63845 disrupted binding of BAK and BAX to MCL1. S63845 killed cancer cells through activation of the BAX/BAK-dependent mitochondrial apoptotic pathway by direct inhibition of MCL1.