Development of indole derived non-orthosteric inhibitors for Proto-oncogene serine/threonine-protein kinase PIM-1

The feature that makes it easier to identify and optimize orthosteric inhibitors-their reliance on a well-defined active site-also limits their selectivity and leads to off-target effects. An example of this is found in the orthosteric inhibitors for the serine/threonine-protein kinase PIM-1. PIM-1 is a pro-oncogenic protein involved in tumor progression and drug resistance development, making it a prime therapeutic target. Indeed, several orthosteric PIM-kinase inhibitors have been developed, however, they interfere with off-target kinases, leading to toxicity. While non-orthosteric inhibitors are challenging to identify, we aimed to discover such an inhibitor for PIM-1 to improve selectivity over current orthosteric inhibitors. Therefore, we designed a luciferase reporter assay to identify potential non-orthosteric PIM-1 inhibitors from a specifically designed 41-compound library. Taking hit compounds through in vitro experiments, in-silico study, and BLI-analysis indicated that our identified inhibitors physically interact with PIM-1, do not inhibit PIM-1 kinase function, and decrease PIM-1 concentration under cellular conditions. These findings suggest these compounds interact with PIM-1 via a non-orthosteric binding site. In vivo study of the top three compounds in the EMT6 mouse tumor model indicated one compound, 6d, significantly reduced tumor burden. With these promising initial findings, we introduce novel non-orthosteric PIM-1 inhibitors as new strategy to target PIM-1 with high potential for low toxicity and, further down the road, for use in combination therapy to overcome resistance caused by PIM-1.