Kinase atlas of electrophysiological cardiotoxicity
Objective
Dysregulated kinase function is central to cancer pathophysiology and underpins the targeted therapeutic use of kinase inhibitors (KIs) as anti-cancer agents. Unfortunately, kinases are ubiquitous and kinase inhibitors poorly selective, leading to treatment-related toxicity, including cardiotoxicity. Such toxicity is not rare with over 70% of approved KIs were associated with treatment-related cardiotoxicity, leading to the development of the ‘cardio-oncology’ as a clinical subspecialty to best support these patients. The underlying cardiotoxicity mechanisms remain poorly elucidated and restricts the development of safer drugs or cardioprotective measures. Here, we utilized human induced pluripotent stem cell derived human cardiomyocytes (hiPSC-CMs) and high-throughput phenotypic screening (HTS) methods to identify kinases associated with the dysrythmic or arrhythmic effects of KIs in cardiomyocytes. We build a cardiotoxicity atlas with the identified kinases and applied this understanding to explain the cardiotoxicity phenotype of clinical KIs. Together, this resource makes way towards understanding cardiotoxicity mechanisms to enable safer drug discovery.
659
Kinases or associated proteins tested
6000+
Traces analysed
2
Healthy donor cell lines
Overview of screening setup for identifying kinases with cardiotoxicity liabilities by electrophysiological read-outs
siRNA knock-down in hiPSC-CMs leveraging high-throughput screening technology enables the determination of the cardiotoxic effects of inhibition or knock-down of individual kinases. A two-stage approach was applied to screen and validate kinases for cardiotoxic potential: A low-power primary screen to identify putative high risk kinases, followed by a high-power secundary screen to validate key kinases.
Primary screen: Kinases were prioritised for follow-up screening based on relative qualification of robustness of primary hit
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Validation screen:
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Overview of screened kinases and related proteins
Classification in cardiac kinase atlas
Protein function
Gene expression
Links
Primary screen
The primary screen of siRNA was conducted on two batches on duplicate wells and scored for multiple membrane potential metrics. Deviation from controls in excess of 3 standard deviations in at least three out of four wells, for any metric was considered as threshold for prioritization for secondary screening
Validation screen
The validation screen of siRNA was conducted on three batches with six replicate wells and scored for multiple membrane potential metrics. Deviation for controls was assessed using t-test statistics, and requiring additionally an effect size in excess of 20%. The area comprising metrics and batches reaching statistical significance was shaded red
Methodology
General
Voltage assay and extracting electrophysiological metrics from membrane potential traces
Example traces showcasing multiple different phenotypes
Multiple different phenotypes can be distinguished based on trace morphology, e.g., based on the beat frequency (tachy or fast, brady or slow), based on the length of the APD (prolongation or shortening), the appearance of early after-depolarizations (EADs), or the absence of activity (cessation).
Primary screen
The primary screen of siRNA was conducted on two batches on duplicate wells and scored for multiple membrane potential metrics. Deviation from controls in excess of 3 standard deviations in at least three out of four wells, for any metric was considered as threshold for prioritization for secondary screening
Validation screen
The validation screen of siRNA was conducted on three batches with six replicate wells and scored for multiple membrane potential metrics. Deviation for controls was assessed using t-test statistics, and requiring additionally an effect size in excess of 20%
Legend of icons used
References
About
Online resource of our paper 'An Atlas of Kinase inhibition-induced cardiomyocyte toxicity in human iPSC-cardiomyocytes'
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