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Use of a Machine Learning-Based High Content Analysis Approach to Identify Photoreceptor Neurite Promoting Molecules

  • John A. Fuller
  • Cynthia A. Berlinicke
  • James Inglese
  • Donald J. Zack
Conference paper
Part of the Advances in Experimental Medicine and Biology book series (AEMB, volume 854)

Abstract

High content analysis (HCA) has become a leading methodology in phenotypic drug discovery efforts. Typical HCA workflows include imaging cells using an automated microscope and analyzing the data using algorithms designed to quantify one or more specific phenotypes of interest. Due to the richness of high content data, unappreciated phenotypic changes may be discovered in existing image sets using interactive machine-learning based software systems. Primary postnatal day four retinal cells from the photoreceptor (PR) labeled QRX-EGFP reporter mice were isolated, seeded, treated with a set of 234 profiled kinase inhibitors and then cultured for 1 week. The cells were imaged with an Acumen plate-based laser cytometer to determine the number and intensity of GFP-expressing, i.e. PR, cells. Wells displaying intensities and counts above threshold values of interest were re-imaged at a higher resolution with an INCell2000 automated microscope. The images were analyzed with an open source HCA analysis tool, PhenoRipper (Rajaram et al., Nat Methods 9:635–637, 2012), to identify the high GFP-inducing treatments that additionally resulted in diverse phenotypes compared to the vehicle control samples. The pyrimidinopyrimidone kinase inhibitor CHEMBL-1766490, a pan kinase inhibitor whose major known targets are p38α and the Src family member lck, was identified as an inducer of photoreceptor neuritogenesis by using the open-source HCA program PhenoRipper. This finding was corroborated using a cell-based method of image analysis that measures quantitative differences in the mean neurite length in GFP expressing cells. Interacting with data using machine learning algorithms may complement traditional HCA approaches by leading to the discovery of small molecule-induced cellular phenotypes in addition to those upon which the investigator is initially focusing.

Keywords

Photoreceptor Neuritogenesis Imaging qHTS High content analysis Machine learning Phenotypic screening Protein kinase Inhibitor 

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Copyright information

© Springer International Publishing Switzerland 2016

Authors and Affiliations

  • John A. Fuller
    • 1
  • Cynthia A. Berlinicke
    • 1
  • James Inglese
    • 2
  • Donald J. Zack
    • 1
  1. 1.Ophthalmology, Molecular Biology & Genetics, Neuroscience, and Institute of Genetic MedicineWilmer Eye Institute, Johns Hopkins University School of MedicineBaltimoreUSA
  2. 2.NCATS, NHGRINational Institutes of HealthRockvilleUSA

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