Application of stem cell derived neuronal cells to evaluate neurotoxic chemotherapy

Abstract

The generation of induced pluripotent stem cells (iPSCs) and differentiation to cells composing major organs has opened up the possibility for a new model system to study adverse toxicities associated with chemotherapy. Therefore, we used human iPSC-derived neurons to study peripheral neuropathy, one of the most common adverse effects of chemotherapy and cause for dose reduction. To determine the utility of these neurons in investigating the effects of neurotoxic chemotherapy, we measured morphological differences in neurite outgrowth, cell viability as determined by ATP levels and apoptosis through measures of caspase 3/7 activation following treatment with clinically relevant concentrations of platinating agents (cisplatin, oxaliplatin and carboplatin), taxanes (paclitaxel, docetaxel and nab-paclitaxel), a targeted proteasome inhibitor (bortezomib), an antiangiogenic compound (thalidomide), and 5-fluorouracil, a chemotherapeutic that does not cause neuropathy. We demonstrate differential sensitivity of neurons to mechanistically distinct classes of chemotherapeutics. We also show a dose-dependent reduction of electrical activity as measured by mean firing rate of the neurons following treatment with paclitaxel. We compared neurite outgrowth and cell viability of iPSC-derived cortical (iCell® Neurons) and peripheral (Peri.4U) neurons to cisplatin, paclitaxel and vincristine. Goshajinkigan, a Japanese herbal neuroprotectant medicine, was protective against paclitaxel-induced neurotoxicity but not oxaliplatin as measured by morphological phenotypes. Thus, we have demonstrated the utility of human iPSC-derived neurons as a useful model to distinguish drug class differences and for studies of a potential neuroprotectant for the prevention of chemotherapy-induced peripheral neuropathy.

Human Induced Pluripotent Stem Cell Derived Sensory Neurons are Sensitive to the Neurotoxic Effects of Paclitaxel

“…Since multiple chemotherapeutic drugs cause sensory peripheral neuropathy, the effect of additional agents on neurite complexity was assessed. Concentrations of each drug needed for robust quantification of changes in neurite area were selected based on published data 13,19 and treatment was restricted to 72 hr based on the data from paclitaxel presented above.…”

“…iCell ® neurons are a mixture of postmitotic GABAergic and glutamatergic cortical neurons that are more characteristic of relatively immature forebrain neurons than the sensory neurons found in the DRG 17,18 . Peri.4U ® neurons are more peripheral-like, expressing IIItubulin, peripherin, MAP2 and vGLUT2, but have been minimally characterized with respect to functional properties 10,19 . Additionally, neurons derived from human fibroblasts, blood and embryonic stem cells that express more canonical nociceptive markers like ISL1, BRN3A, P2RX3, the NTRK receptors and NF200 [20][21][22][23] have also been used to study chemotherapy toxicity.…”

Concise Review: The Current State of Human In Vitro Cardiac Disease Modeling: A Focus on Gene Editing and Tissue Engineering

“…These examples show that the use of hiPSC for in vitro cardiac disease modeling without the presence of a known genetic defect are thus challenging, albeit not impracticable. This has also been demonstrated for other fields of disease, where hiPSC have been used to model noncardiac diseases like sporadic Alzheimer's disease , chemotherapy‐induced neuroticxicity , and was shown as a valuable tool in cancer research and precision oncology .…”

Human-Derived Brain Models: Windows into Neuropsychiatric Disorders and Drug Therapies

“…One primary consideration is to first determine whether the drug negatively impacts cell health and viability, in which case the observed effects to neurons may be caused by cellular stress responses rather than specific drug-induced changes to neuronal physiology. Multiple methods have been used for automated analysis of cell health, including abnormal nuclei, 80 viability dyes, such as calcein-AM, 79,87 and caspase assays for apoptosis. 87 Furthermore, 384-well cell culture plates can negatively impact cell viability and consistency across wells due to media loss through evaporation.…”

“…Multiple methods have been used for automated analysis of cell health, including abnormal nuclei, 80 viability dyes, such as calcein-AM, 79,87 and caspase assays for apoptosis. 87 Furthermore, 384-well cell culture plates can negatively impact cell viability and consistency across wells due to media loss through evaporation. 88 To overcome this limitation, researchers developed a ferromagnetic micro-raft array, where an array of 1,600 microrafts are cultured together and then magnetically transferred to a 384-well plate.…”