Inhibition of DNA Topoisomerase II Selectively Reduces the Threat of Tumorigenicity Following Induced Pluripotent Stem Cell-Based Myocardial Therapy

Abstract

The advent of induced pluripotent stem cell (iPSC) technology creates new opportunities for transplant-based therapeutic strategies. The potential for clinical translation is currently hindered by the risk of dysregulated cell growth. Pluripotent stem cells reprogrammed by three-factor (Sox2, Klf, and Oct4) and four-factor (Sox2, Klf, Oct4, and c-Myc) strategies result in the capacity for teratogenic growth from residual pluripotent progeny upon in vivo transplantation. However, these pluripotent stem cells also have a stage-specific hypersensitivity to DNA-damaging agents that may allow separation of lineage-specific therapeutic subpopulation of cells. We aimed to demonstrate the selective effect of DNA topoisomerase II inhibitor, etoposide, in eliminating pluripotent cells in the early cardiac progenitor population thus decreasing the effect of teratoma formation. Immunodeficient murine hearts were infarcted and received implantation of a therapeutic dose of cardiac progenitors derived from partially differentiated iPSCs. Etoposide-treated cell implantation reduced mass formation in the intracardiac and extracardiac chest cavity compared with the same dose of iPSC-derived cardiac progenitors in the control untreated group. In vivo bioluminescence imaging confirmed the localization and engraftment of transplanted cells in the myocardium postinjection in both groups. Comparatively, the equivalent cell population without etoposide treatment demonstrated a greater incidence and size of teratoma formation. Hence, pretreatment with genotoxic etoposide significantly lowered the threat of teratogenicity by purging the contaminating pluripotent cells, establishing an adjunctive therapy to further harness the clinical value of iPSC-derived cardiac regeneration.

Tumor-Free Transplantation of Patient-Derived Induced Pluripotent Stem Cell Progeny for Customized Islet Regeneration

“…We found that close to 100% of immunocompromised recipient mice developed teratoma-like tumors when pancreatic progenitor cells derived from LV-and TGF-iPSCs were transplanted into kidney capsules of SCID-beige mice, indicating that lineage commitment through our original stepwise differentiation protocol may not be sufficient to ensure safe transplantation in the setting of iPSC-based therapy. Previously, purging undifferentiated pluripotent stem cells in ESC and iPSC products has been achieved through genetic selection, cell sorting of fully differentiated cell populations, or pluripotency-targeted drug treatments [34,[38][39][40][41]. Because human pluripotent stem cells, but not murine PSCs, are highly sensitive to enzymatic dissociation, and single-cell dissociation immediately kills human PSCs [32,33], here we used simple enzymatic dissociation steps to eliminate contaminated pluripotent cells from the iPSC-derived pancreatic progenitor population.…”

Effect of Induced Pluripotent Stem Cell Technology in Blood Banking

“…Most methods rely on mouse embryonic fibroblast (MEF) feeders and serum at some point during their culture. Because both MEF and serum can potentially be contaminated with xenogeneic pathogens, their use increases the risk to recipients; hence, serum‐free and xeno‐free protocols are being developed for generation of iPSCs [36] and redifferentiation to blood cells [37]. Redifferentiation of iPSCs to mature blood cell types seems the most difficult step in blood cell manufacturing from iPSCs [38].…”

Full biological characterization of human pluripotent stem cells will open the door to translational research

“…Small molecule inhibitors like quercetin, YM155 (Lee et al 2013), stearoyl-CoA desaturase inhibitor (SCD1) (Ben-David et al 2013) and etoposide (Wyles et al 2014) showed promising results regarding the depletion of undifferentiated stem cells and teratoma formation. Quercetin and YM155 are inhibitors of the anti-apoptotic factor survivin, thereby inducing cell death in undifferentiated cells, whereas hESC-derived dopaminergic neuronal and smooth muscle cells are not affected.…”

“…Treatment with the DNA topoisomerase inhibitor etoposide results in depletion of undifferentiated hiPSC from cardiac progenitor populations. Upon implantation into infarcted murine hearts, hiPSC-depleted conditions showed engraftment and mass growth, but no teratoma formation as compared to undepleted control injections (Wyles et al 2014). Chemical inhibitors bear the advantage that they can be supplied at stable conditions and in large amounts; however, possible side effects on the viability and functionality of differentiated cells must be thoroughly tested.…”