Selectively eliminating residual human induced pluripotent stem cells (iPSCs) in cell mixtures for cell therapy - Abstract
Human-induced pluripotent stem cells (iPSCs) hold tremendous promise for cell therapy. However, the
significant tumorigenic risk associated with residual iPSCs in cell mixture during cell transplantation remains a
major obstacle to their clinical applications. To address this challenge, we have assembled a multidisciplinary
team consisting of a biomaterials chemist (Xu) and a stem cell biologist (Lu). Our goal is to develop and evaluate
transient intranuclear peptide assemblies (nPAs) to selectively eliminate residual iPSCs without impacting iPSC-
derived cells in cell mixture—an unmet need in the field. We recently achieved selectively eliminating iPSCs
without harming iPSC-derived hematopoietic progenitor cells (HPCs) using transient nPA. This selectivity hinges
on the well-documented overexpression of alkaline phosphatase (ALP) in iPSCs. ALP catalytically
dephosphorylates a phospho-L-peptide precursor, forming transient nPAs capable of eliminating iPSCs. This
nPA precursor exhibits exceptional selectivity, rapidly eradicating iPSCs in cell mixture within 2 hours due to
three key factors: (i) iPSCs express high levels of ALP, (ii) ALP expression decreases significantly in iPSC-
derived and normal cells, and (iii) nPAs comprise L-peptides that undergo proteolysis within the cellular
environment. However, it remains to be verified whether nPA precursors/building blocks would affect iPSC-
derived cells beyond HPCs and whether the elimination of residual iPSCs in the cell mixture is sufficient to
prevent tumor formation from the cell transplantation in vivo. Additionally, the design principles governing nPA
precursors have yet to be established. Therefore, we propose to explore nPAs for eliminating residual iPSCs
from cell mixtures containing iPSCs and iPSC-derived photoreceptor precursor cells (PPCs) by the studies
focusing on three specific aims: Aim 1: Design, synthesize, and characterize nPA precursors to conduct a
preliminary structural-to-activity relationship (SAR) study. Aim 2: Evaluate nPAs on iPSCs and iPSC-derived
photoreceptor precursor cells (PPCs) using cell assays. Aim 3: Examining the efficacy of nPAs to prevent tumor
formation from cell transplantation using a standard murine model. The successful completion of these studies
will provide a novel approach to eliminate residual iPSCs rapidly and selectively in cell mixtures, substantially
enhancing its safety profile of transplantation of iPSC-derived cells and thereby improving the treatment of
various human diseases.
