⦁ Molecular drivers of reprogramming – Our “Project Grandiose” (PG) generated a genome-wide, multi-omics, almost daily resolution characterization of the reprogramming process from somatic cells to induced pluripotent stem cells (iPSCs) [1-6]. Further mining of this unprecedented resource and target validation in the coming years will bring additional insight into the process of cell state change, teaching us how to generate alternative cell types with therapeutic relevance.
⦁ Alternative cell states – We also discovered and characterized a new class of iPSCs called F-class cells . Compared to embryonic stem cells (ESCs) and ESC-like iPSCs, these cells proliferate faster, are less heterogeneous, are amenable to suspension culture expansion, and exhibit a higher propensity to differentiate toward the neuronal lineage. Previously, the Haigh and Nagy labs described another novel cell type called induced vascular progenitor cells (iVPCs) that retain endothelial cell memory . These papers demonstrate that there are more applications for reprogramming beyond simply generating ESC-like pluripotent cells. PG has begun to illuminate the complex process of reprogramming and provides us with an extraordinary opportunity to generate and study alternative cell states of significant therapeutic potential.
⦁ Therapeutic cells – Our research aims to generate mouse and human therapeutic cell types for replacement therapies. We work with several disease models, including diabetes, blindness , arthritis, spinal cord injury , and stroke , and in vitro and in vivo cell types. Building upon decades of experience with transgenic mouse engineering, we are also working to make cells safer and more effective for transplantation by introducing novel functions, such as an inducible suicide switch, secreted therapeutics (e.g. VEGF “sticky-traps” ), anti-inflammatory ability and enhanced homing.
⦁ Gene therapy – Another research focus that has emerged from our extensive experience with transposon-mediated genetic engineering includes novel applications for gene therapy. In collaboration with Ian Alexander’s lab we developed a hybrid recombinant adeno-associated virus (rAAV)/piggyBac transposon vector system combining the liver-targeting properties of rAAV with stable piggyBac-mediated transposition of a transgene into the genome of liver cells.  Using this system, we effectively reversed the disease phenotype in two mouse models of urea cycle defects. Clinical translation of this technology could provide a bridging therapy for infants with severe urea cycle defects while awaiting liver transplantation. We are also exploiting new CRISPR/Cas9-mediated techniques for genome editing to further expand our “toolkit” in this area of research.
⦁ Combining cell and gene therapy – We propose to extend the use of cells beyond their own therapeutic effect by delivering transgene products to the diseases site. Such an extension of cell therapy could maximize the benefit of treatments by synergizing the effects of the two therapies. For example, we are working to introduce local acting anti-angiogenic VEGF “sticky-traps”  into retinal pigmented epithelial (RPE) cells to develop a novel cell therapy for the wet form of age-related macular degeneration. Incorporating VEGF sticky-traps into other therapeutic cell types may also provide improved angiostatic regulation for other diseases involving pathological neovascularisation.
1. Benevento M, et al., Proteome adaptation in cell reprogramming proceeds via distinct transcriptional networks. Nat Commun, 2014. 5: p. 5613. http://www.ncbi.nlm.nih.gov/pubmed/25494451
2. Clancy JL, et al., Small RNA changes en route to distinct cellular states of induced pluripotency. Nat Commun, 2014. 5: p. 5522. http://www.ncbi.nlm.nih.gov/pubmed/25494340
3. Hussein SM, et al., Genome-wide characterization of the routes to pluripotency. Nature, 2014. 516(7530): p. 198-206. http://www.ncbi.nlm.nih.gov/pubmed/25503233
4. Lee DS, et al., An epigenomic roadmap to induced pluripotency reveals DNA methylation as a reprogramming modulator. Nat Commun, 2014. 5: p. 5619. http://www.ncbi.nlm.nih.gov/pubmed/25493341
5. Shakiba N, et al., CD24 tracks divergent pluripotent states in mouse and human cells. Nat Commun, 2015. 6: p. 7329. http://www.ncbi.nlm.nih.gov/pubmed/26076835
6. Tonge PD, et al., Divergent reprogramming routes lead to alternative stem-cell states. Nature, 2014. 516(7530): p. 192-7. http://www.ncbi.nlm.nih.gov/pubmed/25503232
7. Haenebalcke L, et al., The ROSA26-iPSC mouse: a conditional, inducible, and exchangeable resource for studying cellular (De)differentiation. Cell Rep, 2013. 3(2): p. 335-41. http://www.ncbi.nlm.nih.gov/pubmed/23395636
8. Michael IP, et al., Local acting Sticky-trap inhibits vascular endothelial growth factor dependent pathological angiogenesis in the eye. EMBO Mol Med, 2014. 6(5): p. 604-23. http://www.ncbi.nlm.nih.gov/pubmed/24705878
9. Salewski RP, et al., The generation of definitive neural stem cells from PiggyBac transposon-induced pluripotent stem cells can be enhanced by induction of the NOTCH signaling pathway. Stem Cells Dev, 2013. 22(3): p. 383-96. http://www.ncbi.nlm.nih.gov/pubmed/22889305
10. Faiz M, et al., Adult Neural Stem Cells from the Subventricular Zone Give Rise to Reactive Astrocytes in the Cortex after Stroke. Cell Stem Cell, 2015. 17(5): p. 624-34. http://www.ncbi.nlm.nih.gov/pubmed/26456685
11. Cunningham SC, et al., Modeling correction of severe urea cycle defects in the growing murine liver using a hybrid recombinant adeno-associated virus/piggyBac transposase gene delivery system. Hepatology, 2015. 62(2): p. 417-28. http://www.ncbi.nlm.nih.gov/pubmed/26011400