CD90-targeted nanoparticles for in vivo hematopoietic stem cell gene therapy

/ABSTRACT Patients with genetic blood diseases and disorders are commonly treated with blood (hematopoietic) stem cell (HSC) transplants from healthy, human leukocyte antigen (HLA)-matched sibling donors. Unfortunately, for most patients, there is no HLA-identical sibling donor available, and transplantation of HSCs from alternative donors can lead to life-threatening complications such as graft-vs-host disease (GVHD). To avoid severe side effects, especially from GVHD, correcting the patient's own HSCs—a process called gene therapy—has become a promising alternative treatment approach for genetic diseases and disorders affecting the blood. Despite significant progress in the field of HSC gene therapy, currently available treatment strategies are greatly limited by the requirement of highly sophisticated infrastructure in specialized facilities similar to bone marrow transplantations limiting the accessibility of this treatment option for low- and middle-income countries. The main bottleneck is the current inability to safely and efficiently perform HSC gene therapy directly in the patient (in vivo). The ability to genetically modify HSCs in vivo would 1) improve the feasibility of HSC gene therapy, 2) enable portable gene therapy, and 3) avoid expensive infrastructure. With the aim to refine the target for HSC gene therapy, we recently performed comprehensive studies in our nonhuman primate (NHP) stem cell transplantation and gene therapy model. This study identified a highly HSC-enriched cell population that contained all required cells for rapid short-term recovery and robust long-term multilineage engraftment of gene- modified cells providing the ideal target for in vivo gene therapy. Most importantly, we recently reported highly efficient encapsulation of gene-editing nucleases into polymeric and lipid nanoparticles (NPs). NPs can be surface modified for targeting, lyophilized for enhanced portability, and have been successfully used in vivo. Here, we hypothesize that targeting our novel HSC-enriched phenotype in vivo performing injections of NPs directly into the bone marrow stem cell compartment will allow us to overcome current limitations in HSC gene therapy. In Aim 1, we will optimize our NP formulations that can be loaded with next-generation double-strand break independent gene-editing agents and functionalize the surface for targeting. Untargeted as well as targeted NPs will be tested in vitro on cell lines and primary human HSCs for efficiency and specificity. In Aim 2, we will evaluate different versions of NP in the humanized mouse model to determine the biodistribution after intraosseous injections, on-target specificity, and long-term efficiency in vivo. Finally, in Aim 3 we will focus on the scale-up and clinical translation using our preclinical NHP large animal model. We will perform short-term studies to evaluate the biodistribution and safety profile of intramarrow NP injections, long-term studies to monitor persistence of editing, as well as in vivo selection to increase the frequency of gene editing. Project Number: 5R01HL173365-02 | Fiscal Year: 2026 | NIH Institute/Center: National Heart Lung and Blood Institute (NHLBI) | Principal Investigator: HANS-PETER KIEM | Institution: FRED HUTCHINSON CANCER CENTER, SEATTLE, WA | Award Amount: $756,008 | Activity Code: R01 | Study Section: Therapeutic Approaches to Genetic Diseases Study Section[TAG] View on NIH RePORTER: https://reporter.nih.gov/project-details/5R01HL17336502