An innovative hematopoietic stem cell gene therapy approach benefits CLN1 disease in the mouse model

Abstract Hematopoietic stem and progenitor cells (HSPCs) can establish a long‐lasting microglia‐like progeny in the central nervous system of properly myeloablated hosts. We exploited this approach to treat the severe CLN1 neurodegenerative disorder, which is the most aggressive form of neuronal ceroid lipofuscinoses due to palmitoyl‐protein thioesterase‐1 (PPT1) deficiency. We here provide the first evidence that (i) transplantation of wild‐type HSPCs exerts partial but long‐lasting mitigation of CLN1 symptoms; (ii) transplantation of HSPCs over‐expressing hPPT1 by lentiviral gene transfer enhances the therapeutic benefit of HSPCs transplant, with first demonstration of such a dose–effect benefit for a purely neurodegenerative condition like CLN1 disease; (iii) transplantation of hPPT1 over‐expressing HSPCs by a novel intracerebroventricular (ICV) approach is sufficient to transiently ameliorate CLN1‐symptoms in the absence of hematopoietic tissue engraftment of the transduced cells; and (iv) combinatorial transplantation of transduced HSPCs intravenously and ICV results in a robust therapeutic benefit, particularly on symptomatic animals. Overall, these findings provide first evidence of efficacy and feasibility of this novel approach to treat CLN1 disease and possibly other neurodegenerative conditions, paving the way for its future clinical application.

= p < 0.0001 Repeated Measures ANOVA followed by Tukey's post-hoc test. D. Comparison of DSS with different disease manifestations (limbs clasping, stiffness and rotarod deficits) and with survival by Log-rank analysis. See Appendix Table S2 for statistics.
A. Schematic representation of the transplant procedure and animals monitoring post-transplant.

A. Representative fluorescence microscope photomicrographs showing human PPT1
immunoreactivity (hPPT1, red) in different brain regions of Ppt1 -/mice transplanted with hPPT1-LV transduced HSPCs (administered IV, ICV or ICV+IV) at 6-8 weeks of age or at 18-20 weeks of age (sympt tx) analyzed at study termination. Ppt1 -/untreated (UT) mice analyzed at about 250 days, i.e. humane end point, are shown as control of the specificity of hPPT1 staining. DAPI staining (blue) is shown to highlight nuclei. Scale bar = 100 µm. B. Quantification of hPPT1 immunostaining in different brain regions of untreated or transplanted mice analyzed at study termination. p>0.05; Kruskal Wallis followed by Dunn's post-hoc test.   Table S4. Statistics for longitudinal analyses among HSC gene therapy groups.

Fig. 2D. Longitudinal assessment of the DSS
A mixed-effect model was applied. The analysis was performed from day 140 to day 350, since more than 50% of values were missing for some groups afterwards (due to ICDs). The comparison was limited only to groups IV tx, ICV tx, ICV+IV tx and Ppt1 +/+ as for the others too many missing values (due to ICDs) were present after day 217.

Fig. 2G. Longitudinal assessment of Rotarod performance
A mixed-effect model was applied. The analysis was performed from day 140 to day 364, since more than 50% of values were missing for some groups afterwards (due to ICDs). The comparison was limited only to groups IV tx, ICV tx, ICV+IV tx and Ppt1 +/+ as for the others too many missing values (due to ICDs) were present after day 217.

Fig. S5B. Longitudinal assessment of DSS in symptomatic transplanted Ppt1 -/-mice.
A mixed-effect model was applied. The analysis was performed from day 140 to day 273, since more than 50% of values were missing for some groups afterwards (due to ICDs). The comparison was limited only to groups IV tx, ICV+IV tx and Ppt1 -/-UT as for the others too many missing values (due to ICDs) were present after day 250.