ABSTRACT / PROJECT SUMMARY
Sickle-cell disease (SCD) is an autosomal recessive disorder that causes considerable
morbidity and mortality, affecting an estimated 100,000 individuals in the US, and millions more
worldwide. Multiple editing-based cures for SCD are currently in clinical development, however,
there are no clinical-grade laboratory tests available capable of characterizing the biophysical
and rheological properties of RBCs derived from genome-edited SCD HSPCs. Assays capable
of characterizing RBC quality are urgently needed to assess the potency of emerging
editing-based genomic therapies for SCD, regardless of the editing modality.
One of the central challenges that has impeded the development of a highly performant
potency assay for evaluating the functional efficacy of editing-based genomic therapies for SCD
has been the lack of laboratory technologies capable of sensitively, accurately, and precisely
capturing the biophysical properties of SCD-RBCs utilizing only a small number of cells. In
recent years, several innovations have emerged that now make the development and analytical
validation of a potency assay for editing-based SCD genomic therapies feasible. One of these
has been the advent of microfluidics-based diagnostic devices capable of functionally
characterizing the health of RBCs at unprecedented levels of resolution and sensitivity.
This proposal seeks to leverage (1) an existing suite of these aforementioned
next-generation RBC biophysical and functional characterization devices, (2) conventional
hematologic assays, and (3) a well-established machine learning approach to develop and
analytically validate a first-in-kind potency assay for editing-based therapies for SCD. To
achieve this, we will first construct a panel of comprehensively profiled, gold-standard reference
samples of HSPCs that simulate a representative range of genome editing outcomes in SCD
and prepare data for machine learning training. A machine learning model will then be trained to
predict the percentage of RBCs that functionally exhibit a non-SCD phenotype. Once trained,
we will validate the performance of the new potency assay, a panel of HSPCs affected by SCD
will be therapeutically edited using at least three different modalities (e.g. homology-directed
repair, base-editing, etc.).