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Mazur et al. Rare Dis Orphan Drugs J 2023;2:1 https://dx.doi.org/10.20517/rdodj.2022.12 Page 3 of 10

ELANE MUTATIONS LEADING TO NEUTROPENIA
During the promyelocytic stage of granulopoiesis in the bone marrow, NE is synthesized as zymogen and
post-translationally activated in the neutrophil precursors [Figure 1A]. The NE activation steps include N-
terminal trimming by cathepsin C, which is required for NE protease activity. In addition, NE may be
[1]
modified at C-terminus by an as yet unidentified protease . The proteolytically-processed C-terminal is
thought to serve as a binding site for adaptor protein complex 3 (AP3), implicated in the transport of NE to
lysosome-like granules [1,13] .

NE mainly localizes to the primary granules of the neutrophil precursors, where it remains stored in a
catalytically active, ready-to-use form. Neutrophils that are released from the bone marrow into the
circulation, followed by recruitment to peripheral tissues, rely on catalytically active NE for their full
spectrum of antimicrobial, pro-inflammatory and tissue-remodeling functions . In addition to the
[14]
localization of NE to the primary granules in the neutrophil precursors and mature neutrophils, the
presence of this protease out of the granules, including in cytosol, at the cell surface and in the nucleus, has
also been documented [15-17] . In mature, circulating neutrophils, transient release of NE from the granules is
likely to be required for the physiological functions of these cells, such as generating and releasing
neutrophil extracellular traps, or regulating neutrophil migration [15,17] . However, NE misplaced from the
granules in the neutrophil precursors in the bone marrow is associated with neutrophil pathology, such as
SCN (see below).

ELANE-based neutropenia results from the gain-of-function of mutant NE rather than diminished levels of
native protein which might also be a consequence of monoallelic mutations . This is supported by several
[1]
lines of research. For example, a blockade of NE expression leads to the restoration of normal
granulopoiesis and neutrophil differentiation, regardless of the type of underlying genetic mutation .
[18]

The ELANE gene consists of 5 exons and 6 introns, and > 200 mutations, distributed over all exons as well
as some noncoding regions, have been described in association with congenital neutropenia [3,19] . However,
to link genetic ELANE variants with pathologic outcomes, mechanistic studies are required. Mouse models
of ELANE-targeted neutropenia have failed to accurately phenocopy human neutropenia because basal
granulopoiesis was not disrupted in mice upon expression of abnormal murine NE, targeted at a position
orthologous to human pathogenic NE mutation. A lack of suitable experimental neutropenia models has
hampered the testing of ELANE mutants in vivo in the context of this pathology . Human, murine or rat
[20]
promyelocytic cell lines transduced with mutant ELANE, or patient neutrophil or patient-derived induced
pluripotent stem cell (iPSC) lines, are typically used to unravel the biochemical and biological consequences
of different ELANE mutations in myeloid progenitors. On the basis of these studies, several ELANE
mutations have been identified to interfere with neutrophil maturation potential [Table 1].

According to recent studies, a comprehensive gene editing screen in the human hematopoietic stem and
progenitor cells (HSPCs) dissects the genetic bases of ELANE neutropenia pathogenicity. This model, in
which HSPCs were edited with sgRNAs that targeted a broad range of ELANE mutants in vitro, faithfully
[21]
recapitulated the known genetic features of ELANE-based neutropenia . In addition, human-edited HSPCs
have also been shown to produce an ELANE pathogenic variant-dependent abnormal hematopoietic
engraftment function, following infusion into specific immunodeficient mice. Thus, these new xenograft
models of gene-edited human HSPCs also enable the outcome of ELANE mutations to be tested in in vivo
settings, which was not possible before with ELANE transgenic mice .
[21]