Monocytic Differentiation in B-Lymphoblastic Leukemia with CRLF2 rearrangement at Diagnosis: Implications for Pathogenesis and Post-Therapeutic Lineage Switch

The majority of BCR::ABL1-like (Ph-like) B-lymphoblastic leukemia (B-ALL) is characterized by genetic rearrangements involving the CRLF2 gene (CRLF2r). This subtype is enriched in adolescent/young adult patients and those of Hispanic ethnicity. The biologic basis of the relatively unfavorable prognosis and chemotherapeutic resistance in this disease is not well understood. Immunophenotypic evidence of monocytic differentiation in CRLF2r leukemia at diagnosis, detailed in a case report (Choi et al, AJCP 2017) and two examples of lineage switch (LS) to acute myeloid leukemia (AML) of CRLF2r B-ALL at relapse (Silbert et al, submitted), raises the hypothesis that CRFL2r B-ALL arises from a pre-lineage committed progenitor rather than a B-cell progenitor. Here we present evidence of monocytic differentiation in a subset of CRLF2r B-ALL at disease presentation.

At our large clinical flow cytometry practice at Children's Hospital Los Angeles (CHLA, 70% Hispanic patients), multiparametric flow cytometry at new B-ALL diagnosis routinely utilizes antibodies to screen for expression of CRLF2, cytoplasmic myeloperoxidase (cMPO), and monocyte-associated antigens (CD4, CD14, CD33, CD64 and CD371). New leukemias showing co-expression of CRLF2 and one or more monocytic antigens were reviewed (Jan. 2021 - June 2024). Cases with available frozen mononuclear cells underwent cell sorting of the B-cell and monocytic components of the progenitors, and each component was interrogated for CRLF2r by fluorescence in situ hybridization (FISH).

Among 223 de novo B-ALLs, 31 (13.9%) had CRLF2r, 7 (22.6%) of which showed partial monocytic differentiation. These 7 cases were from 4 females and 3 males, all Hispanic, with a median age of 16 years (range 4-20 years). Two had constitutional trisomy 21 (Down syndrome, DS). Five (5) of the 7 cases had CRLF2r partners demonstrated by FISH during the diagnostic clinical work-up of bulk leukemia cells [rearrangement partners: 3 IGH (including 1 DS), 2 P2RY8 (including 1 DS), 2 not tested]. The monocytic component comprised 0.5-1.5% of total leukemic cells in 5 cases and 7.0% in a 6^th case. In a 7^th case, the monocytic component was identified in an end of Induction (EOI) sample evaluated for measurable residual disease (MRD) and represented 62.7% of the 1.2% MRD calculated from mononuclear cells per Children's Oncology Group guidelines (or 0.51% of total nucleated cells); monocytic antigens had not been interrogated on the diagnostic sample. In all 7 cases, the monocytic component was initially suggested by increased CD45 expression and increased forward- and side-scatter properties as well as by decreased CD19 and variable CD10 expression compared to the majority B-lineage component. Monocytic differentiation was confirmed in all 7 cases by variable (maturational) CD14 and/or CD64 expression with or without cMPO, CD4, CD13+33 or CD33 (bright), and CD371. In the 4 cases with available frozen mononuclear cells, CRLF2r was demonstrated by FISH in the myelomonocytic cell-sorted fraction.

The data show that monocytic differentiation in CRLF2r B-ALL at diagnosis is a relatively common and likely underrecognized phenomenon. This observation raises the possibility of disease origin in a primitive hematopoietic cell prior to B-lineage commitment, as well as the possibility that monocytic differentiation may emerge with increasing frequency with growing use of CD19-targeted therapies (leading to possible full LS). Such targeted therapies also may not optimally treat these B-ALL given their decreased CD19 expression. The relative significance of monocytic differentiation in both DS-associated as well as non-syndromic individuals is not clear, as CRLF2r B-ALL is known to have different biology in these genetic settings. Reconciliation of these observations with the myelomonocytic differentiation that may be seen at presentation or following Induction chemotherapy in other subtypes of B-ALL, i.e. the clinically favorable DUX4-rearranged B-ALL, is needed. Moreover, the prevalence of monocytic differentiation in other B-ALL subtypes, including other forms of Ph-like ALL, remains unknown. These findings provide insight into the biology of Ph-like B-ALL and suggest more work is needed to define the boundary between this entity and mixed phenotype acute leukemia (MPAL), B/Myeloid.

Parekh:Pluto: Current equity holder in private company, Patents & Royalties: receives royalties for technology licensed to Pluto that is unrelated to the study in this abstract; Amgen: Other: spouse is Amgen employee and owns Amgen stock. Wood:Amgen: Consultancy; Cellnomics LLC: Current equity holder in private company.