Mixed phenotype acute leukemia: Lineage assignment, immunophenotypic classification and genetic insights

Mixed phenotype acute leukemia (MPAL) is a rare and biologically heterogeneous category characterized by discrete admixed populations of myeloid and lymphoid blasts (bilineal or mixed lineage) or with coexpression of lymphoid and myeloid markers in a single blast population (biphenotypic or mixed phenotype). Despite advances in diagnostic criteria, MPAL remains a significant diagnostic and therapeutic challenge due to its immunophenotypic complexity and genetic diversity. Accurate classification relies heavily on multiparameter flow cytometry immunophenotyping, integrated with cytogenetic and molecular studies. The current World Health Organization classification and International Consensus Classification recognize specific genetic subtypes, including MPAL with t(9; 22)(q34.1; q11.2)/BCR::ABL1 and MPAL with KMT2A rearrangement, reflecting the increasing importance of genomic profiling in disease characterization. Recent genomic studies have revealed additional recurrent alterations involving transcription factors, epigenetic regulators, signaling pathways, and lineage-specifying genes, including ZNF384 rearrangements, BCL11B alterations, mutations involving PHF6, WT1, FLT3, and RUNX1, and mutations affecting chromatin remodeling and cytokine signaling pathways. Importantly, distinct genetic lesions appear to correlate with specific immunophenotypic patterns, suggesting biologically meaningful disease subsets and providing insights into mechanisms of lineage plasticity and leukemogenesis. This review summarizes the current understanding of the immunophenotypic and genetic landscape of MPAL, emphasizing the integration of morphologic, immunophenotypic, cytogenetic, and molecular findings into diagnosis and subclassification.