Disrupted Nuclear Export of Proteins Drives the Development of B-cell Malignancy

Shuttling of proteins between the nucleus and cytoplasm is a tightly regulated process that is essential for normal cellular function. Aberrant nuclear export is frequently observed in blood cancer (e.g., cytoplasmic localization of mutant NPM1 in acute myeloid leukemia), and novel mechanisms by which nuclear-cytoplasmic distribution of proteins might be altered are of great interest. Exportin-1 (XPO1) is the major nuclear export receptor in all eukaryotic cells that functions through identification of nuclear export signals (NESs) in the amino acid sequence of cargo proteins. Expression of XPO1 is reported to be increased in several cancers, and targeted XPO1 inhibitors are in clinical development. Furthermore, recurrent mutations in XPO1 have previously been reported to occur in B-cell malignancies including chronic lymphocytic leukemia (CLL) and Hodgkin lymphoma. However, the mechanism through which XPO1 dysfunction contributes to oncogenesis is unclear. This lack of evidence prompted Dr. Justin Taylor and colleagues to investigate the role of XPO1 mutations in oncogenesis in a study led by the Omar Abdel-Wahab Lab at the Memorial Sloan Kettering Cancer Center.

Analysis of whole-exome and -genome sequencing data from 42,793 patients with cancer identified recurrent and previously unrecognized mutational hotspots in XPO1. These mutations occurred as heterozygous point mutations and showed striking lineage specificity, with enrichment in a variety of B-cell malignancies, notably in CLL, primary mediastinal B-cell lymphoma, and classical Hodgkin lymphoma. The most frequent mutation was a point mutation (E571K) which always occurred alongside a retained wild-type allele, suggesting that the point mutation results in altered, rather than loss of, function. To investigate the functional and molecular impact of this mutation, the researchers developed an impressive array of new research tools including a genome-edited malignant B cell line (NALM-6 cells) engineered to carry the point mutation in XPO1, and a conditional knock-in mouse model allowing inducible expression of the Xpo1 mutation from the endogenous locus. The Xpo1^E571K mutation promoted B-cell proliferation both in vitro and in vivo and initiated the development of clonal B-cell malignancies, in some cases resembling human CLL. As mouse models that recapitulate a CLL phenotype are few and far between, this in itself is a notable contribution of this study. Combination of Xpo1 mutation with overexpression of c-MYC or BCL-2 led to a more aggressive lymphoproliferative phenotype in mouse models.

To evaluate how mutations in XPO1 might drive lymphoid malignancy, investigators carried out biochemical, structural, proteomic, and molecular studies. XPO1^E571K mutations were shown to alter nuclear export recognition in a sequence-specific manner, favoring the export of cargoes with negatively charged C-terminus NES sequences. This resulted in disruption of nucleo-cytoplasmic distribution of hundreds of proteins, including a number known to be implicated in cancer development. Interestingly, most proteins only showed changes in one compartment without the expected reciprocal change; for example, a protein that increased in abundance in the cytoplasmic compartment in the presence of XPO1 mutation did not usually show reciprocal reduced expression in the nucleus. Presence of XPO1^E571K mutation rendered cells sensitive to nuclear export inhibitors KPT-185 and the first-in-class selinexor (KPT-330), which was recently approved by the U.S. Food and Drug Administration.