Ribosome biogenesis and its regulation by the exportin, CSE1L, are targetable vulnerabilities in childhood and young adult Acute Myeloid Leukemia

Abstract There are fundamental differences in the genomic drivers of childhood, adolescent and young adult (AYA) acute myeloid leukemia (AML) compared to AML that develops in older adults. Despite this, little is known about the downstream pathways that regulate leukemogenesis in young AML. Although genetic alterations in the nuclear pore complex are more common in young patients (e.g., NUP98 rearrangements, DEK-NUP214 fusions), nucleocytoplasmic transport has not been exclusively studied in childhood/AYA AML. We identified enrichment of nucleocytoplasmic transport pathways in young (<40 years; mean age 27 in BeatAML & 29 in TGCA) versus older AML patients (>40 years, mean age 64 in BeatAML & 61 in TCGA) (BeatAML FDR=0.01, TCGA FDR=3.46e-3). Young patients with nucleoporin (NUP) rearrangements had high expression of nucleocytoplasmic transport genes, but this pathway was also upregulated in young NUP98r/DEK-NUP214 wildtype patients. Of the nucleocytoplasmic export genes, CSE1L was the top differentially expressed exportin in young AML. Immunoblotting confirmed increased CSE1L protein expression in childhood/AYA (n=10, mean age 23) compared to older (n=10, mean age 77) AML patient samples. Increased CSE1L mRNA expression predicted for decreased remission rates, event free survival and overall survival, exclusively in young, but not older patients with AML (TARGET, Beat AML). To further understand the dependency of childhood/AYA AML on CSE1L, we knocked down CSE1L with shRNA in cell lines derived from young AML patients (NB4, MV4-11, THP-1) and TEX cells, derived from transducing cord blood with a FUS-ERG fusion. CSE1L depletion reduced AML growth, viability and clonogenicity. Loss of AML viability and clonogenicity was confirmed using CSE1L CRISPR knockout. Knockdown of CSE1L in primary young AML cells reduced primary and secondary leukemic engraftment into mouse marrow. In contrast, depletion of CSE1L did not impair engraftment of normal cord blood into mouse marrow. To understand the function of CSE1L, we interrogated a BioID database and discovered that the CSE1L interactome was enriched for ribosome biogenesis processes (PXD007976). We validated the interaction with BioID hits, RPL29 and RPL32 (large ribosome subunit (60S) proteins), using proximity ligation assay and Co-IP. Co-IP further confirmed that CSE1L interacts exclusively with 60S, but not 40S ribosomal subunits. Knockdownof CSE1L in TEX cells led to nuclear accumulation of RPL29 and a 3.3 fold reduction in cytoplasmic RPL29. Using a GFP reporter assay that incorporates into the 60S subunit, we induced depletion of CSE1L and confirmed decreased cytoplasmic RPL29-GFP. Polysome profiling further validated a role for CSE1L in 60S export, as CSE1L depletion decreased cytoplasmic 60S but not 40S formation. Downstream of 60S export, a reduction in protein synthesis was observed in CSE1L depleted AML cells, relative to control shRNA cells. Given increased CSE1L expression in young patients with AML and its role in ribosome biogenesis, next we analyzed the top 20 differentially upregulated gene sets in young versus older AML patient samples and discovered that ribosome biogenesis pathways were the most abundantly expressed genes in AML blasts of young patients. GO processes highly enriched in young AML cells included rRNA processing, pre-ribosome, large ribosome subunit and ribosome biogenesis. Functionally, we demonstrated increased rRNA transcription rates in young (n = 10, mean age = 16.8 years) relative to older primary AML samples (n = 10, mean age = 69.7 years ). Next, we treated these samples with the RNA polymerase I inhibitor, BMH-21, and identified a differential sensitivity to BMH-21 in young AML cells. In summary, nucleocytoplasmic transport and ribosome biogenesis are positively correlated, differentially upregulated pathways in childhood/AYA AML. Depleting CSE1L, leads to impaired large ribosome subunit export, decreased protein synthesis and leukemic cell death. Likewise, AML cells of young patients have increased rates of ribosome biogenesis and are more sensitive to RNA polymerase I inhibitors than AML cells from older patients. Our findings confirm that childhood and older adult AML are fundamentally different diseases, driven by different biologic processes and reinforce the need to develop pediatric-specific treatment approaches to manage this disease in young patients.

Récupéré en direct depuis OpenAlex et désinversé. Les résumés ne sont pas conservés dans cette base de données : les index inversés représentent 8,6 Go des 9,3 Go de texte de la base, et le serveur dispose de 13 Go libres.