Conjoined Genes as Common Events in Childhood Acute Lymphoblastic Leukemia

SIMPLE SUMMARY: Acute lymphoblastic leukemia (ALL) is the most frequent childhood cancer. In recent years, broad application of NGS technologies enabled the discovery of novel genomically defined ALL. In this study, as a proof-of-principle, we applied RNA-seq technology to comprehensively profile the transcriptional landscape of a collection of 10 childhood BCP-ALL cases, and performed a deep bioinformatics analysis including several publicly available datasets, in order to characterize their full spectrum of transcriptional events. The paired-end RNA sequencing of our BCP-ALL pediatric cohort revealed a total of 9001 raw fusion events, which, after filtering, resulted in 245 candidate fusions. Overall, 235 out of 245 events were intra-chromosomal fusions, among which 229 involved two contiguous or overlapping genes, also known as conjoined genes (CGs). Among them, we identified a subset of 14 CGs (6.1%) exclusively expressed in leukemic cases but neither in solid cancers nor in normal samples. These events could be suggestive of a novel mechanism of transcriptional regulation in childhood leukemia and may represent novel potential leukemia-specific biomarkers. ABSTRACT: Acute lymphoblastic leukemia (ALL) is the most frequent childhood cancer. For the last three decades, conventional cytogenetic and molecular approaches allowed the identification of genetic abnormalities having prognostic and therapeutic relevance. Although the current cure rate in pediatric B cell acute leukemia is approximately 90%, it remains one of the leading causes of mortality in childhood. Furthermore, in the contemporary protocols, chemotherapy intensity was raised to the maximal levels of tolerability, and further improvements in the outcome will depend on the characterization and reclassification of the disease, as well as on the development of new targeted drugs. The recent technological advances in genome-wide profiling techniques have allowed the exploration of the molecular heterogeneity of this disease, even though some potentially interesting biomarkers such as conjoined genes have not been deeply investigated yet. In the present study, we performed the transcriptome sequencing (RNA-seq) of 10 pediatric B cell precursor (BCP)-ALL cases with different risk (four standard- and six high-risk patients) enrolled in the Italian AIEOP-BFM ALL2000 protocol, in order to characterize the full spectrum of transcriptional events and to identify novel potential genetic mechanisms sustaining their different early response to therapy. Total RNA was extracted from primary leukemic blasts and RNA-seq was performed by Illumina technology. Bioinformatics analysis focused on fusion transcripts, originated from either inter- or intra-chromosomal structural rearrangements. Starting from a raw list of 9001 candidate events, by employing a custom-made bioinformatics pipeline, we obtained a short list of 245 candidate fusions. Among them, 10 events were compatible with chromosomal translocations. Strikingly, 235/245 events were intra-chromosomal fusions, 229 of which involved two contiguous or overlapping genes, resulting in the so-called conjoined genes (CGs). To explore the specificity of these events in leukemia, we performed an extensive bioinformatics meta-analysis and evaluated the presence of the fusions identified in our 10 BCP-ALL cohort in several other publicly available RNA-seq datasets, including leukemic, solid tumor and normal sample collections. Overall, 14/229 (6.1%) CGs were found to be exclusively expressed in leukemic cases, suggesting an association between CGs and leukemia. Moreover, CGs were found to be common events both in standard- and high-risk BCP-ALL patients and it might be suggestive of a novel potential transcriptional regulation mechanism active in leukemic cells.