Understanding the effect of adaptive mutations on the three-dimensional structure of RNA

Single-nucleotide polymorphisms (SNPs) are variations in the genome where one base pair can differ between individuals.1 SNPs occur throughout the genome and can correlate to a disease-state if they occur in a functional region of DNA.1According to the central dogma of molecular biology, any variation in the DNA sequence will have a direct effect on the RNA sequence and will potentially alter the identity or conformation of a protein product. A single RNA molecule, due to intramolecular base pairing, can acquire a plethora of 3-D conformations that are described by its structural ensemble. One SNP, rs12477830, which was previously shown to harbor signatures of positive selection by Sugden et. al,3 was passed through multiple RNA folding algorithms. The results of SNPfold 2 demonstrate that the SNP significantly alters the structural ensemble, and the significance of this change offers a potential explanation of SWIF(r)’s result.3 Furthermore, the RNAfold Webserver 4-6reveals that the mutant RNA molecule is more stable than the wild-type with a more negative free energy and a higher frequency. These loci of variation should be studied in order to understand the potentially induced conformational changes that could significantly alter the functional capacity of an RNA molecule. Future work aims to assess conformational changes elicited by SNPs previously shown to harbor signatures of positive selection using ancestry-specific reference genomes to better understand motivations behind a locus experiencing positive selective pressure

ProcessDriver: A Computational Pipeline to Identify Copy Number Drivers and Associated Disrupted Biological Processes in Cancer

Copy number amplifications and deletions that are recurrent in cancer samples harbor genes that confer a fitness advantage to cancer tumor proliferation and survival. One important challenge in computational biology is to separate the causal (i.e., driver) genes from passenger genes in large, aberrated regions. Many previous studies focus on the genes within the aberration (i.e., cis genes), but do not utilize the genes that are outside of the aberrated region and dysregulated as a result of the aberration (i.e., trans genes). We propose a computational pipeline, called ProcessDriver, that prioritizes candidate drivers by relating cis genes to dysregulated trans genes and biological processes. ProcessDriver is based on the assumption that a driver cis gene should be closely associated with the dysregulated trans genes and biological processes, as opposed to previous studies that assume a driver cis gene should be the most correlated gene to the copy number of an aberrated region. We applied our method on breast, bladder and ovarian cancer data from the Cancer Genome Atlas database. Our results included previously known driver genes and cancer genes, as well as potentially novel driver genes. Additionally, many genes in the final set of drivers were linked to new tumor events after initial treatment using survival analysis. Our results highlight the importance of selecting driver genes based on their widespread downstream effects in trans

Challenges in identifying cancer genes by analysis of exome sequencing data.

Massively parallel sequencing has permitted an unprecedented examination of the cancer exome, leading to predictions that all genes important to cancer will soon be identified by genetic analysis of tumours. To examine this potential, here we evaluate the ability of state-of-the-art sequence analysis methods to specifically recover known cancer genes. While some cancer genes are identified by analysis of recurrence, spatial clustering or predicted impact of somatic mutations, many remain undetected due to lack of power to discriminate driver mutations from the background mutational load (13-60% recall of cancer genes impacted by somatic single-nucleotide variants, depending on the method). Cancer genes not detected by mutation recurrence also tend to be missed by all types of exome analysis. Nonetheless, these genes are implicated by other experiments such as functional genetic screens and expression profiling. These challenges are only partially addressed by increasing sample size and will likely hold even as greater numbers of tumours are analysed

Plasma Cytokine and Growth Factor Profiling during Free Flap Transplantation

Ischemia and reperfusion (I/R) is an unavoidable condition during free flap transplantation. Restoration of blood flow is usually associated with a profound inflammatory response. Cytokines and growth factors are the functional proteins which exert their specific influence on injury or repair during the healing period. Plasma concentrations of 18 cytokines and growth factor proteins (IL6, IL8, IP10, TNFα, MCP1, Fractalkine, GRO, bFGF, GMCSF, IFNg, MIP1a, VEGF, sCD40L, IL10, TGFα, IL1β, IL12P40, and TNFβ) have been analyzed with respect to I/R status during microsurgery tissue transplantation in both, artery and vein, from patients by multiplexed immunoassay. Both technical feasibility and biostatistics data analysis approaches were thoroughly assessed. It has been found that, from all investigated proteins, the venous plasma levels of IL6 significantly increased during the ischemia period and mostly sustained their high levels during reperfusion, while venous plasma levels of IL8 showed in general a significant increase in the ischemia period followed by a rapid decrease in the reperfusion period. In conclusion, these findings direct toward an active involvement of tissue-resting leukocytes which may become therapeutic targets for concomitant medication in flap surgery to improve wound healing

Detection of novel fusion-transcripts by RNA-Seq in T-cell lymphoblastic lymphoma

Fusions transcripts have been proven to be strong drivers for neoplasia-associated mutations, although their incidence in T-cell lymphoblastic lymphoma needs to be determined yet. Using RNA-Seq we have selected 55 fusion transcripts identified by at least two of three detection methods in the same tumour. We confirmed the existence of 24 predicted novel fusions that had not been described in cancer or normal tissues yet, indicating the accuracy of the prediction. Of note, one of them involves the proto oncogene TAL1. Other confirmed fusions could explain the overexpression of driver genes such as COMMD3-BMI1, LMO1 or JAK3. Five fusions found exclusively in tumour samples could be considered pathogenic (NFYG-TAL1, RIC3-TCRBC2, SLC35A3-HIAT1, PICALM MLLT10 and MLLT10-PICALM). However, other fusions detected simultaneously in normal and tumour samples (JAK3-INSL3, KANSL1-ARL17A/B and TFG-ADGRG7) could be germ-line fusions genes involved in tumour-maintaining tasks. Notably, some fusions were confirmed in more tumour samples than predicted, indicating that the detection methods underestimated the real number of existing fusions. Our results highlight the potential of RNA-Seq to identify new cryptic fusions, which could be drivers or tumour-maintaining passenger genes. Such novel findings shed light on the searching for new T-LBL biomarkers in these haematological disorders.The authors would like to thank the Spanish Biobanks integrated in the Spanish Hospital Biobanks Network (RetBioH; www.redbiobancos.es) for providing us with the necessary T-LBL samples to elaborate this work. We thank all patients who were willing to donate their samples without their support the research work would not be possible. And to Isabel Sastre for her technical support. This work was supported by the Spanish Ministry of Economy and Competitiveness (SAF2015-70561-R; MINECO/FEDER, EU); the Autonomous Community of Madrid, Spain (B2017/BMD-3778; LINFOMAS-CM) and the Spanish Association Against Cancer (AECC, 2018; PROYE18054PIRI). Institutional grants from the Fundación Ramón Areces and Banco de Santander are also acknowledged.S

ASSOCIATION BETWEEN LEUKEMIC EVOLUTION AND UNCOMMON CHROMOSOMAL ALTERATIONS IN PEDIATRIC MYELODYSPLASTIC SYNDROME

Background and objective: Pediatric myelodysplastic syndrome (pMDS) is a group of rare clonal neoplasms with a difficult diagnosis and risk of progression to acute myeloid leukemia (AML). The early stratification in risk groups is essential to choosing the treatment and indication for allogeneic hematopoietic stem cell transplantation (HSCT). According to the Revised International Prognostic Scoring System, cytogenetic analysis has demonstrated an essential role in diagnosis and prognosis. In pMDS, abnormal karyotypes are present in 30-50% of the cases.  Monosomy 7 is the most common chromosomal alteration associated with poor prognosis. However, the rarity of specific cytogenetic alterations makes its prognosis uncertain. Thus, this study aimed to describe uncommon cytogenetic alterations in a cohort of 200 pMDS patients and their association with evolution to AML. Methods: The cytogenetic analysis was performed in 200 pMDS patients by G-banding and fluorescence in situ hybridization between 2000 to 2022. Results: Rare chromosome alterations were observed in 7.5% (15/200) of the cases. These chromosome alterations were divided into four cytogenetic groups: hyperdiploidy, biclonal chromosomal alterations, translocations, and uncommon deletions, which represented 33.3%, 33.3%, 20%, and 13.3%, respectively. Most of these patients (10/15) were classified with advanced MDS (MDS-EB and MDS/AML) and the initial subtype was present in five patients (RCC). The leukemic evolution was observed in 66.66% (10/15) of the patients. Most patients had poor clinical outcomes and they were indicated for HSCT.  Conclusion: The study of uncommon cytogenetic alterations in pMDS is important to improve the prognosis and guide early indication of HSCT.  Keywords: Pediatric MDS; Leukemic evolution; rare chromosomal altwerations; HSCT, Childre

Diagnostic utility of whole genome sequencing in adults with B-other acute lymphoblastic leukemia

Genomic profiling at diagnosis of B-cell precursor Acute Lymphoblastic Leukemia (BCP-ALL) in adults is used to guide disease classification, risk stratification and treatment decisions. Patients for which diagnostic screening fails to identify disease defining or risk stratifying lesions are classified as B-other ALL. We screened a cohort of 652 BCP-ALL cases enrolled in UKALL14 to identify and perform whole genome sequencing (WGS) on paired tumor-normal samples. For 52 B-other patients we compared WGS findings to data from clinical and research cytogenetics. WGS identifies a cancer associated event in 51/52 cases, this includes an established subtype defining genetic alteration in 5/52 that were previously missed by standard-of-care genetics. Of the 47 true B-other ALL we identified a recurrent driver in 87% (41). Complex karyotype by cytogenetics emerges as a heterogeneous group, including distinct genetic alterations associated with either favorable (DUX4-r) or poor outcomes (MEF2D-r, IGK::BCL2). For a subset of 31 cases, we integrate findings from RNA-sequencing (RNA-seq) analysis to include fusion gene detection, and classification by gene expression. Compared to RNA-seq, WGS was sufficient to detect and resolve recurrent genetic subtypes, however RNA-seq can provide orthogonal validation of findings. In conclusion, we demonstrate that WGS can identify clinically relevant genetic abnormalities missed by standard-of-care testing and identify leukemia driver events in virtually all cases of B-other ALL