Biology subjects

Hong, Y.

Publications and source records attributed to Hong, Y..

At least 127 records · Page 7Linked to original sources

bioRxiv2022

Global Spatial Transcriptome of Macaque Brain at Single-Cell Resolution

Global profile of gene expression at single-cell resolution remains to be determined for primates. Using a recently developed technology ("Stereo-seq"), we have obtained a comprehensive single-cell spatial transcriptome map at the whole-brain level for cynomolgus monkeys, with [~]600 genes per cell for 10 m-thick coronal sections (up to 15 cm2 in size). Large-scale single-nucleus RNA-seq analysis for [~]1 million cells helped to identify cell types corresponding to Stereo-seq gene expression profiles, providing a 3-D cell type atlas of the monkey brain. Quantitative analysis of Stereo-seq data revealed molecular fingerprints that mark distinct neocortical layers and subregions, as well as domains within subcortical structures including hippocampus, thalamus, striatum, cerebellum, hypothalamus and claustrum. Striking whole-brain topography and coordinated patterns were found in the expression of genes encoding receptors and transporters for neurotransmitters and neuromodulators. These results pave the way for cellular and molecular understanding of organizing principles of the primate brain.

cell biology↗

bioRxiv2022

Class-Controlled Copy-Paste Based Cell Segmentation for CoNIC Challenge

Muti-class cell segmentation in histopathology images is a challenging task. Here, we propose a copy-paste augmentation-based method for CoNIC challenge. As the challenge train data is severely class imbalanced. To deal with it, we copy all cell objects of train data and paste them to the train image on the fly while training model. The paste strategy is that we paste more cell objects of the insufficient classes and paste less cell objects for the sufficient classes. We experimented the method by stratified splitting train data in 4:1 ratio, the result shows the copy paste method can reach PQ 64.84 and mPQ 53.72, which improved and 0.66 compared to without copy pasted. Moreover, the improvements in those insufficient classes is more obvious.

pathology↗

bioRxiv2022

Loss of β-ketoacyl acyl carrier protein synthase III activity restores multidrug-resistant Escherichia coli sensitivity to previously ineffective antibiotics

Antibiotic resistance is one of the most prominent threats to modern medicine. In the latest World Health Organization list of bacterial pathogens that urgently require new antibiotics, nine out of 12 are Gram-negative, with four being of Critical Priority. One crucial barrier restricting antibiotic efficacy against Gram-negative bacteria is their unique cell envelope. While fatty acids are a shared constituent of all structural membrane lipids, their biosynthesis pathway in bacteria is distinct from eukaryotes making it an attractive target for new antibiotic development that remains less explored. Here, we interrogated the redundant components of the bacterial Type II Fatty Acid Synthesis (FAS II) pathway, showing that disrupting FAS II homeostasis in Escherichia coli through deletion of the fabH gene damages the cell envelope of antibiotic susceptible and antibiotic resistant clinical isolates. The fabH gene encodes the {beta}-ketoacyl acyl carrier protein synthase III (KAS III), which catalyzes the initial condensation reactions during fatty acid biosynthesis. We show that fabH null mutation potentiated the killing of multi-drug resistant E. coli by a broad panel of previously ineffective antibiotics, despite the presence of relevant antibiotic resistance determinants, for example, carbapenemase kpc2. Enhanced antibiotic sensitivity was additionally demonstrated in the context of eradicating established biofilms and treating established human cell infection in vitro. Our findings showcase the potential of FabH as a promising target that could be further explored in the development of therapies that may repurpose currently ineffective antibiotics or rescue failing last-resort antibiotics against Gram-negative pathogens. IMPORTANCEGram-negative pathogens are a major concern for global public health due to increasing rates of antibiotic resistance and the lack of new drugs. A major contributing factor towards antibiotic resistance in Gram-negative bacteria is their formidable outer membrane, which acts as a permeability barrier preventing many biologically active antimicrobials from reaching the intracellular targets and thus limiting their efficacy. Fatty acids are the fundamental building blocks of structural membrane lipids and their synthesis constitutes an attractive antimicrobial target as it follows distinct pathways in prokaryotes and eukaryotes. Herein, we identified a component of fatty acid synthesis, FabH, as a gate-keeper of outer membrane barrier function. Without FabH, Gram-negative bacteria become susceptible to otherwise impermeable antibiotics and are re-sensitised to killing by last-resort antibiotics. This study supports FabH as a promising target for inhibition in future antimicrobial therapies.

microbiology↗

bioRxiv2022

uniPort: a unified computational framework for single-cell data integration with optimal transport

Single-cell data integration can provide a comprehensive molecular view of cells. Here we introduce uniPort, a unified single-cell data integration framework which combines a coupled Variational Autoencoder (coupled-VAE) and Minibatch Unbalanced Optimal Transport (Minibatch-UOT). It leverages both highly variable common and dataset-specific genes for integration and is scalable to large-scale and partially overlapping datasets. uniPort jointly embeds heterogeneous single-cell multi-omics datasets into a shared latent space. It can further construct a reference atlas for online prediction across datasets. Meanwhile, uniPort provides a flexible label transfer framework to deconvolute spatial heterogeneous data using optimal transport space, instead of embedding latent space. We demonstrate the capability of uniPort by integrating a variety of datasets, including single-cell transcriptomics, chromatin accessibility and spatially resolved transcriptome data. uniPort software is available at https://github.com/caokai1073/uniPort.

bioinformatics↗

bioRxiv2022

Bile Acid Restrained T Cell Activation Explains Cholestasis Aggravated Hepatitis B Virus Infection

Cholestasis is a common complication of Hepatitis B Virus (HBV) infection, characterized by increased intrahepatic and plasma bile acid levels. Cholestasis was found negatively associated with hepatitis outcome, however; the exact mechanism by which cholestasis impact on anti-viral immunity and impede HBV clearance remains elusive. Here, we found that cholestatic mice are featured with dysfunctional T cell response, and bile acids inhibit the activation and metabolic reprogramming of CD4+ T cells. Mechanistically, bile acids disrupt intracellular calcium homeostasis via inhibiting mitochondria calcium uptake and elevating cytoplasmic Ca2+ concentration of CD4+ T cells, leading to STIM1 and ORAIL1 decoupling and impaired store-operated Ca2+ entry which is essential for NFAT signaling and T cell activation. Moreover, in a transgenic mouse model of HBV infection, it was confirmed that cholestasis compromised T cells activation resulting in poor viral clearance. Collectively, our results suggest that bile acids play pivotal roles in anti-HBV infection via controlling T cells activation and metabolism, and that targeting regulation of bile acids may be a therapeutic strategy for host virus defense.

immunology↗

bioRxiv2022

Lineage-mosaic and mutation-patched spike proteins for broad-spectrum COVID-19 vaccine

The widespread SARS-CoV-2 in humans results in the continuous emergence of new variants. Recently emerged Omicron variant with multiple spike mutations sharply increases the risk of breakthrough infection or reinfection, highlighting the urgent need for new vaccines with broad-spectrum antigenic coverage. Using inter-lineage chimera and mutation patch strategies, we engineered a recombinant monomeric spike variant (STFK1628x), which showed high immunogenicity and mutually complementary antigenicity to its prototypic form (STFK). In hamsters, a bivalent vaccine comprised of STFK and STFK1628x elicited high titers of broad-spectrum antibodies to neutralize all 14 circulating SARS-CoV-2 variants, including Omicron; and fully protected vaccinees from intranasal SARS-CoV-2 challenges of either the ancestral strain or immune-evasive Beta variant. Strikingly, the vaccination of hamsters with the bivalent vaccine completely blocked the within-cage virus transmission to unvaccinated sentinels, for either the ancestral SARS-CoV-2 or Beta variant. Thus, our study provides new insights and antigen candidates for developing next-generation COVID-19 vaccines.

microbiology↗

bioRxiv2022

Regulators of proteostasis are translationally repressed in fibroblasts from sporadic and LRRK2-G2019S Parkinson's patients

Parkinsons disease (PD) is characterized by heterogeneous motor and non-motor symptoms. The majority of PD cases are sporadic, whereas the leucine-rich repeat kinase 2 (LRRK2) G2019S mutation is the most common monogenic cause of familial PD and symptomatically indistinguishable from the sporadic form. Deficits in protein synthesis have been reported in PD. However, it is not known which proteins are affected or if there are synthesis differences between patients with sporadic and LRRK2 G2019S PD. Here we used non-canonical amino acid labeling and mass-spectrometry based proteomics to perform a quantitative comparison of nascent proteins in fibroblasts from sporadic and G2019S mutation PD patients compared to healthy individuals. We found that several proteins were under-represented among nascent proteins in cells from PD patients. Among these were regulators endo-lysosomal sorting, mRNA processing and translation itself. We further validated which of these proteins were dysregulated at the static proteome level using targeted proteomics. Key downregulated proteins included the autophagy regulator ATG9A and translational stability regulator YTHDF3. Notably, 68% of the identified proteins were downregulated in both sporadic and LRRK2-G2019S forms of PD. We identify translational repression of key regulators of proteostasis in patient fibroblasts that are common to both sporadic and LRRK2-G2019S PD.

neuroscience↗

bioRxiv2022

Dynamic Turnover of PI4P and PI(4,5)P2 under Hypoxia Controls Electrostatic Plasma Membrane Targeting

Phosphatidylinositol 4-phosphate (PI4P) and phosphatidylinositol 4,5-biphosphate (PIP2) are key phosphoinositides that determine the identity of the plasma membrane (PM) and regulate numerous key biological events there. To date, mechanisms regulating the homeostasis and dynamic turnover of PM PI4P and PIP2 in response to various physiological conditions and stresses remain to be fully elucidated. Here we report that hypoxia in Drosophila induces acute and reversible depletion of PM PI4P and PIP2 that severely disrupts the electrostatic PM targeting of multiple polybasic polarity proteins. Genetically encoded ATP sensors confirmed that hypoxia induces acute and reversible reduction of cellular ATP levels which showed a strong real-time correlation with the levels of PM PI4P and PIP2 in cultured cells. By combining genetic manipulations with quantitative imaging assays we showed that PI4KIII, as well as Rbo/EFR3 and TTC7 that are essential for targeting PI4KIII to PM, are required for maintaining the homeostasis and dynamic turnover of PM PI4P and PIP2 under normoxia and hypoxia. Our results revealed that in cells challenged by energetic stresses triggered by hypoxia, ATP inhibition and possibly ischemia, dramatic turnover of PM PI4P and PIP2 could have profound impact on many cellular processes including electrostatic PM targeting of numerous polybasic proteins.

cell biology↗

bioRxiv2022

The salivary, metal-binding peptide histatin-5 buffers extracellular copper availability

Antimicrobial peptides (AMPs) are key components of diverse host innate immune systems. The family of human salivary AMPs known as histatins bind Zn and Cu. Fluctuations in Zn and Cu availability play significant roles in the host innate immune response (so-called "nutritional immunity"). Thus, we hypothesised that histatins contribute to nutritional immunity by influencing host Zn and/or Cu availability. We posited that histatins limit Zn availability (promote bacterial Zn starvation) and/or raise Cu availability (promote bacterial Cu poisoning). To test this hypothesis, we examined the interactions between histatin-5 (Hst5) and Group A Streptococcus (GAS), which colonises the human oropharynx. Our results showed that Hst5 does not strongly influence Zn availability. Hst5 did not induce expression of Zn-responsive genes in GAS, nor did it suppress growth of mutant strains that are impaired in Zn transport. Biochemical examination of purified peptides confirmed that Hst5 binds Zn only weakly. By contrast, Hst5 bound Cu tightly and it strongly influenced Cu availability. However, Hst5 did not promote Cu toxicity. Instead, Hst5 suppressed expression of Cu-inducible genes, stopped intracellular accumulation of Cu, and rescued growth of a {Delta}copA mutant strain that is impaired in Cu efflux. We thus proposed a new role for salivary histatins as major Cu buffers in saliva that contribute to microbial homeostasis in the oral cavity and oropharynx by reducing the potential negative effects of Cu exposure (e.g. from food) to microbes. Our results raise broad questions regarding the physiological roles of diverse metal-binding AMPs and the management of host metal availability during host-microbe interactions.

microbiology↗

bioRxiv2021

High Resolution Nanostructure with Two-stages of Exponential Energy Dissipation at the Ultrathin Osteochondral Interface Tissue of Human Knee Joint

Cartilage adheres to subchondral bone via a specific osteochondral interface tissue where forces are transferred from soft cartilage to hard bone without fatigue damage over a lifetime of load cycles. However, the fine structure and mechanical properties of osteochondral interface tissue remain unclear. Here, we identified an ultrathin [~]20-30 m calcified region with two-layered micro-nano structures of osteochondral interface tissue in human knee joint, which exhibited characteristic biomolecular compositions and complex nanocrystals assembly. Within this region, an exponential increase of modulus (3 orders of magnitude) was conducive to the force transmission which was verified by finite element simulations. The nanoscale heterogeneity of hydroxyapatite, along with enrichment of elastic-responsive protein-titin which is usually present in muscle, endowed the osteochondral tissue with excellent energy dissipation and fatigue resistance properties. Our results provide potential design for high-performance interface materials for osteochondral interface regeneration and functional coatings.

bioengineering↗

bioRxiv2021

Mechanically active integrins direct cytotoxic secretion at the immune synapse

The secretory output of cell-cell interfaces must be tightly controlled in space and time to ensure functional efficacy. This is particularly true for the cytotoxic immune synapse (IS), the stereotyped junction formed between a cytotoxic lymphocyte and the infected or transformed target cell it aims to destroy1. Cytotoxic lymphocytes kill their targets by channeling a mixture of granzyme proteases and the pore forming protein perforin directly into the IS2, 3. The synaptic secretion of these toxic molecules constrains their deleterious effects to the target cell alone, thereby protecting innocent bystander cells in the surrounding tissue from collateral damage. Despite the importance of this process for immune specificity, the molecular and cellular mechanisms that establish secretory sites within the IS remain poorly understood. Here, we identified an essential role for integrin mechanotransduction in cytotoxic secretion using a combination of single cell biophysical measurements, ligand micropatterning, and functional assays. Upon ligand-binding, the L{beta}2 integrin LFA-1 functioned as a spatial cue, attracting lytic granules containing perforin and granzyme and inducing their fusion at closely adjacent sites within the synaptic membrane. LFA-1 molecules were subjected to pulling forces within these secretory domains, and genetic or pharmacological suppression of these forces abrogated cytotoxicity. We conclude that lymphocytes employ an integrin-dependent mechanical checkpoint to enhance both the potency and the security of their cytotoxic output.

immunology↗

bioRxiv2021

PhosPiR: An automated phospho-proteomic pipeline in R

Large-scale phospho-proteome profiling using mass spectrometry (MS) provides functional insight that is crucial for disease biology and drug discovery. However, extracting biological understanding from this data is an arduous task requiring multiple analysis platforms that are not adapted for automated high-dimensional data analysis. Here, we introduce an integrated pipeline that combines several R packages to extract high-level biological understanding from largescale phosphoproteomic data by seamless integration with existing databases and knowledge resources. In a single run, PhosPiR provides data clean-up, fast data overview, multiple statistical testing, differential expression analysis, phospho-site annotation and translation across species, multi-level enrichment analyses, proteome-wide kinase activity and substrate mapping and network hub analysis. Data output includes graphical formats such as heatmap, box-, volcano- and circos-plots. This resource is designed to assist proteome-wide data mining of pathophysiological mechanism without a need for programming knowledge.

bioinformatics↗

bioRxiv2021

A-Prot: Protein structure modeling using MSA transformer

In this study, we propose a new protein 3D structure modeling method, A-Prot, using MSA Transformer, one of the state-of-the-art protein language models. For a given MSA, an MSA feature tensor and row attention maps are extracted and converted into 2D residue-residue distance and dihedral angle predictions. We demonstrated that A-Prot predicts long-range contacts better than the existing methods. Additionally, we modeled the 3D structures of the free modeling and hard template-based modeling targets of CASP14. The assessment shows that the A-Prot models are more accurate than most top server groups of CASP14. These results imply that A-Prot captures evolutionary and structural information of proteins accurately with relatively low computational cost. Thus, A-Prot can provide a clue for the development of other protein property prediction methods.

bioinformatics↗

bioRxiv2021

A TET1-PSPC1-Neat1 molecular axis modulates PRC2 functions in controlling stem cell bivalency

TET1 maintains hypomethylation at bivalent promoters through its catalytic activity in embryonic stem cells (ESCs). However, whether and how TET1 exerts catalytic activity-independent functions in regulating bivalent genes is not well understood. Using a proteomics approach, we mapped the TET1 interactome in mouse ESCs and identified PSPC1 as a novel TET1 partner. Genome-wide location analysis reveals that PSPC1 functionally associates with TET1 and Polycomb repressive complex-2 (PRC2) complex. We establish that PSPC1 and TET1 repress, and Neat1, the PSPC1 cognate lncRNA, activates the bivalent gene expression. In ESCs, Neat1 tethers the TET1-PSPC1 pair with PRC2 at bivalent promoters. During the ESC-to-formative epiblast-like stem cell (EpiLC) transition, PSPC1 and TET1 promote PRC2 chromatin occupancy at bivalent gene promoters while restricting Neat1 functions in facilitating PRC2 binding to bivalent gene transcripts. Our study uncovers a novel TET1-PSPC1-Neat1 molecular axis that modulates PRC2 binding affinity to chromatin and bivalent gene transcripts in controlling stem cell bivalency. In BriefTET1 is a transcriptional repressor for bivalent genes in pluripotent stem cells, but its mechanistic action on stem cell bivalency is unclear. Huang et al. use proteomics and genetic approaches to reveal that catalytic activity-independent functions of TET1, coordinated with the paraspeckle components PSPC1 and its cognate lncRNA Neat1, dynamically regulates stem cell bivalency by modulating PRC2 binding affinity to chromatin and bivalent gene transcripts in pluripotent state transition. HighlightsO_LIThe TET1 interactome identifies PSPC1 as a novel partner in ESCs C_LIO_LITET1 and PSPC1 repress bivalent genes by promoting PRC2 chromatin occupancy C_LIO_LINeat1 facilitates bivalent gene activation by promoting PRC2 binding to their mRNAs C_LIO_LINeat1 bridges the TET1-PSPC1 and PRC2 complexes in regulating bivalent gene transcription C_LI

developmental biology↗

bioRxiv2021

Modeling Western Pacific Amyotrophic Lateral Sclerosis and Parkinsonism dementia Complex with Microglia Containing Cerebral Organoids Derived from Induced Pluripotent Stem Cells

Western Pacific Amyotrophic Lateral Sclerosis and Parkinsonism-dementia Complex (ALS-PDC) is a neurodegenerative disease linked to the traditional consumption of cycad seeds by the Chamorro people of Guam. Little is known about the etiological role of cycad toxin in ALS-PDC. Patient-derived induced pluripotent stem cells were derived from age- and sex-matched affected and unaffected patient lymphoid cells then differentiated into cerebral organoids. After three months, the ALS-PDC affected organoids were smaller, their neurons had less extensive neurite outgrowth, and the organoids had more reactive astrocytes and M1 microglia, fewer resting and M2 microglia, and more open extracellular space. Most of these phenomena could be recapitulated by exposing unaffected organoids to {beta}-methylamino L-alanine (BMAA), a toxic amino acid produced by cyanobacteria living with cycad plants. Furthermore, ALS-PDC affected organoids exhibited an exacerbated neuroinflammatory response to BMAA exposure via activation of caspase1/NLRP3 inflammasome. A genome-wide transcriptome analysis of the organoids showed that the most down-regulated pathways were taurine, alanine, aspartate, and glutamate metabolism; protein digestion; and absorption. The most down-regulated biological processes were type I interferon signaling, regulation of neuron differentiation and extracellular matrix organization. Our results suggested that the etiology of ALS-PDC is due to metabolic disorders that shifted microglia to a more pro-inflammatory M1 state instead of a non-inflammatory, repairing M2 state, which exacerbated inflammation and reduced extracellular matrix strength. Supplementation of transforming growth factor beta (TGF-{beta}) to ALS-PDC affected organoids increased the expression of interferon-induced transmembrane proteins (IFITMs) and restored M2 microglia populations and extracellular matrix organization. Organoids containing networks of neurons, astrocytes, microglia derived from iPSC with our protocol provides an excellent cellular model for neurodegenerative disease modeling. Significance StatementO_LIWestern Pacific Amyotrophic Lateral Sclerosis and Parkinsonism-dementia Complex (ALS-PDC) cerebral organoids containing networks of neurons, astrocytes, and microglia were generated from patient specific lymphoid derived induced pluripotent stem cells. C_LIO_LIALS-PDC affected organoids were smaller, with neurons had less extensive neurite outgrowth, more reactive astrocytes and M1 microglia, fewer resting and M2 microglia, and more open extracellular matrix spaces when compared to ALS-PDC unaffected organoids. C_LIO_LIGenome-wide transcriptome analysis indicated that ALS/PDC affected organoids had significantly lower expression of genes related to vitamin B6, amino acid and protein glycation metabolisms, down-regulated type I interferon signaling, the regulation of neuron differentiation and extracellular matrix production. C_LIO_LIOur results implicated that the etiology of ALS-PDC is due to metabolic disorders that led the shift of microglia to more pro-inflammatory M1 state and less non-inflammatory resting, repairing M2 state of microglia subpopulation, which primed the exacerbated inflammation and reduced extracellular matrix strength. TGF-{beta} promoted interferon-induced transmembrane protein (IFITMs) expression and restored the repairing M2 state of microglia population and extracellular matrix organization. C_LI

neuroscience↗

bioRxiv2021

The genetic architecture of phenotypic diversity in the betta fish (Betta splendens)

The Betta fish displays a remarkable variety of phenotypes selected during domestication. However, the genetic basis underlying these traits remain largely unexplored. Here, we report a high-quality genome assembly and re-sequencing of 727 individuals representing diverse morphologies of the betta fish. We show that current breeds have a complex domestication history with extensive introgression with wild species. Using GWAS, we identify the genetic basis of multiple traits, including several coloration phenotypes, sex-determination which we map to DMRT1, and the long-fin phenotype which maps to KCNJ15. We identify a polygenic signal related to aggression with many similarities to human psychiatric traits, involving genes such as CACNB2 and DISC1. Our study provides a resource for developing the Betta fish as a genetic model for morphology and behavior in vertebrates.

genetics↗

bioRxiv2021

Notch-Induced Endoplasmic Reticulum-Associated Degradation Governs Thymocyte β-Selection

Signals from the pre-T cell receptor and Notch coordinately instruct {beta}-selection of CD4-CD8- double negative (DN) thymocytes to generate {beta} T cells in the thymus. However, how these signals ensure a high-fidelity proteome and safeguard the clonal diversification of the pre- selection TCR repertoire given the considerable translational activity imposed by {beta}-selection is largely unknown. Here, we identify the endoplasmic reticulum (ER)-associated degradation (ERAD) machinery as a critical proteostasis checkpoint during {beta}-selection. Expression of the SEL1L-HRD1 complex, the most conserved branch of ERAD, is directly regulated by the transcriptional activity of the Notch intracellular domain. Deletion of Sel1l impaired DN3 to DN4 thymocyte transition and severely impaired {beta} T cell development. Mechanistically, Sel1l deficiency induced unresolved ER stress that triggered thymocyte apoptosis through the PERK pathway. Accordingly, genetically inactivating PERK rescued T cell development from Sel1l- deficient thymocytes. Our study reveals a critical developmental signal controlled proteostasis mechanism that enforces T cell development to ensure a healthy adaptive immunity.

immunology↗

bioRxiv2021

B/tMSI-CAST: a systematic approach for microsatellite instability detection in both tumor tissue and plasma specimens from next-generation sequencing data

Microsatellite instability (MSI) is a well-established prognostic and predictive biomarker in certain types of cancers. MSI detection using tumour tissue is often limited by the availability of specimens. Next generation sequencing (NGS)-based MSI detection in plasma cell-free DNA (cfDNA) is challenged by a much lower signal-to-noise ratio. We developed a highly accurate cfDNA MSI detection method called bMSI-CAST (blood MSI Caller Adjusted with Sequence duplicaTes), with improvement on three features including a set of locus selection principles ensuring loci with high robustness and compatibility across sequencing platforms, an MSI-specific duplicate removal strategy, and a calling algorithm that dynamically matches baselines with a broad range of duplication levels. Analytical validation via MSI-high (MSI-H) cell gDNA showed an LOD of 0.15%. Furthermore, in an analysis of 95 evaluable cfDNA samples from patients with gastrointestinal cancers, bMSI-CAST exhibited a positive predictive agreement (PPA) of 92.9% (39/42) and negative predictive agreement (NPA) of 100% (53/53) with tissue MSI-PCR. In conclusion, bMSI-CAST provides novel and advanced solutions to key aspects fundamental to cfDNA MSI calling but not sufficiently addressed by existing methods, and it is a validated method ready to be applied to aid clinical decisions for cancer patients.

bioinformatics↗