Genomic Copy Number Research Articles

A Delicate Balance: CYFIP1 Dosage Drives Divergent Phenotypes

Genomic copy number variants (CNVs) that result in altered gene dosage have emerged as an important risk factor for neuropsychiatric disorders such as autism spectrum disorder (ASD) and schizophrenia (SCZ). Interestingly, patients harboring deletions or duplications of the same genomic region can present with shared and unique clinical phenotypes (1). One such example is at the 15q11.2 locus, which is associated with neurocognitive phenotypes including developmental delay, intellectual disability, and motor impairments.

Combination of iron flocculation and qPCR for quantitative evaluation of virus-shedding intensity of goldfish Carassius auratus infected with cyprinid herpesvirus 2 in the water and the effect of sodium chlorite powder in blocking waterborne horizontal viral transmission

Cyprinid herpesvirus 2 (CyHV-2), a member of the Alloherpesviridae family belonging to the genus Cyprinivirus, was initially isolated from goldfish (Carassius auratus) and has been recently emerging as a virulent pathogen for cultured prussian carp (Carassius auratus gibelio) world-wide. In this study, a novel and effective method for concentration and quantification of live CyHV-2 virions from water was successfully established through coupling the iron flocculation with real time qPCR assay. Then, the shedding intensity of CyHV-2 in fish-tank water from artificially-challenged goldfish (25 fish/20 L) was monitored continuously for 7 days on a daily basis through quantitating viral genomic copy numbers by qPCR, and the maximum shedding level was determined to be 105 copies/L. Horizontal transmission research system was established by inoculating healthy goldfish in water spiked with serial dilution of CyHV-2 virions ranging from 107 to 103 copies/L. Our results indicated that water-borne CyHV-2 efficiently caused the infection of tested goldfish even in a concentration of 103 copies/L, and the overall transmission efficacy was not linearly correlated with the level of input virus in the fish tank. Commercial disinfectant Composite Sodium Chlorite Powder (CSCP) has been widely applied in aquaculture to control microbial infection through direct spill in the water, and its effect in inactivating the CyHV-2 infectivity remains unknown. We further determined that the EC50 of CSCP against 3.89 TCID50/mL CyHV-2 was close to 15.625 μg/mL in vitro, and application of CSCP in a level as high as 60 μg/mL (the safety concentration of CSCP for goldfish) couldn't protect goldfish from CyHV-2 challenge through immersion. Thus, the disinfectant CSCP was regarded as none-effective for blocking CyHV-2 transmission in water during epidemic. Overall, our data provided quantitative data to demonstrate the shedding intensity of CyHV-2 in water, and CSCP was shown to be not effective in blocking water-borne horizontal transmission of CyHV-2 in goldfish. The virus-concentration protocol and virus-inhibition assay established here also paved the way for evaluating more commercial disinfectants in their effects in blocking water-borne horizontal transmission of CyHV-2.

Genetics Abnormalities with Clinical Impact in Primary Cutaneous Lymphomas.

Simple SummaryThe genetic landscape of cutaneous T-cell lymphomas analyzed by sequencing high throughput techniques shows a heterogeneous somatic mutational profile and genomic copy number variations in the TCR signaling effectors, the NF-κB elements, DNA damage/repair elements, JAK/STAT pathway elements and epigenetic modifiers. A mutational and genomic stratification of these patients provides new opportunities for the development or repurposing of (personalized) therapeutic strategies. The genetic heterogeneity in cutaneous B-cell lymphoma parallels with the specific subtype. Damaging mutations in primary cutaneous diffuse large B-cell lymphoma of the leg type, involving MYD88 gene, or BCL6 and MYC translocations or CDKN2A deletions are useful for diagnostic purposes. The more indolent forms, as the primary cutaneous lymphoma of follicle center cell (somatic mutations in TNFRSF14 and 1p36 deletions) and the cutaneous lymphoproliferative disorder of the marginal zone cells (FAS gene), present with a more restricted pattern of genetic alterations.Primary cutaneous lymphomas comprise a heterogeneous group of extranodal non-Hodgkin lymphomas (NHL) that arise from skin resident lymphoid cells and are manifested by specific lymphomatous cutaneous lesions with no evidence of extracutaneous disease at the time of diagnosis. They may originate from mature T-lymphocytes (70% of all cases), mature B-lymphocytes (25–30%) or, rarely, NK cells. Cutaneous T-cell lymphomas (CTCL) comprise a heterogeneous group of T-cell malignancies including Mycosis Fungoides (MF) the most frequent subtype, accounting for approximately half of CTCL, and Sézary syndrome (SS), which is an erythrodermic and leukemic subtype characterized by significant blood involvement. The mutational landscape of MF and SS by NGS include recurrent genomic alterations in the TCR signaling effectors (i.e., PLCG1), the NF-κB elements (i.e., CARD11), DNA damage/repair elements (TP53 or ATM), JAK/STAT pathway elements or epigenetic modifiers (DNMT3). Genomic copy number variations appeared to be more prevalent than somatic mutations. Other CTCL subtypes such as primary cutaneous anaplastic large cell lymphoma also harbor genetic alterations of the JAK/STAT pathway in up to 50% of cases. Recently, primary cutaneous aggressive epidermotropic T-cell lymphoma, a rare fatal subtype, was found to contain a specific profile of JAK2 rearrangements. Other aggressive cytotoxic CTCL (primary cutaneous γδ T-cell lymphomas) also show genetic alterations in the JAK/STAT pathway in a large proportion of patients. Thus, CTCL patients have a heterogeneous genetic/transcriptional and epigenetic background, and there is no uniform treatment for these patients. In this scenario, a pathway-based personalized management is required. Cutaneous B-cell lymphoma (CBCL) subtypes present a variable genetic profile. The genetic heterogeneity parallels the multiple types of specialized B-cells and their specific tissue distribution. Particularly, many recurrent hotspot and damaging mutations in primary cutaneous diffuse large B-cell lymphoma of the leg type, involving MYD88 gene, or BCL6 and MYC translocations and BLIMP1 or CDKN2A deletions are useful for diagnostic and prognostic purposes for this aggressive subtype from other indolent CBCL forms.

Development of a droplet digital PCR method for the sensitive detection and quantification of largemouth bass ranavirus.

Largemouth bass ranavirus (LMBRaV), also known as largemouth bass virus (LMBV), is a high mortality pathogen in largemouth bass. A rapid, sensitive, specific and convenient diagnosis method is an urgent requirement for the prevention of virus transmission. In the present study, a droplet digital PCR (ddPCR) method based on the major capsid protein (mcp) gene was established to detect and quantify the virus genome copy number. Oligonucleotide primers were designed based on the LMBRaV mcp gene sequence. The specificity and sensitivity of ddPCR assay were analysed. The other aquatic virus including Chinese giant salamander iridovirus (GSIV), Cyprinid herpesvirus II (CyHV-2) and infectious spleen and kidney necrosis virus could not be detected by LMBRaV ddPCR assay. The detection limit of ddPCR assay was 2 ± 0.37 copies/μl DNA sample. And this ddPCR assay had great repeatability and reproducibility. In clinical diagnosis of 50 largemouth bass, 43 positive samples were detected by ddPCR, whereas only 34 positive samples were detected by quantitative PCR (qPCR). This LMBRaV detection assay provided a specific and sensitive method for the rapid diagnosis of LMBRaV infection in largemouth bass as well as quantification of the virus load.

SARS-CoV-2 disrupts host epigenetic regulation via histone mimicry.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) emerged at the end of 2019 and caused the devastating global pandemic of coronavirus disease 2019 (COVID-19), in part because of its ability to effectively suppress host cell responses1–3. In rare cases, viral proteins dampen antiviral responses by mimicking critical regions of human histone proteins4–8, particularly those containing post-translational modifications required for transcriptional regulation9–11. Recent work has demonstrated that SARS-CoV-2 markedly disrupts host cell epigenetic regulation12–14. However, how SARS-CoV-2 controls the host cell epigenome and whether it uses histone mimicry to do so remain unclear. Here we show that the SARS-CoV-2 protein encoded by ORF8 (ORF8) functions as a histone mimic of the ARKS motifs in histone H3 to disrupt host cell epigenetic regulation. ORF8 is associated with chromatin, disrupts regulation of critical histone post-translational modifications and promotes chromatin compaction. Deletion of either the ORF8 gene or the histone mimic site attenuates the ability of SARS-CoV-2 to disrupt host cell chromatin, affects the transcriptional response to infection and attenuates viral genome copy number. These findings demonstrate a new function of ORF8 and a mechanism through which SARS-CoV-2 disrupts host cell epigenetic regulation. Further, this work provides a molecular basis for the finding that SARS-CoV-2 lacking ORF8 is associated with decreased severity of COVID-19.

Microbial molecule ingress promotes neuroinflammation and brain CCR5 expression in persons with HIV-associated neurocognitive disorders

BackgroundSystemic inflammation accompanies HIV-1 infection, resulting in microbial translocation from different tissues. We investigated interactions between lentivirus infections, neuroinflammation and microbial molecule presence in the brain. MethodsBrain tissues from adult humans with (n = 22) and without HIV-1 (n = 11) infection as well as adult nonhuman primates (NHPs) with (n = 11) and without (n = 4) SIVmac251 infection were investigated by RT-PCR/ddPCR, immunofluorescence and western blotting. Studies of viral infectivity, host immune gene expression and viability were performed in primary human neural cells. FindingsAmong NHPs, SIV DNA quantitation in brain showed increased levels among animals with SIV encephalitis (n = 5) that was associated with bacterial genomic copy number as well as CCR5 and CASP1 expression in brain. Microbial DnaK and peptidoglycan were immunodetected in brains from uninfected and SIV-infected animals, chiefly in glial cells. Human microglia infected by HIV-1 showed increased p24 production after exposure to peptidoglycan that was associated CCR5 induction. HIV-1 Vpr application to human neurons followed by peptidoglycan exposure resulted in reduced mitochondrial function and diminished beta-III tubulin expression. In human brains, bacterial genome copies (250–550 copies/gm of tissue), were correlated with increased bacterial rRNA and GroEL transcript levels in patients with HIV-associated neurocognitive disorders (HAND). Glial cells displayed microbial GroEL and peptidoglycan immunoreactivity accompanied by CCR5 induction in brains from patients with HAND. InterpretationIncreased microbial genomes and proteins were evident in brain tissues from lentivirus-infected humans and animals and associated with neurological disease. Microbial molecule translocation into the brain might exacerbate neuroinflammatory disease severity and represent a driver of lentivirus-associated brain disease.

Analysis results of 579 cases of genomic copy number variation sequencing of pregnant women in prenatal diagnosis.

This study explored the usefulness of genomic copy number variation sequencing (CNV-Seq) in the prenatal diagnosis of pregnant women. Based on prenatal diagnostic indications, CNV-Seq analysis was done in the samples from the 579 pregnant women of the 7 subgroups that included advanced maternal age (group A), high risk noninvasive prenatal test (NIPT) (group B), high risk Down's (Group C), abnormal ultrasound findings (Group D), adverse pregnancy history (Group E), chromosome abnormalities in couples (Group F), and the mixed group (Group G). A total of 57 (9.84%) cases have abnormal CNV-Seq results. Among them, 21 cases were aneuploid chromosomal number abnormalities (3.63%, 21/579), and 36 cases were CNV abnormalities (6.22%, 36/579), including 7 cases of pathogenic copy number alteration (pCNA) (1.21%, 7/579) and 29 cases variants of uncertain significance (VUS) (5.01%, 29/579). The total detection rates of abnormal CNV-Seq in Group G and Group B were 20.27% (15/74) and 15.91% (14/88), which were significantly higher than those in other groups (p < 0.05). Among 36 cases of abnormal CNV-Seq, 7 cases were chromosome fragment deletion or duplication, which were pathogenic CNV, and some rare chromosomal diseases were detected. Patients with a high risk of NIPT or multiple indications of prenatal diagnosis are highly suspected of chromosomal diseases. CNV-Seq is a useful tool for detecting chromosome abnormalities for prenatal diagnosis of pregnant women more accurately and provides more comprehensive information for prenatal diagnosis to reduce birth defects.

A general approach to identify low-frequency variants within influenza samples collected during routine surveillance.

Influenza viruses exhibit considerable diversity between hosts. Additionally, different quasispecies can be found within the same host. High-throughput sequencing technologies can be used to sequence a patient-derived virus population at sufficient depths to identify low-frequency variants (LFV) present in a quasispecies, but many challenges remain for reliable LFV detection because of experimental errors introduced during sample preparation and sequencing. High genomic copy numbers and extensive sequencing depths are required to differentiate false positive from real LFV, especially at low allelic frequencies (AFs). This study proposes a general approach for identifying LFV in patient-derived samples obtained during routine surveillance. Firstly, validated thresholds were determined for LFV detection, whilst balancing both the cost and feasibility of reliable LFV detection in clinical samples. Using a genetically well-defined population of influenza A viruses, thresholds of at least 104 genomes per microlitre and AF of ≥5 % were established as detection limits. Secondly, a subset of 59 retained influenza A (H3N2) samples from the 2016-2017 Belgian influenza season was composed. Thirdly, as a proof of concept for the added value of LFV for routine influenza monitoring, potential associations between patient data and whole genome sequencing data were investigated. A significant association was found between a high prevalence of LFV and disease severity. This study provides a general methodology for influenza LFV detection, which can also be adopted by other national influenza reference centres and for other viruses such as SARS-CoV-2. Additionally, this study suggests that the current relevance of LFV for routine influenza surveillance programmes might be undervalued.

280. MULTI-OMIC FEATURES OF OESOPHAGEAL ADENOCARCINOMA PATIENTS PRE-TREATED WITH PREOPERATIVE NEOADJUVANT THERAPY

Abstract The incidence of oesophageal adenocarcinoma (OAC) is rising in Western countries, with a 5-year overall survival (OS) rate of 14%. Curative treatment based on oesophagectomy is only suitable for ~50% of patients due to late-stage diagnosis. While the addition of preoperative chemotherapy or chemoradiotherapy has improved OS in OAC patients, little is known about the molecular basis of treatment response and patient outcomes. We investigated multi-omics data including whole-genome sequencing, RNA sequencing, methylation profiles and immunohistochemistry from 115 OAC patients mostly from DOCTOR phase II clinical trial that utilized neoadjuvant therapy (ANZCTR-ACTRN12609000665235). We identified genomic features associated with poor OS, such as the APOBEC mutational signature. We also showed that positron emission tomography non-responders have more sub-clonal genomic copy number alterations. RNA sequencing and methylation profiles suggested four immune clusters associated with OS and progression-free survival. The immune-suppressed cluster was associated with worse survival and epithelial-mesenchymal transition signature and enriched with myeloid-derived cells. The immune hot cluster was associated with better survival and enriched with T-cells, myeloid-derived cells, interferon-gamma and alpha responses, and immune markers such as CCL5, CD8A, and NKG7. We investigated multi-omic features of OAC tumours from patients who were part of phase II clinical trial. We discovered prognostic features such as mutational signatures and complex genomic events. We identified distinct immune clusters associated with patient outcomes and the potential of immunotherapy for select clusters. We characterized molecular features of OAC patients which has the potential to predict responses to neoadjuvant therapy and develop better-selected therapy, monotherapy, or combination therapy in the future.

Vaccination Decreases the Infectious Viral Load of Delta Variant SARS-CoV-2 in Asymptomatic Patients.

The Delta variant of SARS-CoV-2 has caused many breakthrough infections in fully vaccinated individuals. While vaccine status did not generally impact the number of viral RNA genome copies in nasopharyngeal swabs of breakthrough patients, as measured by Ct values, it has been previously found to decrease the infectious viral load in symptomatic patients. We quantified the viral RNA, infectious virus, and anti-spike IgA in nasopharyngeal swabs collected from individuals asymptomatically infected with the Delta variant of SARS-CoV-2. Vaccination decreased the infectious viral load, but not the amount of viral RNA. Furthermore, vaccinees with asymptomatic infections had significantly higher levels of anti-spike IgA in their nasal secretions compared to unvaccinated individuals with asymptomatic infections. Thus, vaccination may decrease the transmission risk of Delta, and perhaps other variants, despite not affecting the amount of viral RNA measured in nasopharyngeal swabs.

Copy number variation association studies for sheep tail-relevant traits in Hulunbuir sheep.

Tail type is an important trait that influences meat quality and consumer purchasing attitudes. As a novel genetic marker, the study of genomic copy number variations (CNVs) provides a new research method to study the genetic mechanisms underlying trait formation. In the present paper, we conducted CNV-based association studies for sheep tail type and growth traits in Hulunbuir sheep. A total of 167 CNVs and 288 individuals were analyzed at the marker and individual level, respectively. Association studies of 10 traits were carried out using two mixed linear models. We found that three, one and one CNV loci were significantly associated with tail type, tail length and tail fat weight, respectively, which together contained a total of 52 candidate genes. Out of these genes, five have been documented to be associated with fat metabolism in sheep. Our findings provide a reference for further studies on fat deposition in sheep as well as a theoretical basis for breeding Hulunbuir sheep with the required tail type.

Multi-omics subtyping of hepatocellular carcinoma patients using a Bayesian network mixture model.

Comprehensive molecular characterization of cancer subtypes is essential for predicting clinical outcomes and searching for personalized treatments. We present bnClustOmics, a statistical model and computational tool for multi-omics unsupervised clustering, which serves a dual purpose: Clustering patient samples based on a Bayesian network mixture model and learning the networks of omics variables representing these clusters. The discovered networks encode interactions among all omics variables and provide a molecular characterization of each patient subgroup. We conducted simulation studies that demonstrated the advantages of our approach compared to other clustering methods in the case where the generative model is a mixture of Bayesian networks. We applied bnClustOmics to a hepatocellular carcinoma (HCC) dataset comprising genome (mutation and copy number), transcriptome, proteome, and phosphoproteome data. We identified three main HCC subtypes together with molecular characteristics, some of which are associated with survival even when adjusting for the clinical stage. Cluster-specific networks shed light on the links between genotypes and molecular phenotypes of samples within their respective clusters and suggest targets for personalized treatments.

Clinicopathological and genomic copy number variation analysis in nodular hidradenoma and hidradenocarcinoma with focus on prognostically important features

Nodular hidradenoma is a cutaneous adnexal tumor of sweat gland origin, characterized by its diverse but overlapping histomorphologic features with other skin tumors. In addition, distinction of benign hidradenoma and its malignant counterpart hidradenocarcinoma can be challenging, especially in prognostic prediction. We retrospectively reviewed pathological features of 29 cases, including benign nodular hidradenoma (n=17) and hidradenocarcinoma (n=12), with clinical follow-up ranging from 18 to 216 months. Genomic copy number variation (CNV) was studied in selected cases (n=18) by single nucleotide polymorphismmicroarray. None of the benign hidradenomas (0/17) or low-grade hidradenocarcinomas (0/6) had recurrence or metastasis after complete excision, whereas all 6 high-grade hidradenocarcinomas (6/6) showed locally destructive disease, recurrence, or local metastases. In benign hidradenomas, CNV abnormality was absent in all clear cell hidradenomas (0/5) but was detected in a considerable portion of poroid hidradenoma (3/5), with number of abnormalities ranging 2, 4, and 9. In malignant cases, regardless of morphological classification, both low-grade hidradenocarcinomas demonstrated limited CNV abnormalities in 2 areas (2/2), whereas all high-grade hidradenocarcinomas contained 8 or more CNV abnormalities (6/6). No disease-associated death was recorded in the cohort except one case was lost to follow-up after the development of metastatic disease. Overall, the findings support that genomic CNV abnormalities may serve as a sensitivebut less specific tool in detecting malignancy in these tumors, and potentially have a role in predicting clinical behavior particularly in the tumors of nonporoid morphology.

Expanding the Phenotype of 8p23.1 Deletion Syndrome: Eight New Cases Resembling the Clinical Spectrum of 22q11.2 Microdeletion

To report the effectiveness of early molecular diagnosis in the clinical management of rare diseases, presenting 8 patients with 8p23.1DS who have clinical features that overlap the phenotypic spectrum of 22q11.2DS. This report is part of a previous study that aims to provide a precocious molecular diagnosis of the 22q11.2 deletion syndrome in 118 infants with congenital heart disease. To confirm the clinical diagnosis, patients underwent comparative genomic screening by the multiplex ligation-dependent probe amplification (MLPA) assay with the SALSA MLPA probemix kits P064-B2, P036-E1, P070-B2, P356-A1, and P250- B1. Subsequently, the patients performed the genomic microarray using the Infinium CytoSNP-850K BeadChip to confirm the deletion, determine the breakpoints of the deletion, and search for genomic copy number variations. MLPA performed with 3 different kits revealed the 8p23.1 typical deletion involving the PPP1R3B, MSRA, and GATA4 genes in the 5 patients. The array analysis was performed on these 5 patients and 3 other patients (8 patients) who also had clinical suspicion of 22q11 deletion (8 patients) allowed a precise definition of the breakpoints and excluded other genomic abnormalities. Cytogenomic screening was efficient in establishing a differential diagnosis and ruling out the presence of other concomitant syndromes. The clinical picture of the 8p23.1 deletion syndrome is challenging; however, cytogenomic tools can provide an exact diagnosis and help to clarify the genotype-phenotype complexity of these patients. Our reports underline the importance of early diagnosis and clinical follow-up of microdeletion syndromes.

1661MO Genomic somatic copy number alterations drive adaptive tumor immune suppression and primary resistance to anti-PD1 + anti-angiogenics in pleural mesothelioma

In patients with advanced pleural malignant mesothelioma (MM), Pembrolizumab [P] in combination with Nintedanib [N] synergize with a response rate of 24% (PEMBIB trial initial results reported at ESMO Congress 2021), superior to [P] alone (8% in MM in the KN158 trial). Here we report new data from the biomarker analysis performed in this trial (NCT02856425). 30 MM patients were treated with [N] (150mg bid) & [P] (200mg IV Q3W). Fresh tumor biopsies were collected in media and immediately processed at baseline and at C2D1 for measuring proteins released in the biopsy supernatant and immune cell infiltrates (by Flow Cytometry). Frozen and fixed tumor biopsies were also analyzed by WES, FISH, RNAseq and IHC. Circulating cytokines & immune cells were measured on plasma and fresh whole blood at baseline and on-treatment. At 6 months, patients remaining in stable disease or objective tumor response per RECIST1.1 were categorized as having Durable Clinical Benefit (DCB). All differences reported here are statistically significant with a p value at least <0,05. Higher PD-L1 on tumor cells, higher tumor CD45+, CD3+ & CD8+ T-cells characterized the DCB group at baseline. Unexpectedly, active recruitment of CD8+ T-cells, with higher gene expression of EOMES, T-BET, IFNg, & GzmB, occurred in No DCB tumors upon treatment. At C2D1, same levels of tumor infiltrating CD8+ T-cells were found in DCB & No DCB tumors. This tumor T-cell recruitment occurred in a context of high blood and tumor CXCL9 increases upon [N]+[P] treatment. However, No DCB patients presented more IL6, VEGF, IL8, in their blood & tumor secretome and they increased their tumor content in CD4+ CD25+ Tregs compared to DCB patients upon treatment. Tumor somatic copy number alterations (SCNA) were higher in tumors with no DCB and correlated with higher plasmatic IL-6 rate levels in MM patients. MM primary resistance to anti-PD1+antiangiogenics is not due to a lack of CD8+ T-cell tumor response but to an adaptive secretion of IL6 by tumors, driven by genomic SCNA, leading to VEGF and IL8 secretion, and recruitment of tumor infiltrating Tregs.

Analysis of antitumor immunity in mouse glioma microenvironment through intercellular communication.

This study aimed to map the antitumor immunity in the glioma microenvironment by analyzing intercellular communication. The single-cell RNA-sequencing (scRNA-Seq) data were obtained from fresh mouse gliomas. Tumor cells were inferred by estimating genomic copy number profiles. CellMarker database was used to identify cell types. Intercellular communication was inferred using CellChat. Flow cytometry was used to detect the effect of microglia or stroma-educated monocytes on CD4+ T cell proliferation. Mouse glioma contained at least eight cell populations, and T cells were the only infiltrating immunocytes. Whether in signal outgoing or signal incoming, intercellular communication could be divided into four patterns by which cell populations in the tumor microenvironment (TME) cooperate with each other. By analyzing the complex communication between brain cell populations and infiltrating T cells in TME, we found that the brain cell populations used 25 signaling pathways to connect to T cells, and T cells used 21 signaling pathways to connect to brain cell populations. We also found that microglia from normal mice and brain stroma-educated monocytes exhibited immunosuppressive activity against CD4+ T cell proliferation. We described the previously underestimated complex communication between infiltrating T cells and brain cell populations. Our data suggest that the tolerogenic property of glioma TME is related to the immune privilege of CNS.

576P Genome instability index used for the optimization of molecular classification of endometrial carcinoma

The molecular classification of endometrial carcinoma (EC) has divided patients into four distinct prognostic subtypes based on the genomic abnormalities. However, in clinical application, the genomic copy number index lacks a simple and accurate detection method, and the method of using p53 immunohistochemistry or TP53 mutation as a surrogate for copy number status needs to be optimized. Here, we explored the application of genome instability (GI) index in molecular classification of EC. We assessed a retrospective cohort of EC from Beijing Cancer Hospital between 2011 and 2020. Genomic profiling of tumor tissues from EC patients was sequenced by OncoScreen Plus Panel of 520 cancer-related genes, and tumor immune microenvironment was evaluated using multiplex immunofluorescence staining. GI index was calculated as the sum of loss of heterozygosity, telomeric allelic imbalance, and large-scale state transitions scores. 143 patients were enrolled in this study, with the median follow-up time of 38.77 months. Molecular analysis firstly identified four different prognostic subgroups: POLE mutations (mut), MSI-H, TP53 wildtype, and TP53 mut. We also divided patients into two groups based on GI index: GI-high and GI-low, and patients with GI-high were found in three subgroups except for POLE mut. Further analysis showed that GI status had a more important effect on prognosis than TP53 mutation. Therefore, we proposed a three-group molecular classification based on POLE mutation and GI index: POLE mut (n=9, 6.5%), GI-low (n=116, 82.9%), and GI-high (n=15, 10.7%). Patients with GI-high had much poorer survival than those with GI-low [disease-free survival: hazard ratio (HR), 17.87, 95% CI, 6.67-50.00, P < 0.001; overall survival: HR, 16.75, 95% CI, 4.76-50.00, P < 0.001]. The application in TCGA-UCEC cohort containing 481 EC patients confirmed that this new method worked well for risk stratification of EC. Further analysis showed that patients in GI-high group had higher tumor grade, aneuploidy and lower T cell infiltrations. The one-stop method of GI index combined with POLE mutation offers more accurate and stable disease risk stratification and demonstrates the potential for guiding therapy for EC patients.

Cuproptosis scoring model predicts overall survival and assists in immunotherapeutic decision making in pancreatic carcinoma.

Background: Cuproptosis is a newly identified form of non-apoptotic cell death that is associated with the progression and treatment responses in pancreatic adenocarcinoma (PAAD). However, its impact on oncology and tumor microenvironment (TME) remains unclear. Methods: Hub genes were identified using least absolute shrinkage and selection operator (LASSO) Cox regression for 25 newly reported cuproptosis-related regulators and subjected to stepwise regression to obtain cuproptosis-related score (CuRS). Additionally, the clinical significance, functional status, role on TME, and genomic variation of CuRS were further examined systematically. Results: A CuRS model incorporating TRAF2, TRADD, USP21, FAS, MLKL, TNFRSF10B, MAPK8, TRAF5, and RIPK3 was developed. The stability and accuracy of this risk model as an independent prognostic factor for PAAD were confirmed in the training and external validation cohorts. Patients in the high-CuRS group had “cold” tumors with active tumor proliferation and immunosuppression, whereas those in the low-CuRS group comprised “hot” tumors with active immune function and cell killing capacity. Additionally, patients in the high-CuRS group carried fewer genomic copy number variations (CNVs) and greater somatic mutations. Furthermore, patients in the low- and high-CuRS groups exhibited increased sensitivity to immunotherapy and chemotherapy, respectively. Conclusion: We developed and validated a robust CuRS model based on cuproptosis to assess patients’ prognoses and guide clinical decision-making. Overall, the findings of this study are expected to contribute to the comprehensive understanding of cuproptosis and facilitate precise treatment of PAAD.

Deletion of the CD2v Gene from the Genome of ASFV-Kenya-IX-1033 Partially Reduces Virulence and Induces Protection in Pigs.

Infection of pigs with the African swine fever virus (ASFV) leads to a devastating hemorrhagic disease with a high mortality of up to 100%. In this study, a CD2v gene deletion was introduced to a genotype IX virus from East Africa, ASFV-Kenya-IX-1033 (ASFV-Kenya-IX-1033-∆CD2v), to investigate whether this deletion led to reduced virulence in domestic pigs and to see if inoculation with this LA-ASFV could induce protective immunity against parental virus challenge. All pigs inoculated with ASFV-Kenya-IX-1033-ΔCD2v survived inoculation but presented with fever, reduced appetite and lethargy. ASFV genomic copies were detected in only one animal at one time point. Seven out of eight animals survived subsequent challenge with the pathogenic parental strain (87.5%) but had mild to moderate clinical symptoms and had a gross pathology compatible with chronic ASFV infection. All mock-immunised animals developed acute ASF upon challenge with ASFV-Kenya-IX-1033 and were euthanised upon meeting the humane endpoint criteria. ASFV genome copy numbers after challenge were similar in the two groups. ASFV-Kenya-IX-1033-∆CD2v is therefore a useful tool to investigate the development of immunity to ASFV genotype IX, but safety concerns preclude its use as a candidate vaccine without further attenuation.

Four-copy number alteration (CNA)-related lncRNA prognostic signature for liver cancer

The objective of this study was to identify CNA-related lncRNAs that can better evaluate the prognosis of patients with liver cancer. Prognostic molecular subtypes were identified, followed by tumor mutation and differential expression analyses. Genomic copy number anomalies and their association with lncRNAs were also evaluated. A risk model was built based on lncRNAs, as well as a nomogram, and the differences in the tumor immune microenvironment and drug sensitivity between the High_ and Low_risk groups were compared. Weighted gene co-expression network analysis was used to identify modules with significant enrichment in prognostic-related lncRNAs. In total, two subtypes were identified, TP53 and CTNNB1 were common high-frequency mutated genes in the two subtypes. A total of 8,372 differentially expressed (DE) mRNAs and 798 DElncRNAs were identified between cluster1 and cluster2. In addition, a four-lncRNA signature was constructed, and statistically significant differences between the Low_ and High_risk groups were found in terms of CD8 T cells, resting memory CD4 T cells, etc. Enrichment analysis showed that prognostic-related lncRNAs were involved in the cell cycle, p53 signaling pathway, non-alcoholic fatty liver disease, etc. A prognostic prediction signature, based on four-CNA-related lncRNAs, could contribute to a more accurate prognosis of patients with liver cancer.