Transcriptomic clustering of critically ill COVID-19 patients

Infections caused by SARS-CoV-2 may cause a severe disease, termed COVID-19, with significant mortality. Host responses to this infection, mainly in terms of systemic inflammation, have emerged as key pathogenetic mechanisms, and their modulation is the only therapeutic strategy that has shown a mortality benefit. Herein, we used peripheral blood transcriptomes of critically-ill COVID-19 patients obtained at admission in an Intensive Care Unit (ICU), to identify two transcriptomic clusters characterized by expression of either interferon-related or immune checkpoint genes, respectively. These profiles have different ICU outcome, in spite of no major clinical differences at ICU admission. A transcriptomic signature was used to identify these clusters in an external validation cohort, yielding similar results. These findings reveal different underlying pathogenetic mechanisms and illustrate the potential of transcriptomics to identify patient endotypes in severe COVID-19, aimed to ultimately personalize their therapies.

IFIH1 rs1990760 variants, systemic inflammation and outcome in critically-ill COVID-19 patients

RationaleOutcomes in patients with severe SARS-CoV-2 infection (COVID-19) are conditioned by virus clearance and regulation of inflammation. Variants in IFIH1, a gene coding the cytoplasmatic RNA sensor MDA5, regulate the response to viral infections. ObjectiveTo characterize the impact of IFIH1 rs199076 variants on host response and outcomes after severe COVID-19. MethodsPatients admitted to an intensive care unit (ICU) with confirmed COVID-19 were prospectively studied and rs1990760 variants determined. Peripheral blood gene expression, cell populations and immune mediators were measured. Peripheral blood mononuclear cells from healthy volunteers were exposed to an MDA5 agonist and dexamethasone ex-vivo, and changes in gene expression assessed. ICU discharge and hospital death were modelled using rs1990760 variants and dexamethasone as factors in this cohort and in-silico clinical trials. Measurements and Main Results227 patients were studied. Patients with the IFIH1 rs1990760 TT variant showed a lower expression of inflammation-related pathways, an anti-inflammatory cell profile and lower concentrations of pro-inflammatory mediators. Cells with TT variant exposed to a MDA5 agonist showed an increase in IL6 expression after dexamethasone treatment. All patients with the TT variant not treated with steroids (N=14) survived their ICU stay (HR 2.49, 95% confidence interval 1.29-4.79). Patients with a TT variant treated with dexamethasone (N=50) showed an increased hospital mortality (HR 2.19, 95% confidence interval 1.01-4.87) and serum IL-6. In-silico clinical trials supported these findings. ConclusionsCOVID-19 patients with the IFIH1 rs1990760 TT variant show an attenuated inflammatory response and better outcomes. Dexamethasone may reverse this anti-inflammatory phenotype.

The CCR5-delta32 variant might explain part of the association between COVID-19 and the chemokine-receptor gene cluster

A polymorphism in the LZTFL1 gene located in the chemokine-receptor gene cluster (chromosome 3p) has been associated with the risk of developing COVID-19. The chemokine receptor-5 (CCR5) maps to this region, and the common 32 bp deletion variant ({Delta}32) has been associated with the extent of inflammatory disease and the outcome in several viral diseases. Several studies have also suggested that the pharmacological targeting of CCR5 could reduce the impact of SARS-CoV-2 infection and the severity of COVID-19. We sought to investigate whether this polymorphism was associated with the risk of moderate-severe COVID-19. We genotyped 294 patients who required hospitalization due to COVID-19 (85 were severe cases) and 460 controls. We found a significantly lower frequency of CCR5-{Delta}32 among the COVID-19 patients (0.10 vs 0.18 in controls; p=0.002, OR=0.48, 95%CI=0.29-0.76). The difference was mainly due to the reduced frequency of CCR5-{Delta}32 carriers in the severe, significantly lower than in the non-severe patients (p=0.036). Of note, we did not find deletion-homozygotes among the patients compared to 1% among controls. We also confirmed the association between a LZTFL1 variant and COVID-19. Our study points to CCR5 as a promising target for treatment of COVID-19, but requires validation in additional large cohorts. In confirmed by others, the genetic analysis of CCR5-variants (such as {Delta}32) might help to identify patients with a higher susceptibility to severe COVID-19.

Angiotensin-converting enzyme (ACE1, ACE2) gene variants are associated with COVID19 severity depending on the hypertension status.

BackgroundThe Angiotensin system is implicated in the pathogenesis of COVID19. First, ACE2 is the cellular receptor for SARS-COv-2, and expression of the ACE2 gene could regulate the individuals susceptibility to infection. In addition, the balance between ACE1 and ACE activity has been implicated in the pathogenesis of respiratory diseases and could play a role in the severity of COVID19. Functional ACE1/ACE2 gene polymorphisms have been associated with the risk of cardiovascular and pulmonary diseases, and could thus also contribute to the outcome of COVID19. MethodsWe studied 204 COVID19 patients (137 non-severe and 67severe-ICU cases) and 536 age-matched controls. The ACE1 insertion/deletion and ACE2rs2285666 polymorphism were determined. Variables frequencies were compared between the groups by logistic regression. We also sequenced the ACE2 coding nucleotides in a group of patients. ResultsSevere COVID19 was associated with hypertension male gender (p<0.001), hypertension (p=0.006), hypercholesterolaemia (p=0.046), and the ACE1-DD genotype (p=0.049). In the multiple logistic regression hypertension (p=0.02, OR=2.26, 95%CI=1.12-4.63) and male gender (p=0.002; OR=3.15, 95%CI=1.56-6.66) remained as independent significant predictors of severity. The ACE2 polymorphism was not associated with the disease outcome. The ACE2 sequencing showed no coding sequence variants that could explain an increased risk of developing COVID19. ConclusionsAdverse outcome of COVID19 was associated with male gender, hypertension, hypercholesterolemia and the ACE1 genotype. The ACE1-I/D was a significant risk factor for severe COVID19, but the effect was dependent on the hypertensive status.

Capillary Electrophoresis of PCR fragments with labelled primers for testing the SARS-Cov-2

BackgroundDue to the huge demand for SARS-Cov-2 determination, alternatives to the standard qtPCR tests are potentially useful for increasing the number of samples screened. Our aim was to develop a direct fluorescent PCR capillary-electrophoresis detection of the viral genome. We validated this approach on several SARS-Cov-2 positive and negative samples. Study designWe isolated the naso-pharingeal RNA from 20 positive and 10 negative samples. The cDNA was synthesised and two fragments of the SARS-Cov-2 were amplified. One of the primers for each pair was 5-end fluorochrome labelled. The amplifications were subjected to capillary electrophoresis in ABI3130 sequencers to visualize the fluorescent peaks. ResultsThe two SARS-Cov-2 fragments were successfully amplified in the positive samples, while the negative samples did not render fluorescent peaks. ConclusionWe describe and alternative method to identify the SARS-Cov-2 genome that could be scaled to the analysis of approximately 100 samples in less than 5 hours. By combining a standard PCR with capillary electrophoresis our approach would overcome the limits imposed to many labs by the qtPCR (lack of reactive and real-time PCR equipment) and increase the testing capacity.

Nefropatía familiar hiperuricemiante: nueva mutación familiar del gen de la uromodulina / Familial hyperuricaemic nephropathy: New mutation in uromodulin gen

No disponible

Pharmacogenetics of tacrolimus after renal transplantation: analysis of polymorphisms in genes encoding 16 drug metabolizing enzymes.

BACKGROUND: Tacrolimus (Tac) is an immunosuppressive drug used to prevent post-transplant (PT) organ rejection. Continuous Tac monitoring is necessary to adjust the dose and prevent toxicity or rejection. Tac is metabolized by cytochrome-P450 (CYP) enzymes, and variation at the CYP and other drug metabolizing enzymes could influence Tac bio-availability and dose requirements. Our aim was to define the effect of DNA variants at 16 drug metabolising enzymes on Tac dose in patients with kidney transplants. METHODS: The REDINREN Pharmacogenetics Project was a multicenter study designed to evaluate the effect of DNA polymorphisms on Tac dose requirements. A total of 200 patients who received a first cadaveric kidney and Tac as primary immunosuppressive drug were genotyped for 96 DNA polymorphisms on 16 genes. Significant associations were further replicated in a second group of 200 patients. The Tac daily dose was adjusted to achieve a blood concentration of 10-15 ng/mL in the period 0-3 months PT, and 5-10 ng/mL thereafter. The dose of tacrolimus dose and blood concentrations were compared between genotypes at 1 week, 6 months, and 1 year PT. RESULTS: The CYP3A5 genotype (SNP rs776746) was the strongest predictor of Tac dose requirements. Patients who were CYP3A5*3*3 (CYP3A5 non-expressors) received significantly higher Tac dose at 1 week, 6 months, and 1 year PT (p<0.0001). At 1 week, 41% of the CYP3A5 non-expressors achieved target blood concentrations compared to 26% of the CYP3A5 expressors (p=0.007). We also found a significant effect of CYP3A4 genotype (SNP rs2740574) on Tac dose requirements in patients who were CYP3A5 non-expressors. None of the other polymorphisms were related to Tac dose requirements or modified the effect of the CYP3A5 genotype. CONCLUSIONS: rs776746 (CYP3A5) and rs2740574 (CYP3A4) were the only SNPs associated with Tac dosage. The genotyping of these polymorphisms could be a useful pharmacogenetic tool to determine the Tac dose immediately after transplantation.

Association between genetic variation in the Y chromosome and hypertension in myocardial infarction patients.

A single nucleotide polymorphism (SNP) in chromosome Y has been associated with blood pressure. In men, the risk of suffering from cardiovascular diseases, including coronary artery disease, could be influenced by one or more loci on chromosome Y. We genotyped 208 men who had suffered an early episode of myocardial infarction (MI) (< or =55 years) and 178 healthy control men for two Y-polymorphisms (a HindIII polymorphism in an alphoid satellite in the centromeric non-recombining region and the -2627 T/C in the SRY gene). Frequencies were compared through a chi(2)-test. Frequencies for the two polymorphisms did not differ between patients and controls. The alphoid-HindIII polymorphism was not related to blood pressures in our population (HindIII+: diastolic, 80 +/- 2; systolic, 129 +/- 5. HindIII-: diastolic, 80 +/- 2; systolic, 128 +/- 3). Seventy-six patients (37%) were hypertensives and had a significantly higher frequency of the HindIII+ allele compared to the normotensive patients (46 and 26%, respectively; P = 0.028). According to our data, the alphoid-HindIII polymorphism in chromosome Y was not associated with differences in blood pressure in men from Asturias (Northern Spain). However, the HindIII+ allele increased the risk of suffering an early episode of MI among hypertensives.

Single-nucleotide polymorphisms in the promoter region of the PARKIN gene and Parkinson's disease.

Mutations in the PARKIN gene have been identified in families with recessively inherited Parkinson disease (PD). Common DNA-polymorphisms at the PARKIN gene could contribute to the risk for PD in the general population. Here we searched for DNA-polymorphisms in the PARKIN promoter. We found two single nucleotide polymorphisms (-324 A/G and -797 A/G). In order to analyse the association of PD with these and two previously described polymorphisms (1281 G/A, Asp394Asn, and 601 G/A, Ser167Asn) we genotyped 105 patients and 150 healthy controls. Allele and genotype frequencies for the four polymorphisms did not differ between patients and controls, or between patients with an early-onset (< or =40 years; n = 20) and a late-onset (>40 years; n = 85). According to our data, the genetic variation at the PARKIN gene (including promoter polymorphisms) did not contribute to the risk of developing PD in the general population.