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GENOMEWIDEASSOCIATIONSTUDY OFSEVERECOVID-19 WITHRESPIRATORYFAILURE

Estudo de associação ampla do genoma com Covid-19

grave e insuficiência respiratóriaSeminar 3 - Genetics

Seminário 3 - Genética

Ana Clara Germano TostesDiego Ferreira CaldasGabriel Garcia SchmittHenrique LarssenKamilla Dolgoff Santana

Profº Jeremy Andrew SquireThe Severe Covid-19 GWAS GroupFrom the NEW ENGLAND JOURNAL of MEDICINE

Summary

◦ Abstract

◦ Background

◦ Methods

◦ Results

◦ Conclusions

◦ Methods

◦ Study participants and recruitment

◦ Ethics committee aproval

◦ Sample processing, genotyping and imputation

◦ Statistical analysis

◦ Results

◦ Patients, genotyping and quality control

◦ Genomewide association analysis

◦ Chromosome 3P21.31

◦ ABO Locus

◦ HLA Analysis

◦ Discussion

AbstractResumo

BACKGROUND METHODS RESULTS CONCLUSIONS

Contexto Métodos Resultados Conclusões

Presenter: Kamilla Dolgoff Santana

BackgroundContexto

Severe acuterespiratory syndrome

coronavirus 2 wasdiscovered in Wuhan,

China, in late 2019;

Covid-19 rapidlyevolved into a global

pandemic;

More than 8.03 millionconfirmed cases

worldwide, with total deaths exceeding 435

thousand;

The relative role ofclinical risk factors in

determining theseverity of Covid-19 has

not yet been clarified.


◦ It was performed a genomewide association study

(GWAS) in an attempt to delineate host genetic factors

contributing to severe Covid-19 with respiratory failure.

◦ Since there is considerable variation in disease behavior

among patients infected with severe acute respiratory

syndrome coronavirus 2, genomewide association

analysis may allow for the identification of potential

genetic factors involved in the development of Covid-19.

Methods; Results and ConclusionsMétodos; Resultados e Conclusões

◦ 1980 patients with Covid-19 and

severe disease at seven hospitals

in the Italian and Spanish

epicenters of the SARS-CoV-2

pandemic;

◦ 8,582,968 singlenucleotide

polymorphisms analyzed and a

meta-analysis of the two case-

control panels conducted.


◦ Higher risk in blood group A than

in other blood groups;

◦ Protective effect in blood group

O as compared with other blood

groups;

◦ Cross-replicating associations

with rs11385942 at locus 3p21.31

and with rs657152 at locus 9q34.2

detected:

◦ Locus 3p21.31: the association

signal spanned the genes

SLC6A20, LZTFLl, CCR9,

FYCOl, CXCR6 and XCRl;

◦ Locus 9q34.2: coincided with

the ABO blood group locus.

Confirmed a potential involvementof the ABO blood-group system.

MethodsMétodos

Study participants and recruitment

Ethics committee aproval

Sample processing, genotyping and imputation

Statistical analysis

Participantes do estudo e recrutamento

Processamento de amostras, genotipagem e imputação

Análises estatísticas

Aprovação do Comitê de Ética

Presenter: Diego Ferreira Caldas

Study participants and recruitment

◦ 7 hospitals in four cities in pandemic epicenters in Italy and Spain

◦ 1980 patients with severe Covid-19 (hospitalization, respiratory failure and confirmed SARS-CoV-2 RNA

PCR test)

◦ Respiratory failure defined as use of oxygen supplementation and mechanical ventilation, and severity

graded according to the maximum respiratory support received at any time (supplemental oxygen therapy,

noninvasive ventilatory support, invasive ventilatory support and extracorporeal membrane oxygenation)

◦ 2381 control participants from Italy and Spain (998 randomly selected blood donors and two previous

studies` control panels)


Ethics Committee Approval

◦ No additional project related procedures (clinically indicated venipunctures), anonimity

(minimal data set)

◦ Local differences due to local regulations or larger biobanking efforts

◦ Written informed consent obtained when possible, verbal and from next of kin in some

instances, exeption from informed consent for some severely ill patients afforded by local

regulations.


Sample Processing, Genotyping and Imputation

◦ DNA extraction with Chemagic 360

◦ Genotyping with Global Screening Array 2.0

◦ Single-nucleotide polymorphism (SNP) on genome build GRCh38 with Michigan Imputation

Server and haplotypes from the Trans-Omics for Precision Medicine program

◦ Exclusion of samples in quality control mostly due to population outliers, resulting in 835

patients and 1255 control participants from Italy and 775 patients and 950 control

participants from Spain.


Statistical Analysis

◦ Testing for phenotypic associations with allele dosage date separately for both Italianand Spanish case-control panels to account for imputation uncertainty

◦ Conducting two genomewide tests of association with adjustments to control forpotential population stratification and potential population stratification and age andsex

◦ Definition of 5x10^-8 threshold for joint P values for statistical significance

◦ Selection of three ABO SNPs to infer ABO blood type and calculated odds ratio

◦ Assessment of allelic distribution according to no mechanical ventilation compared tomechanical ventilation of any kind


ResultsResultados

Presenter: Gabriel Garcia Schmitt

Patients, genotyping and quality control

• Median age: 64 –69 • Sex prevalence: male• Most frequent: supplemental

oxygen only and ventilator

• High rate of hypertension• Low rate of coronary artery disease• Medium rate of diabetes


Genomewide association analysis

• P Values suggest 2 group of genes associated with severe Covid-19 and respiratory failure• rs11385942 ------ locus 3p21.31 ------ P = 1.15 x 10^-10• rs657152 --------- locus 9q34.2 ------- P = 4.95 x 10^-8


• P Value: indicates how likely a null hypothesys could happen, given the experimental data. The lower the P Value, the more likely is an alternative hypothesys .

SNP rs11385942, locus 3p21.31

◦ Comprises a group of six genes located in chromosome 3: SLC6A20, LZTFLI, CCR9, FYCOI,

CXCR6, and XCRl

◦ Risk allele GA of rs11385942:

◦ reduced expression of CXCR6 and increased expression of SLC6A20 and LZTFLI (in lung cells)

◦ higher frequency among pacients who needed ventilation than those who needed oxygen only

◦ homozygotic patients for the risk allele with respiratory failure were younger


Conclusion: the risk allele GA of rs11385942 is associated with more severe Covid-19

SNP rs657152, locus 9q34.2

◦ ABO blood group complex of genes

◦ Higher risk for blood group A

◦ Lower risk for blood group O

◦ Why?

◦ Antigen A interacts with viral protein S

◦ Anti-A antibodies limit viral interaction with host blood cell


OBS: HLA analysis (region in chromosome 6)Despite its important role in several viralinfections, the study did not find anysignificant SNPs related to Covid-19 severityor respiratory failure.

DISCUSSIONDISCUSSÃO

Presenters: Ana Clara Germano Tostes and Henrique Larssen

Locus 3p21.31

Several genes are associated

with this locus, such

as SLCGA20, LZTFL1,

CCR9, FYCO1, CXCR6 e

XCR1.

Principal candidates:

• SLCGA20

• CCR9

• CXCR6

Presenter: Henrique Larssen

The Covid-19 Host Genetics Consortium

◦ SLC6A20 functionally interacts with ACE2, the

receptor of the SARS-CoV-2 Spike protein that

is the key host gene for viral entry;

◦ CCR9 encodes the CC motif chemokine

receptor 9 , maturation of T cells;

◦ CXCR6 regulates the localization of resident

memory T (TRM) cells in the lung and

maintains a pool of airway TRM cells, critical

for cellular immunity against respiratory

pathogens.


Currently studies about locus 3p21.31

◦ To date, the locus with the most robust human

genetic association to COVID-19 severity is 3p21.31.

◦ The study identify SLC6A20 andCXCR6 as putative

causal genes that modulate COVID-19 risk


System ABO influence on Covid-19

Natural anti-A antibodies against SARS-CoV-2 infection:

◦ Group B or O has anti-A antibodies that slightly inhibits the entry of the virus into the host

cell.

ABO antigens in SARS-CoV-2:

◦ ABO antigens can trigger the virus spike protein attachment to the host cell receptor by

interactions with cell surface receptors, facilitating its entry, which occurs mainly with A

antigens.

Presenter: Ana Clara Germano Tostes

VonWillebrandfactor (vWF) andfactorVIII inCovid-19 risk:

◦ Coagulation factors such as factor VIII and von

Willebrand factor (vWF) increase the risk of thrombosis and

pulmonary embolism when in high levels in the blood plasma,

aggravating Covid-19;

◦ Pulmonary thromboembolism stops blood flow to the lungs,

making breathing difficult;

◦ In addition, VWF serves as a potential marker to predict the

endothelial activation that expresses the ACE2 receptor

(membrane protein through which the SARS-CoV-2 virus

enters);


https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8035048/

Impacts on blood donation Research difficulties

◦ The influence of the ABO system in

diseases such as Covid-19 can result

in a segregation of blood donors, with

greater appreciation for those with

type O, however, there are studies

that show that there is no

interference of donated blood in the

manifestation of the disease.


◦ Short period (recent studies);

◦ Minimum of clinical metadata was requested –extensive genotype-phenotype elaboration ofcurrent findings could not be conducted;

◦ Limited information about the SARS-CoV-2infection status in the control participants;

◦ Few restrictions imposed during the formationof control group;

◦ Factors present at lower levels in type O individuals that has lower risk of thromboembolism

◦ This hypothesis supports FVIII and VWF in the ABO (H) determinants also shows the associationof the vulnerability of COVID-19 to the ABO blood group

ConclusionConclusão

It is necessary further exploration of

current findings to their usefulness in

clinical risk profiling of patients and

toward a mechanicist understanding of the

underlying pathophysiology.


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