CC BY 4.0 · Glob Med Genet
DOI: 10.1055/s-0041-1731068
Original Article

Thromboses and Hemostasis Disorders Associated with Coronavirus Disease 2019: The Possible Causal Role of Cross-Reactivity and Immunological Imprinting

1  Department of Biosciences, Biotechnologies and Biopharmaceutics, University of Bari, Bari, Italy
› Author Affiliations
Funding None.
 

Abstract

By examining the issue of the thromboses and hemostasis disorders associated with severe acute respiratory syndrome-coronavirus-2 (SARS-CoV-2) through the lens of cross-reactivity, it was found that 60 pentapeptides are shared by SARS-CoV-2 spike glycoprotein (gp) and human proteins that— when altered, mutated, deficient or, however, improperly functioning— cause vascular diseases, thromboembolic complications, venous thrombosis, thrombocytopenia, coagulopathies, and bleeding, inter alia. The peptide commonality has a relevant immunological potential as almost all of the shared sequences are present in experimentally validated SARS-CoV-2 spike gp-derived epitopes, thus supporting the possibility of cross-reactions between the viral gp and the thromboses-related human proteins. Moreover, many of the shared peptide sequences are also present in pathogens to which individuals have previously been exposed following natural infection or vaccinal routes, and of which the immune system has stored imprint. Such an immunological memory might rapidly trigger anamnestic secondary cross-reactive responses of extreme affinity and avidity, in this way explaining the thromboembolic adverse events that can associate with SARS-CoV-2 infection or active immunization.


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Introduction

Clinical studies have shown that severe acute respiratory syndrome-coronavirus-2 (SARS-CoV-2) infection can lead to an increased incidence of disorders such as thrombosis, venous thrombosis, and pulmonary embolism.[1] [2] [3] A main conclusion of these studies is that, although it cannot be proven that the hypercoagulable state is a direct causative effect of SARS-CoV-2 infection, nonetheless it is apparent that patients with SARS-CoV-2 could have a predilection to the occurrence of thromboembolic events.[1]

However, currently there are no hypotheses or data that might suggest a molecular mechanism that relates to such SARS-CoV-2-related thromboembolic events. Searching for possible mechanisms, the present study analyzes the SARS-CoV-2 spike glycoprotein (gp) for peptide sharing, that is, molecular mimicry, with human proteins, alterations of which may cause thromboses and hemostasis diseases. The underlying scientific rationale is that peptides common to a pathogen and the human host may lead to autoimmune pathologies through cross-reactivity phenomena following pathogen infection.[4] [5] [6] The results indicate that several linear sequences shared between the SARS-CoV-2 spike gp and human proteins related to thromboembolic events can possibly generate pathogenic autoantibodies via cross-reactivity and immunologic imprinting phenomena, in this way leading to thromboses and hemostasis disorders.


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Materials and Methods

Peptide sharing between spike gp (NCBI, GenBank Protein Accession, ID: QHD43416.1) from SARS-CoV-2 and human proteins related to thromboses and hemostasis disorders was analyzed as previously detailed.[4] [5] [6] In brief, pentapeptides were used as sequence probes since a peptide grouping formed by five amino acid (aa) residues defines a minimal immune determinant that can (1) induce highly specific antibodies, and (2) determine antigen–antibody specific interaction.[7] [8] Human proteins linked to thromboses and hemostasis disorders were retrieved from UniProtKB database ( www.uniprot.org ).[9] Methodologically the spike gp primary sequence was dissected into pentapeptides offset by one residue (i.e., MFVFL, FVFLV, VFLVL, FLVLL, and so forth) and the resulting viral pentapeptides were analyzed for occurrences within the human proteins related to thromboses and hemostasis disorders. Then, the shared peptides were also controlled for occurrences in the pathogens Bordetella pertussis, Corynebacterium diphtheriae, Clostridium tetani, Haemophilus influenzae, and Neisseria meningitidis.

The immunological potential of the peptides shared between SARS-CoV-2 spike gp and thrombosis-related proteins was analyzed by searching the Immune Epitope DataBase (IEDB [ www.iedb.org/ ])[10] for immunoreactive SARS-CoV-2 spike gp-derived epitopes hosting the shared pentaptides.


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Results and Discussion

Peptide Sharing between SARS-CoV-2 Spike Glycoprotein and Thromboses-Related Human Proteins

[Table 1] shows that 60 minimal immune determinants are shared between SARS-CoV-2 spike gp and 44 human proteins that—when altered, mutated, deficient or, however, improperly functioning—may cause diseases that include coagulation disorders, bruising, bleeding, hemorrhages, retinal vessel occlusion, cerebral thrombosis, venous thrombosis, ischemic stroke, and thrombophilia, inter alia.

Table 1

Pentapeptide sharing between SARS-CoV-2 spike gp and human proteins linked when altered, mutated, or deficient to blood diseases

Shared peptides

Human proteins and associated functions/pathologies[a] [b]

References

MTKTS, NLLLQ

ADTRP (androgen-dependent TFPI-regulating protein)

Regulates the anticoagulant activity of the tissue factor pathway inhibitor, dysfunctions of which lead to vascular diseases

[11]

TQLPP, PRTFL

ALG12: Dol-P-Man: Man(7)GlcNAc(2)-PP-Dol α-1,6-mannosyltransferase

Psychomotor retardation, hypotonia, coagulation disorders, and immunodeficiency

[12]

SAIGK

ALG8: Dolichyl pyrophosphate Glc1Man9GlcNAc2 α-1,3-glucosyltransferase

Pathologies: see ALG12 above

[13]

AEIRA

ANXA6 (annexin A6)

Anticoagulant protein from human placenta

[14]

QLIRA, IRASA

AP3B1 (AP-3 complex subunit β-1)

Associates with Hermansky–Pudlak syndrome. Bleeding diathesis resulting in bruising, epistaxis, gingival bleeding, postpartum hemorrhage, bleeding

[15]

LIGAE

APLP2 (amyloid-like protein 2)

The soluble form may have inhibitory properties toward coagulation factors and regulates cerebral thrombosis

[16]

VLLPL

B3AT (band 3 anion transport protein)

Involved in venous thrombosis of unknown origin

[17]

FGGVS

B4GT1 (β-1,4-galactosyltransferase 1)

Defects in the nervous system development, psychomotor retardation, dysmorphic features, hypotonia, coagulation disorders

[18]

KGYHL

C4BPB (C4b-binding protein β chain)

Controls complement activation; binds as a cofactor to C3b/C4b inactivator; possibly involved in the susceptibility to venous thrombosis

[19] [20]

LTVLP

CBS (cystathionine β-synthase)

CBS-deficient patients are prone to vascular thrombosis

[21]

NSVAY

CO1A1 (collagen α-1(I) chain)

Connective tissue disorders characterized by fragile, bruisable skin

[22] [23]

PGQTG, NGLTG

CO1A2 (collagen α-2(I) chain)

Pathology: see CO1A1 above

[22] [23]

TQSLL, GTGVL

COG1 (conserved oligomeric Golgi's complex subunit 1)

Psychomotor retardation, hypotonia, coagulation disorders, and immunodeficiency

[24]

STNLV, GAISS

COG2. (conserved oligomeric Golgi's complex subunit 2)

Pathology: as for COG1

[25]

PINLV

COG5 (conserved oligomeric Golgi's complex subunit 5)

Pathology: as for COG1

[26]

LPFQQ, PFQQF, IGKIQ

ENTP1 (ectonucleoside triphosphate diphosphohydrolase 1)

Implicated in the prevention of platelet aggregation

[27] [28]

YTSAL

EPHB2 (ephrin type-B receptor 2)

Regulation of platelet activation and blood coagulation

[29]

VLNDI

F13A (coagulation factor XIII A chain)

Relates to hematologic disorders characterized by bleeding tendency

[30]

DPLQP

FA5 (coagulation factor V)

Central regulator of hemostasis. Parahemophilia, i.e., poor clotting; pregnancy loss, ischemic stroke, thrombophilia

[31] [32] [33] [34]

PPLLT, FVTQR

FA8 (coagulation factor VIII)

Hemophilia

[35]

NSYEC

FA9 (coagulation factor IX)

Hemophilia

[35]

SSANN

FIBA (fibrinogen α chain)

Bleeding, amyloidosis, arterial hypertension, hepatosplenomegaly, cholestasis, petechial skin rash; thromboembolic complications

[36] [37] [38]

GAGAA

GATA4 (transcription factor GATA-4)

Regulates factor X, a vitamin K-dependent serine protease that functions in blood coagulation. Can predispose to dilated cardiomyopathy, and to premature death

[39] [40] [41]

NDPFL

GP1BA (platelet glycoprotein Ib α chain)

Epistaxis; hemorrhage; menorrhagia; purpura; congenital bleeding diathesis; large platelets; thrombocytopenia; long bleeding time

[42]

ALLAG

GPIX (platelet glycoprotein IX)

Epistaxis; hemorrhage; menorrhagia; purpura; congenital bleeding diathesis; large platelets; thrombocytopenia; long bleeding time

[42]

KLIAN

HABP2 (hyaluronan-binding protein 2)

Serine protease involved in coagulation fibrinolysis and inflammatory pathways

[43]

TQLPP

HPS4 (Hermansky–Pudlak syndrome 4 protein)

Epistaxis; reduced visual acuity; horizontal nystagmus; iris transillumination; restrictive lung disease; bruising; bleeding tendency; menorrhagia; absence of platelet dense bodies; lack of secondary aggregation response of platelets

[44]

HTSPD

HPS5 (Hermansky–Pudlak syndrome 5 protein)

As HPS4 above

[45]

FNATR, DRLIT

HS3S5 (heparan sulfate glucosamine 3-O-sulfotransferase 5)

Catalyzes a crucial step in the biosynthesis of the anticoagulant heparan sulfate

[46]

SASFS

ITA2 (integrin α-2)

Associates with increased ischemic stroke risk; thrombophilia

[47] [48]

VRDLP

ITB3 (integrin β-3)

Thrombasthenia, characterized by mucocutaneous bleeding

[49]

FGTTL, YDPLQ, GDISG

JAK2 (tyrosine-protein kinase JAK2)

Thrombophilia, thrombocytosis

[50] [51]

VNLTT, GDSSS, VTYVP

MMRN1 (multimerin-1)

Deficiency in multimerin-1 associates with bleeding disorder

[52]

LLPLV

PLF4 (Platelet factor 4)

Involved in thrombosis

[53]

TFGAG

PLMN (plasminogen) may be associated with susceptibility to thrombosis

[54]

TVEKG, TGTGV

PROS: vitamin K-dependent protein S

Anticoagulant plasma protein. Helps to prevent coagulation and stimulates fibrinolysis. Deficiency leads to impaired blood coagulation and a tendency to venous thrombosis

[55] [56]

LALHR

PROZ: vitamin K-dependent protein Z

Helps hemostasis by binding thrombin and promoting its association with phospholipid vesicles. Deficiency may be a risk factor for retinal vessel occlusion

[57]

IDRLI

PTPRJ: receptor-type tyrosine-protein phosphatase η

Lack of PTPRJ leads to a bleeding tendency and defective arterial thrombosis

[58]

VFAQV

TF (tissue factor)

Initiates blood coagulation by forming a complex with circulating factor VII or VIIa

[59]

LFRKS

THRB: Prothrombin

Functions in blood homeostasis.

[60]

AGAAL, GAALQ

TRBM (thrombomodulin)

Relates to thrombophilia, venous thrombosis, and thromboembolic disease. TRBM administration is beneficial in sepsis-induced coagulopathy and in disseminated intravascular coagulations

[61] [62] [63]

TLLAL

TSP1 (thrombospondin-1)

Coronary artery disease

[52] [64]

TLLAL, SCGSC

TSP2 (thrombospondin-2)

Coronary artery disease

[52] [64]

VSSQC, LQYGS

VWF (von Willebrand factor)

Von Willebrand's disease is characterized by deficiency of circulating VWF that is otherwise structurally and functionally normal. Clinical features: impaired platelet aggregation, cardiovascular diseases, mucocutaneous bleeding, epistaxis, menorrhagia

[52] [65] [66] [67]

Abbreviations: gp, glycoprotein; SARS-CoV-2, severe acute respiratory syndrome-coronavirus-2.


a Human proteins given by Uniprot accession and name in italics.


b Functions and/or pathologies: data and further references from Uniprot, PubMed, and OMIM databases



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Immunological Potential of the Viral versus Human Peptide Sharing

The data shown in [Table 1] are quantitatively impressive and become strikingly preeminent from a pathological perspective when analyzed for their immunological potential. Indeed, exploration of the IEDB[10] reveals that nearly all the shared pentapeptides described in [Table 1] are also disseminated among SARS-CoV-2 spike gp-derived epitopes that have been experimentally validated as immunoreactive and are cataloged at the IEDB database ( http://www.iedb.org ).[10]

That is, [Table 2] concretely supports the possibility that autoimmune cross-reactions may be triggered by SARS-CoV-2 infection/active immunization and hit human proteins related to thrombotic/thromboembolic disorders and coagulopathies, inter alia. Clinically, the vastity of the potential immunological cross-reactivity that emerges from [Table 2] indicates that mild-to-moderate and severe forms of thrombosis and coagulopathy may unavoidably accompany SARS-CoV-2 infection/active immunization.

Table 2

Distribution of peptides shared between SARS-CoV-2 spike gp and human proteins related to thromboses and hemostasis disorders among 94 experimentally validated SARS-CoV-2 spike gp-derived epitopes

ID[a]

Epitope[b]

ID[a]

Epitope[b]

1069137

aqYTSALLAGtitsg

1309555

qcVNLTTrTQLPPaytnsft

1069290

ctlksfTVEKGiyqt

1309558

qfnSAIGKIQdslsstasal

1071585

nlVRDLPqgfsalep

1309564

qtragcLIGAEhvnNSYECd

1071723

patvcgpkkSTNLVknkc

1309573

rLFRKSnlkpferdisteiy

1072807

skhtPINLVRDLPqg

1309595

tnftisvtteilpvsMTKTS

1072965

svtteilpvsMTKTS

1309598

tvYDPLQPeldsfkeeldky

1073281

tesnkkfLPFQQFgrdia

1309599

Tyvpaqeknfttapaichdg

1073938

vqIDRLITgrlqslq

1309600

tyvtqQLIRAAEIRASAnla

1074201

ylyrLFRKSnlkpfe

1309602

vcgpkkSTNLVknkcvnfnf

1074838

AEIRASAnlaatk

1309603

vknkcvnfnfNGLTGTGVLt

1074925

hVTYVPaqeknf

1309604

VLNDIlsrldkveaevqidr

1074969

lgaeNSVAYsnn

1309621

yskhtPINLVRDLPqgfsal

1074974

lLALHRsyl

1310254

aeNSVAYsnnsiaip

1075005

nqKLIANqf

1310281

aphgvvflhVTYVPa

1075031

rLFRKSnlk

1310303

caqkfngLTVLPpll

1075039

rqiaPGQTGkiadynykl

1310336

dskTQSLLivnnatn

1075066

sVLNDIlsrl

1310392

FGTTLdskTQSLLiv

1075079

tPINLVrdl

1310401

fkiyskhtPINLVrd

1075085

tvYDPLQPeldsfk

1310415

fngLTVLPPLLTdem

1075094

vlPPLLTdemiaqyt

1310434

GAISSVLNDIlsrld

1075125

ysvlynSASFStfk

1310444

givnntvYDPLQPel

1075131

yyvgylqPRTFLl

1310487

iginitrfqTLLALh

1087680

PINLVRDLPqgfsalepl

1310506

irgwiFGTTLdsktq

1125063

gLTVLPpll

1310513

itrfqTLLALHRsyl

1309117

ggnynylyrLFRKSn

1310592

lLALHRsyltpgdss

1309118

gpkkSTNLVknkcvn

1310611

lPPLLTdemiaqyts

1309123

khtPINLVRDLPqgf

1310633

lyenqKLIANqfnsa

1309140

tdemiaqYTSALLAG

1310787

SASFStfkcygvspt

1309147

ylqPRTFLl

1310828

svlynSASFStfkcy

1309418

AEIRASAnlaatkmsecvlg

1310852

tlvkqlssnfGAISS

1309442

ayyvgylqPRTFLlkyneng

1310865

trfqTLLALHRsylt

1309450

dplsetkctlksfTVEKGiy

1310899

VLLPLVSSQCVNLTT

1309451

dsfkeeldkyfknHTSPDvd

1310909

VNLTTrTQLPPaytn

1309461

ehvnNSYECdipigagicas

1310927

vtqnvlyenqKLIAN

1309464

esnkkfLPFQQFgrdiadtt

1310947

wTFGAGAALQipfam

1309469

fknHTSPDvdlGDISGinas

1310979

yvgylqPRTFLlkyn

1309470

fknidgyfkiyskhtPINLV

1311657

ccSCGSCckfdeddsepvlkgvkl

1309475

gccSCGSCckfdeddsepvl

1311813

rLFRKSnlkp

1309492

ilditpcsFGGVSvitpgtn

1313244

nSASFStfk

1309506

kvggnynylyrLFRKSnlkp

1313285

PINLVRDLPqgfsal

1309515

lhrsyltpGDSSSgwtagaa

1313286

PINLVRDLPqgfwal

1309516

litgrlqslqtyvtqQLIRA

1314023

ynylyrLFRKSnlkp

1309523

lssnfGAISSVLNDIlsrld

1317916

gylqPRTFLl

1309524

lyenqKLIANqfnSAIGKIQ

1321084

lPPLLTdem

1309531

NGLTGTGVLtesnkkflpfq

1327418

vYDPLQPeldsf

1309532

ngLTVLPPLLTdemiaqyts

1327923

yenqKLIANqf

1309534

nitrfqTLLALHRsyltpgd

1328800

ytmslgaeNSVAY

Abbreviations: gp, glycoprotein; SARS-CoV-2, severe acute respiratory syndrome-coronavirus-2.


a Epitopes listed as the Immune Epitope DataBase ID.


b Shared peptides given in capital letters.



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Autoimmunity Potential and the Immunological Memory

As already highlighted also in other infection models,[68] [69] [70] [71] one has to consider that immunologic memory can powerfully enhance and amplify the autoimmune cross-reactivity potential because of interpathogen peptide sharing. Indeed, as a rule, the immune system recalls preexisting memory responses toward past infections rather than inducing ex novo responses toward the recent ones since hallmark of the immune system is the memory for the immune determinants it has previously encountered.[72] [73]

Here, comparative sequence analyses show that 31 out of the 60 minimal immune determinants common to SARS-CoV-2 spike gp and human proteins related to thromboses are also widespread in pathogens, such as B. pertussis, C. diphtheriae, C. tetani, H. influenzae, and N. meningitidis, that are in pathogens with which, in general, an individual has already come into contact during his life due to infections or by vaccination ([Table 3]).

Table 3

Occurrence in microbial organisms of pentapeptides common to SARS-CoV-2 spike gp, SARS-CoV-2 spike gp-derived epitome, and human proteins related to thromboses, coagulopathies, and hemostasis disorders

Organism

Shared peptides

Bordetella pertussis

AEIRA, AGAAL, ALLAG, GAALQ, GAGAA, LLPLV, PFQQF, QLIRA, SSANN, TGTGV, VLLPL, YDPLQ

Corynebacterium diphtheriae

AEIRA, AGAAL, ALLAG, DPLQP, GAALQ, GAGAA, GTGVL, LLPLV, TVEKG

Clostridium tetani

AGAAL, LQYGS

Haemophilus influenzae

AEIRA, AGAAL, FGGVS, GAALQ, GAGAA, GTGVL, KLIAN, LALHR, LLPLV, LPFQQ, LTVLP, NLLLQ, NSVAY, TLLAL, TQSLL, VLLPL, VLNDI, VNLTT, YTSAL

Neisseria meningitidis

AEIRA, AGAAL, ALLAG, DRLIT, GAALQ, GAGAA, IDRLI, KLIAN, LALHR, LTVLP, PINLV, TLLAL, VLLPL, VLNDI

Abbreviations: gp, glycoprotein; SARS-CoV-2, severe acute respiratory syndrome-coronavirus-2.


Hence, [Table 3] indicates the possibility that a preexisting immune response to previously encountered pathogens (in the present case: B. pertussis, C. tetani, C. diphtheriae, H. influenzae, and/or N. meningitidis) might be magnified and intensified following SARS-CoV-2 infection/active immunization. That is, immunological imprinting can start a chain of events according to which followings can be measured:

  • Following exposure to SARS-CoV-2, the primary response to the virus can turn into a secondary response to previously encountered pathogens of which the immune system has stored an immunological memory.

  • The anamnestic secondary and, by definition, extremely powerful response against immune determinants previously encountered implies not only that a low or no immune response will be evoked against the pathogen lastly encountered, that is, SARS-CoV-2, but also entails that the anamnestic secondary reaction against the early sensitizing pathogens—in the case in point, B. pertussis, C. tetani, C. diphtheriae, and/or N. meningitidis—will fail because those early sensitizing pathogens are no more present in the organism.

  • As a final result, the anamnestic, high affinity, high avidity, and extremely powerful secondary immune response triggered by the lastly encountered pathogen (SARS-CoV-2) and addressed toward past infections may find an outlet by hitting available human targets, that is, in the case in object, the human proteins related to thromboses and hemostasis diseases ([Table 1]).


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Conclusion

The last decades have seen the emerging of infectious diseases and, consequently, intensive application of immunization procedures. Concomitantly, concerns about possible adverse events have increased. A recent crucial example is the immunization campaign with the dengue vaccine that highlighted the risk of enhanced disease after vaccination.[74]

Today, the clinical context associated with SARS-CoV-2 infection/active immunization is no different. Actually, understanding whether undesired collateral events, such as the thrombotic manifestations and bleeding disorders discussed in this study, may causally associate with the viral infection/active immunization is a fundamental step for fighting the current pandemic. In this context, the present study:

  • Analyzed the hypothesis that infectious agents can induce cross-reactive autoantibodies capable of hitting and altering human proteins that regulate hemostasis and coagulation.

  • Showed that numerous peptides endowed with an immunologic potential are common to SARS-CoV-2 spike gp and human proteins, when mutated, altered, deficient or improperly functioning, are associated with thromboses and hemostasis diseases ([Tables 1] and [2]).

  • Documented that the peptide commonality extends to pathogens that usually have been already encountered by an individual during his life ([Table 3]).

Scientifically, the data indicate that peptide sharing–associated cross-reactivity and, in conjunction, immunological imprint might help explain some of the thromboembolic events that rapidly, massively, and violently may arise following SARS-CoV-2 infection/active immunization.

Clinically, the present data warrant testing of patients' sera for autoantibodies against the peptide targets described in [Tables 1] [2] and [3], and reiterate the suggestion advanced already in 2000[75] that immunotherapies should take advantage of the principle of peptide uniqueness, that is, of peptides present in the antigen of interest and absent in the human proteome.[71] [76] [77] [78] [79] [80] [81]


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Conflict of Interest

None declared.


Address for correspondence

Darja Kanduc, PhD
Department of Biosciences, Biotechnologies, and Biopharmaceutics, University of Bari
Via Orabona 4, Bari 70125
Italy   

Publication History

Publication Date:
26 June 2021 (online)

© 2021. The Author(s). This is an open access article published by Thieme under the terms of the Creative Commons Attribution License, permitting unrestricted use, distribution, and reproduction so long as the original work is properly cited. (https://creativecommons.org/licenses/by/4.0/)

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