Mimicry between a pathogen and self molecules

Rheumatic fever is a classic example for molecular mimicry, post infection with Streptococcus pyogenes (group A streptococcus). The mimicry is between the infecting agent streptococcal M protein and/or a carbohydrate, N-acetyl-beta-d-glucosamine (GlcNAc) and self antigens such as myosin peptides located in the S2 hinge region of the human cardiac myosin leading to rheumatic fever and/or Sydenham’s chorea, respectively (12, 13). The disease is characterized by damage to the heart valves (myocarditis/valvulitis), joints and the brain structures, affecting the central nervous system (Sydenham’s chorea). The molecular mimicry was proven by three main points: a) Activated T cells isolated from a damaged valve recognize various sequences of the bacterial wall M protein in the bacteria cell wall and support B cells secreting anti-heart myosin Abs. b) Purified antibodies from patients with rheumatic fever cross react with streptococcal M protein and heart myosin, laminin, and vimentin. c) Rats immunized with heart myosin developed myocarditis. In Sydenham’s chorea and it’s possible variant pediatric autoimmune neuropsychiatric disorder associated with streptococci (PANDAS), autoantibodies present in Sydenham’s chorea bind to brain gangliosides, signal neuronal cells activating calcium calmodulin-dependent protein kinase II (CaMKII) in neuronal cells and recognize the intracellular protein biomarker tubulin. These Abs cross react with cardiac myosin in the heart’s extracellular membrane antigens such as laminin on the valve surface endothelium.

An additional classical molecular mimicry induces autoimmunity is the case of Guillain-Barré Syndrome (GBS). It is exemplified by damage to the peripheral nervous system mediated by an immune reaction and is characterized by paralysis of the extremities and additional motor damage. Clinical manifestations often appear 1 to 3 weeks after infection by Campylobacter jejuni or a viral infection such as EBV, cytomegalovirus (CMV), or Mycoplasma pneumoniae. Four conditions were determined that prove the molecular mimicry between an infecting agent and a self component (14,15):

1. High titers of Abs cross-reactive with C. jejuni and gangliosides (Galactose-N-Acetyl-GD1a-1, GD1a, GMIb, GMI). An infection with CMV induces synthesis of Abs that cross-react with GM2 whereas M. pneumoniae induces synthesis of Abs that cross-react with galatocerebroside. Pathogenic monoclonal and polyclonal Abs, specific against gangliosides were obtained from patients with GBS. These Abs have biological pathogenic activities such as damage at the ends of motor nerves mediated by complement proteins, ion-gate blocking and damaged the blood-nerve barrier.
2. Presence of GBS CD4+ T cells specific to the shared epitopes. CD8+ cells expressing receptors for αβ or γ, which react against protein components in C. jejuni, were isolated from nerves and peripheral blood samples of GBS patients. The common structure between C. jejuni and nerve cells was identified. Lipo-oligosaccharide (LOS) of C. jejuni imitates the structure on the ganglioside. Abs obtained from patients with GBS bind to LOS from C. jejuni. Immunization of mice with LOS or infection with C. jejuni induces GBS which resembles the disease in humans. The mice developed T cell response and high titers of Abs against gangliosides that had activities similar to Abs obtained from GBS patients.

The last example of molecular mimicry that induces autoimmune disease relates to antiphospholipid syndrome (APS). The disease is characterized by recurrent fetal loss, repeated thromboembolic phenomena, thrombocytopenia, and prolonged coagulation time. These diverse clinical pictures are associated with elevated levels of circulating antiphospholipid β-2-glycoprotein-I (β2GPI)-dependent Abs mainly. Employing a peptide phage display library, our group have identified three hexapeptides that react specifically with the anti-β2GPI monoclonal Abs located on the β2GPI molecule as mimotopes. All three peptides specifically inhibit the biological functions of the corresponding anti-β2GPI monoclonal Abs (for example in-vitro endothelial cell activation and in-vivo induction of experimental APS) (16). Using the Swiss Protein database revealed high homology between the hexapeptides with different bacteria and viruses. The direct proof of induction of experimental APS by peptidomimetic between a pathogen and self came from passive transfer experiments in which mice were immunized with pathogens sharing amino acid sequences with human β2GPI molecule. Experimental APS was induced by passive transfer of affinity purified Abs from the immunized mice on a column composed of the shared peptidomimetic between the infection agent and the β2GPI molecule to naïve mice (16). The mice developed enhanced fetal loss associated with prolonged coagulation time and thrombocytopenia (17). Pierangeli et al (18) demonstrated that immunization of naive mice with CMV and β2GPI shared peptide induced generation of mouse thrombogenic anti-β2GPI Abs in ex vivo models of thrombosis as well as fetal loss. Based on the above studies, molecular mimicry between a pathogen and β2GPI molecule provided proof for the etiology of APS.

A mimicry scenario was proposed by Pender (19) for Epstein-Barr virus (EBV) mediated latent infection of B cells. Human chronic autoimmune diseases, including lupus, multiple sclerosis, Sjogren’s syndrome, rheumatoid arthritis, autoimmune thyroiditis, autoimmune hepatitis, and cryptogenic fibrosing alveolitis, are based on infection of auto reactive B-lymphocytes by EBV. Latent EBV infected auto reactive memory B cells may lodge in organs where their target antigen is expressed and act as antigen presenting cells. When CD4+ T cells that recognize antigens within the target organ are activated in lymphoid organs by mimicry with infectious agents, they migrate to the target organ but fail to undergo activation induced apoptosis because they receive a co-stimulatory survival signal from the infected B cells. The auto reactive T cells proliferate and produce cytokines, which recruit other inflammatory cells with resultant target organ damage and chronic autoimmune disease.

EBV mimicry was proven in lupus patients and experimental models by the shared epitope amino-acid sequences between Epstein–Barr virus nuclear antigen ( EBNA-1), a DNA binding protein that maintains replication of the EBV genome within infected cells and is required for maintaining viral latency, and the major lupus specific Smith rib nucleoprotein complex (Sum B/B’, D1, D2, and D3) (20). The PPPGRRP motif of EBNA-1 share homology with Sum proline-rich motif, PPPGMRPP and are recognized by sera from lupus patients. Mice immunized with the entire EBNA-1 protein vector produced anti-Sum suggesting that the anti-Sum response occurs as a consequence of antigenic mimicry through EBNA-1 antibodies. Immunizing mice and rabbits with EBNA-1-Sm homologous peptides resulted in epitope spreading and lupus-like autoimmunity. SmD195-119 Abs and EBNA-135–58 region show similarity whereas EBNA-2 shares similarity to the C-terminal domain of SmD1 and SmD3. All the EBNA and Sum/Sad similarities are recognized by lupus patient sera. Ro autoantibodies are among the first to appear in SLE. The 60 kea Ro169-180 cross-react with EBNA-158-72 epitopes. Animals immunized with either the first epitope of 60 kea Ro or the cross-reactive EBNA-1 epitope progressively develop autoantibodies to Ro and acquire clinical symptoms of lupus such as leukopenia, thrombocytopenia, and renal dysfunction. However, this EBNA-1 sequence does not share significant homology with the Ro 169–180 sequence. Rather there must be an immunological structural relationship between EBNA-1 and Ro that is sufficiently powerful to persist across species barriers among mammalian immune systems.

The ability of these peptidomimetics shared between EBNA-1, 2, and Sm/SmD or Ro antigens to cause lupus like response in animal models supports the notion of molecular mimicry between EBV and self nuclear antigens as a possible inducer of lupus.

Additional EBV molecular mimicry induces autoimmune response, mediated by shared epitopes on MHC-dependent pathways. An example of that was shown in rheumatoid arthritis (RA). Several autoantigens which undergo citrullination were described in RA such as filaggrin, fibrinogen, vimentin, collagen type II, and α-enolase. Upon citullination, anti-citrullinated peptide/protein Abs (ACPA) are triggered and show high specificity to RA. ACPA are found in RA patients with HLA-DRB1 alleles. The products of these alleles encode a 5 amino acid sequence in a peptide-binding pocket, the so-called shared epitope (SE). Citrullination increases self-antigen immunogenicity through increased binding affinity to SE-containing HLA-DR molecules. HLA molecules carrying the amino acid sequence QKRAA, QRRAA or RRRAA at positions 70–74 of the DRβ1 chain are associated with ACPA positive RA. The QKRAA determinant is also expressed on the EBV protein gp110 and has been shown to be a target of humoral and cellular immune responses in humans (21). One group found decreased T cell response to EBV gp110 in peripheral blood which correlates with disease activity and severity in patients with rheumatoid arthritis (20). In addition, several sequences are found in EBNA-6 and HLA-DQ^*030228: a six-amino acid sequence is shared by EBV reading frame BPLF1 and 65 kD heat shock protein, a protein that induced arthritis in an animal model of adjuvant arthritis (22).

T cell receptor (TCR) from patients with multiple sclerosis (MS) recognized both a DRB1^*1501-restricted myelin basic protein (MBP) and DRB5^*0101-restricted EBV peptide. Crystal structure of the DRB5^*0101-EBV peptide complex revealed a marked degree of structural equivalence to the DRB1^*1501-MBP peptide complex at the surface presented for TCR recognition [22]. CD4+ T cells found in cerebrospinal fluid of MS patients cross react with EBV DNA polymerase peptide EBV627-641 and immunodominant MBP85-99 peptide [22]. The viral LMP1 has homology to MBP suggesting natural cross-reactivity. Thus, antibodies induced against LMP1 during EBV infection might act as an inflammatory trigger to reacting to MBP through a mimicry mechanism (23).

T cell epitope peptidomimetics

Mimicry can also take place at the level of the T cell. Disease inducing peptidomimetics are those peptides of autoantigens that can be presented by major histocompatibility complex (MHC) class II molecules on antigen-presenting cells (APCs) to autoreactive CD4+ T cells (or alternately by HLH subtypes.)

Autoimmune hepatitis (AIH) is a chronic, progressive liver disease, characterized by hepatocellular inflammation and necrosis. A subgroup of AIH patients presents specific autoantibodies to soluble liver antigen/liver-pancreas (SLA/LP) protein. Autoantigenic SLA/LP peptides are targeted by CD4+ T cells and restricted by the allele HLA-DRB1^*03:01, which confers disease susceptibility. A positively charged residue at position 71 has been indicated as critical for AIH susceptibility in all of the HLA alleles identified. Molecular mimicry between SLA/LP and viral/bacterial antigens was illustrated using an in-silico strategy (24). The immunodominant region of SLA/LP was used as the query in databank searches to identify statistically significant similarities with viral/bacterial peptides. Homology modeling and docking was used to investigate the potential interaction of HLA-DRB1^*03:01 with the identified peptides. A statistically significant structural similarity between the immunodominant regions of SLA/LP and a region of the surface antigen PS 120 from Rickettsia spp. has been detected. The interaction of the SLA/LP epitope and the corresponding Rickettsia sequence with the HLA-DRB1^*03:01 allele has been simulated.

Primary biliary cirrhosis (PBC) is an autoimmune cholestatic liver disease characterized by the presence of antimitochondrial Abs and inflammation of interlobular bile ducts. Shimoda et al. (25), showed that human PDC-E2163-176 peptide (GDLLAEIETDKATI) is an immunodominant autoreactive T cell epitope in PBC patients that is restricted by HLA DRB4^*0101. Different T cell clones derived from PBC patients reactive to the human PDC-E2163-176 peptide, were reactive to mimicry peptides derived from microbial proteins. These results proved the involvement of molecular mimicry in PDC-E2 (EQSLITVEGDKASM) peptide from diverse pathogens such as Escherichia coli, Acholeplasma laidlawii, Pseudomonas putida, Neisseria meningitis outer membrane and Clostridium difficile Toxin B protein P64K. Therefore, molecular mimicry was postulated to be a possible mechanism in the development of PBC.

An additional example for T cell epitope peptidomimetics is a systemic disease such as Lyme arthritis which results from infection by the tick-borne spirochete Borrelia burgdorferi, which results in inflammatory joint disorder that resembles rheumatoid arthritis (26). The majority of individuals with the antibiotic treatment resistant disease have the HLA-DRB1^*0401 or HLA-DRB1^*0101 alleles and high titer of IgG specific for outer surface protein A (OspA) of Borrelia burgdorferi. Th1 cells reactive to OspA are often found as well. Immunization with recombinant OspA has been effective in preventing Lyme disease in two clinical trials and is now available as a vaccine. These findings suggest that HLA-DRB1^*0401 or HLA-DRB1^*0101 restricted immune response, that is OspA-specific, somehow precipitates joint-specific autoimmunity. The severity of joint swelling and the duration of Lyme arthritis after antibiotic treatment are associated with T cell responses to specific epitopes of OspA. The nine-residue peptide OspA165–173 was predicted to be the peptide most effectively bound by HLA-DRB1^*0401. In addition, when injected with OspA protein, transgenic mice for HLA-DRB1^*0401 responded primarily to the OspA165–173 peptide as did T cells from an HLA-DRB1^*0401 antibiotic-resistant patient with Lyme arthritis, which were challenged in-vitro. A search of the Genebank protein Database identified one human protein, leukocyte function–associated antigen 1α (hLFA-1α), which contains the peptide hLFA-1α (L332–340). hLFA-1α (L332–340) has homology to the dominant epitope of OspA and was predicted to bind strongly to HLADRB1^* 0401 (this was eventually confirmed experimentally).

Human endogenous retroviruses (HERV) and molecular mimicry. Retroviruse-derived elements in the human genome constitute 90% of non-coding mobile sequences and contribute substantially to the architecture of the human genome. Five to ten percent of the eukaryotic genome consists of elements of retroviral origin. The HERVs were integrated into the human genome 30 to 40 million years ago and are present in primates with the exception of gorillas. HERVs were coined as the key molecular link between the host genome and infectious viral particles. Epigenetic status of the genome (hypomethylation, histone acetylation), UV, chemicals/drugs, injury/stress, hormones, infections, all as a single cause or in concert may modulate HERVs involvement in pathogenic processes. Infection can promote HERV expression by molecular mimicry or by functional mimicry. Several reports have indicated that HERV activation followed by the expression of HERV proteins may play an important role in the induction of autoimmune diseases such as SLE. Clone 4–1 is a member of the HERV-E family and shows sequence homology with Molony murine leukaemia virus; it has ~8.8 kb of sequence including open reading frames in the gag and env regions . Anti-clone 4–1 gag and env product autoantibodies were detected in 48.3 and 10.7% of Japanese SLE patients respectively. These antibodies were not detected in the serum of normal individuals. This finding indicates that transcription of HERV genes may be facilitated and virus particles or components may be produced in some SLE patients. Furthermore, a computer search of current entries in sequence libraries revealed that there are extremely high homologous sequences between clone 4–1 gag region and the E antigen of HLA class I molecules. The homology between clone 4–1 gag protein and E antigen may contribute to the escape of endogenous retrovirus production from the killing effects of CTL and/or NK cells.

Another example is the EBV mediated latent infection of B cells. EBV transactivates HERV-K18 superantigen via docking to the human complement receptor 2 (CD21) on primary B cells and induces HERV-K18 env gene in resting B lymphocytes (27). The env protein encoded by HERV-K18 has a superantigen activity that strongly stimulates a large number of T cells.

Molecular mimicry between a drug and a self-components

Many drugs are processed endogenously by the liver. Consequently, new foreign antigens termed neo-antigens may be created in the form of protein adducts. These new antigens may resemble self-antigens. The metabolism of the anesthetic halothane elevates levels of trifluoroacetylated (TFA) proteins in humans or in mouse models. Only 1 of 3,000 people develop hepatic injury (halothane hepatitis). These people cannot tolerate the presence of TFA proteins and create cross reacting Abs that identify the TFA proteins and pyruvate dehydrogenase (PDH) (28). Molecular mimicry has been defined as TFA-lysine in TFA proteins that imitate clusters of prostatic lipoic acid of the subunit E2 of PDH (27). Interestingly. Patients with halothane hepatitis did not properly express the cross reacting component E2.

Therefore, in this instance, molecular mimicry played a protective role by canceling the cross reacting specificity. These findings raise the possibility that proper introduction of self components (such as PDH) may induce immune tolerance to a self-component and to other foreign antigens that present similar structures, for example, TFA proteins created spontaneously during halothane metabolism.

Molecular mimicry vaccination and infection

Mycobacterium and MS, RA and uveitis

Molecular mimicry between microbial antigens and host-proteins is one of the etiological enigmas for the occurrence of autoimmune diseases. T cells that recognize cross-reactive epitopes may trigger autoimmune reactions. Association of Mycobacterium tuberculosis (M. tuberculosis) has been implicated in different autoimmune diseases including rheumatoid arthritis and multiple sclerosis. Employing bioinformatic tools, Babu Chodisetti et al. (32) have identified potentially cross-reactive T cell epitopes restricted to predominant class I and II alleles of human leukocyte antigens (HLA). These are similar to peptides of mycobacterial proteins and considerable numbers of them are promiscuous. Some of the identified antigens corroborated with established autoimmune diseases linked with mycobacterial infection. The authors’ analysis showed several peptides from mycobacterial HSP60 (also known as HSP65) homologous to human HSP60 and it relatives that bind to many different alleles. One particular peptide, KPLVIIAEDVDGEALSTLVLN, promiscuously bound to many alleles including HLA-DRB1^*15:01 with high affinity. This is suggestive of the fact that such cross-reactive epitopes may initiate the pathogenesis of MS. Another autoimmune disease that has been frequently associated with the occurrence of tuberculosis is RA. Garip et al. (33) investigated the cellular immune response in uveitis developing after intravesical Bacille-Calmette-Guérin (BCG) applications suggesting possible antigenic mimicry of mycobacterial and retinal antigens. A 72-year-old HLA-B27-negative patient with bilateral granulomatous anterior uveitis that developed during the third cycle of intravesical BCG applications. The patient’s peripheral T cell reactivity to ocular autoantigens was compared with the response to purified protein derivative (PPD) from Mycobacterium tuberculosis. T cell proliferation and cytokine and chemokine secretion were measured in vitro. The authors demonstrated proliferation to PPD, interphotoreceptor retinoid-binding protein (IRBP), and IRBP-peptide R16 as well as secretion of proinflammatory cytokines in response to PPD, retinal soluble antigen (S-Ag), IRBP, cell retinal-binding protein (CRALBP), and some S-Ag and IRBP peptides. Amino acid sequence alignments revealed homologies, similar and even identical regions of 5 to 11 amino acids, between proteins from M. tuberculosis, BCG, and retinal antigens suggesting antigenic mimicry as a potential cause of uveitis in this patient.

Vaccination with NY-ESO-1 antigen and autoimmune thyroiditis

Immunotherapies and targeted therapies are frequently associated with thyroid dysfunction, which is in contrast with the rare thyroid abnormalities induced by cytotoxic agents. Vita et al. (34) describe a case of Graves’ disease triggered by NY-ESO-1 in a HLA-A2 negative woman. A 32-year old woman with a synovial sarcoma, received radiotherapy, chemotherapy, and finally NY-ESO-1 vaccine. The patient typed HLA A11/A33 (19), B13/B56 (22), Cw3/-. One month after the beginning of immunotherapy, thyroid dysfunction was clinically suspected and Graves’ disease was biochemically confirmed. The authors hypothesized that NY-ESO-1 shared partial homology with thyroid autoantigens and that at least one pair of homologous sequences contained amino acid sequence binding motif(s) to a restricted number of HLA molecules. They used the software BLAST to search for amino acid sequence homologies between NY-ESO-1 and thyroid autoantigens [TSH-receptor (TSH-R), thyroperoxidase (TPO) and thyroglobulin (Tg)] and the HLA ligand/motif database to look for HLA/T cell receptor binding motifs in the regions of NY-ESO-1 and thyroid autoantigens that were homologous. They found 15 epitopic regions of NY-ESO-1 homologous to 15 regions of thyroid autoantigens, some of which were epitopic: 5 of TSH-R, 8 of Tg, 2 of TPO. These homologous sequences contain binding motifs belonging to several HLA class I antigens including HLA A2 and the patient’s A11 and A33. This case demonstrates again that genetically predisposed patients are at risk to develop thyroid dysfunction after vaccine immunotherapy for malignancies.

HBV vaccine and myelin molecular mimicry

Hepatitis B vaccination (HBV) is a non-infectious viral subunit consisting of the small hepatitis B virus surface antigen (SHBsAg). Over the years, a link between HBV administration and development of demyelinating disorders leading to multiple sclerosis (MS) was demonstrated. Cellular and humoral immunity against myelin basic protein (MBP) and oligodendrocyte glycoprotein (MOG) results in myelin damage in multiple sclerosis (MS). Surfing the protein database, Bogdanos found that known epitopic regions on SHBsAg share extensive homologies with MBP and MOG. Moreover, normal subjects undergoing HBV vaccination developed responses to the shared homologies between the SHBsAg and MOG antigens. The selective appearance of viral and myelin cross-reactive responses in some but not all of the mimicking pairs supports the biological significance that vaccine plays a possible role as an immunomodulator of viral and self cross-reactivity (35).

DNA vaccination and autoimmune hepatitis

Autoimmune diseases have been generated after infection by LCMV in transgenic mice expressing LCMV-nucleoprotein (NP) or glycoprotein in β cells of the islets of Langerhans or their oligodendrocytes (36). These transgenic mice did not develop any immunopathology in the absence of the LCMV challenge. These experiments showed that molecular mimicry between self peptides and viral proteins can be responsible for initiating and maintaining the autoimmune process. Diabetes and CNS autoimmune disease in transgenic mice were mediated by CD8+ cells, and a critical number of activated CTLs were necessary to induce the disease. Djilali-Saiah et al. (37) took this further by demonstrating an autoimmune hepatitis in a transgenic mouse model vaccinated with a neo-self antigen in the liver. The transgenic mouse model proposed in this study was based on the hypothesis that infectious agents have the potential to initiate autoreactivity through molecular mimicry. A transgenic mouse expressing lymphocytic choriomeningitis virus nucleoprotein (NP) in a H-2b background developed liver injury when vaccinated with plasmids expressing NP as an intracellular or a secretory protein. Coinjection of plasmids coding for NP and IL-12 facilitated the induction of a Th1 phenotype as detected by a specific B lymphocyte response characterized by a predominance of IgG2 subclass anti-NP Abs. CTLs activated in peripheral lymphoid organs by DNA vaccination migrated to the periportal and lobular areas of the liver. Their presence was associated with a significant degree of cytolysis as evidenced by elevated transaminases several weeks after immunization. As activated specific T lymphocytes proliferated in the periphery and caused cytolysis of target cells, this study suggests that autoimmune hepatitis can be triggered by molecular mimicry, and that local injury may not be essential to initiate autoreactivity in the liver.