Atherosclerosis. 2016 Jan;244:44-7. doi: 10.1016/j.atherosclerosis.2015.11.001.

Aptamer BC007 for neutralization of pathogenic autoantibodies directed against G-protein coupled receptors: A vision of future treatment of patients with cardiomyopathies and positivity for those autoantibodies.
 

Johannes Müller, Gerd Wallukat and Ingolf Schimke*

Berlin Cures GmbH, Berlin, Germany

*Corresponding author; schimke@berlincures.de

 

Abstract

Cardiomyopathies such as idiopathic dilated cardiomyopathy (DCM), Chagas’ cardiomyopathy and Peripartum cardiomyopathy present with autoantibodies against G-protein coupled receptors (GPCR-AABs) that agonistically activate their receptors. For the treatment of “agonistic autoantibody diseases” and in particular DCM, the removal of the GPCR-AABs by immunoadsorption (IA) has been studied with convincing patient benefit. To overcome cost and logistics problems of IA, the application of the aptamer BC007 for in vivo neutralization of GPCR-AABs could help. We demonstrate here, that the aptamer neutralized, in vitro, the presently known cardiovascular-pathogenic GPCR-AABs. In spontaneously hypertensive rats, the aptamer demonstrated its GPCR-AAB neutralizing potency in vivo. In the serum of DCM patients, the same GPCR-AAB reduction was achieved when patients were either immunoadsorbed or patient’s serum was ex vivo treated with the aptamer. In our view, aptamer BC007 treatment in GPCR-AAB-positive patients would have a comparable benefit as that seen after IA. Not knowing all that interfering with our idea of aptamer-dependent neutralization of GPCR-AABs, the first preliminary steps have been taken for bringing the idea closer to patients.

KEYWORDS: Aptamer; Autoantibodies; Beta1-adrenergic receptor; Cardiomyopathy; G-protein coupled receptor

PMID: 26584137

 

Supplements:

Cardiomyopathy is a disease of the heart muscle usually leading to heart failure, in which the heart is unable to pump enough blood needed by the other organs. Typical symptoms of heart failure patients are shortness of breath (dyspnea), weakness and swelling (edema) in legs, ankles and feet, and reduced ability to exercise, often combined with rapid or irregular heartbeat. Among the types of cardiomyopathy, dilated cardiomyopathy (DCM), which is characterized by the stretching and dilating of the heart chambers, is most common. Dilated cardiomyopathy affects people of all ages, including children, but is most common in men aged 20–60. Annually in the US, 10,000 deaths and 46,000 hospitalizations are related to DCM. For nearly 50% of DCM patients, the reason for the disease is unknown. However, in this group of patients (idiopathic dilated cardiomyopathy, IDCM) as well as in other forms of dilated cardiomyopathy, such as Chagas cardiomyopathy (nearly 3 million patients in Latin America) and peripartum cardiomyopathy, an uncommon disorder associated with pregnancy (1 case per 2500–4000 live births in Northern America and Europe), autoantibodies were found. Additionally to DCM, such autoantibodies were found in many other diseases of the cardiovascular system and in diseases associated with vascular alteration ([1]).

In contrast to classic autoantibodies, which destroy their target tissues, the autoantibodies indicated in Table 1 bind to receptors, specifically those belonging to the class of G-protein coupled receptors (GPCRs). These receptors are involved in the regulation of almost all of the body system, from sensoric reception to regulation of cell activity, movement and death. The interaction of GPCR-AABs with their related receptors is, for the GPCR-AABs directed against the beta1-adrenergic receptor (beta1-AABs), schematically demonstrated in Figure 1.

 

 

fig1

Figure 1

The human G-protein-coupled receptor targeted by the corresponding autoantibody. The N-terminal domain usually contains fewer than 50 amino acids and is located in the extracellular space, whereas the C-terminal part of the protein varies from 23 (muscarinic2 receptor) to about 100 amino acids (beta2-adrenergic receptor). The transmembrane regions usually contain between 23 and 24 amino acids, limited by the helical secondary structure and the thickness of the hydrophobic lipid bilayer. The homology varies throughout the entire superfamily. However, the homology is higher (35%; 90%) in the transmembrane helices and functionally important side chains in the transmembrane helices, and the loops are strongly conserved between different vertebrate species. Agonists bind to a hydrophobic cave formed by the transmembrane helices. Indicated epitopes on the second extracellular loop are the targets of beta1-adrenergic receptor autoantibodies present in patients with idiopathic dilated cardiomyopathy (DCM), Chagas’ cardiomyopathy (Chagas’ CM), and peripartum cardiomyopathy (peripartum CM). Reproduced from ([2]) with permission of Pabst Science Publishers, Lengerich.

 

GPCR-AABs don’t destroy their targets but functionally affect the related receptors just as it do physiologic hormones and neurotransmitters. However, the receptor activation due to binding of neurotransmitters and hormones are embedded in a complex protection system (e.g. receptor desensitization, receptor down-regulation) to prevent over-boarding receptor activation. In contrast, GPCR-AABs miss such a protection system after receptor binding, which results in excessive and over-boarding effects. As demonstrated in Figure 1, receptor dimerization is typical after GPCR-AAB binding, which is probably responsible for missing receptor desensitization and down-regulation.

 

Focusing on cardiomyopathy, GPCR-AABs such as directed against the beta1-adrenergic receptor (beta1-AABs) and muscarinic 2- receptors (M2-AABs) are frequently present. However, a couple of further GPCRs can be targeted by related AABs which are clearly also pathogenic. After the binding of beta1-AABs and M2-AABs, the overstimulation of the related receptors, due to being uncontrolled, leads to disturbed heart function and later to the death of active heart cells, which are than replaced by inactive connective tissue. Due to affecting cell functions, the related GPCR-AABs can be named “functional autoantibodies”. The diseases presenting with functional autoantibodies should be consequently summarized to a new class of autoimmune diseases named “functional autoantibody diseases”., Functional autoantibodies were found in many of the diseases characterized by disturbances in the cardiovascular system; among these, hypertension, diabetes mellitus and even Alzheimer’s disease are most prominent.

GPCR-AABs were found in small quantities (<10%) in healthy subjects. However, GPCR-AAB positivity is particularly pronounced in patients suffering from diseases of the cardiovascular system or from diseases associated with vascular alterations. The discussed pathogenic consequences of GPCR-AABs are summarized in (1,2,[3],[4],[5],[6]).

 

With respect to DCM patients and despite continuously optimized medical treatment, heart transplantation is often the last chance. Fortunately, it has been evidenced for those DCM patients carrying functional autoantibodies, mainly those directed against the beta1-adrenergic receptor, that their long-time survival without needing heart transplantation is importantly increased if the patients’ blood is cleared of the autoantibodies using the immunoapheresis technique (IA) ([7]). Until now, IA has been successful applied for patients testing positive for functional autoantibodies and suffering from DCM ([8]), DCM associated with diabetes mellitus ([9]), pulmonary hypertension ([10]) and thrombangiitis obliterans ([11]). IA treatment has also been suggested for patients with Chagas’ heart disease. Presently, to the best of our knowledge, it is planned to study IA in Alzheimer`s patients.

Unfortunately, high cost, logistic problems and patient burden associated with IA could be a major issue, especially in developing countries, meaning that they prevent the wider use of IA technology as a treatment option. This could explain why IA for GPCR-AABs, despite the clearly evidenced patient benefit, has been haltingly used in the treatment of DCM patients positive for GPCR-AABs, and even worse has not entered the therapy of the millions of patients with Chagas’ heart disease, which is one of the main causes of death in Latin America.

To overcome the limitations associated with IA treatment, one possible strategy could be to aim at the in vivo neutralization of patients’ GPCR-AABs by treatment with an aptamer binding and neutralizing these AABs. Aptamers (chemical antibodies) are single- or double-stranded oligonucleotides that – if well selected – bind their target molecules in a similar way to antibodies. However, in contrast to autoantibodies, oligonucleotides possess low toxicity and a lack of immunogenicity, and can be easily synthetized and modified for stability and for improvement of their pharmacokinetic and pharmacodynamic properties ([12],[13]).

In view of the aptamer concept for in vivo GPCR-AAB neutralization, the Berlin Cures GmbH, Berlin, Germany (https://berlincures.de) owns the aptamer BC007 ([14]) protected by patents held by or exclusively licensed to Berlin Cures for commercial exploitation. Berlin Cures GmbH is a spin-off company founded, among others, by a group of scientists, formerly from the Max Delbrück Center (MDC) and the Charité – Universitätsmedizin Berlin in September 2014 in order to bring BC007 to the market. BC007 is a single-strand 15 mer DNA aptamer that was recently successfully tested in cell culture and animal studies for its GPCR-AAB neutralization. Thereafter, comprehensive preclinical studies have evidenced the safety of BC007. Sponsored by Berlin Cures GmbH, a phase-1 clinical trial of BC007 for the indication of dilative cardiomyopathy is currently in preparation.

To bring Berlin Cures’s idea step by step closer to humans, we first demonstrated that BC007 is able to inhibit all the functional autoantibodies presently discussed as being pathogenic in human cardiovascular diseases ([15]). For this purpose, we prepared GPCR-AABs from the blood of patients suffering from diseases of the cardiovascular system. For these GPCR-AABs we demonstrated in vitro that their pathogenic activity was strongly reduced in the presence of BC007. In the second line of experiments, we treated rats carrying a functional autoantibody directed against the beta1-receptor (a specific receptor necessary to guarantee normal heart function) with BC007 (9). In this study, the blood’s autoantibody inactivation could be evidenced in vovo. Importantly, there was no substantial autoantibody return until study end. No toxic side effects of the drug were seen. Finally, we demonstrated for DCM patients who presented after IA in correlation to the loss of beta1-AABs with clear clinical benefit, that a comparable autoantibody loss could be achieved in cases where the autoantibodies of the same patients were treated with BC007.

With respect to these findings, BC007 treatment in patients carrying GPCR-AABs should have a comparable benefit to that seen after IA treatment. Consequently, BC007 could offer a hopeful treatment strategy for patients with cardiomyopathy, mainly for those with DCM, Chagas’ cardiomyopathy and peripartum cardiomyopathy. Due to the evidenced potency of BC007 to neutralize the most or even all of the GPCR-AABs found in patients with cardiovascular diseases or with diseases associated with vascular alterations, we postulate the potential of BC007 treatment for many other diseases, among which clearly the most important is Alzheimer’s disease.

 

References:

[1].       Wallukat G, Schimke I. Agonistic autoantibodies directed against G-protein-coupled receptors and their relationship to cardiovascular diseases. Seminars in Immunopathology 2014;36:35163.

[2].       Munoz-Saravia SG, Haberland A, Wallukat G, Schimke I. Chronic Chagas’ heart disease – From pathogenesis to treatment regimes. Applied Cardiopulmonary Pathophysiology 2012;16:55-81.

[3].       Wallukat G, Nissen E, Müller J, Brinckmann R, Schimke I, Kunze R. The pathophysiological role of autoantibodies directed to G-protein coupled receptors and therapeutic strategies of antibody removal. In: Kunze R, Brinkmann R (eds.) Affina academy: G-protein coupled receptors and autoantibodies, 7–47. Lengerich: Pabst Science Publishers; 2002. http://​www.​pabst-publishers.​de/​Medizin/​buecher/​3936142939.​htm

[4].       Xia Y, Kellems RE. Receptor-activating autoantibodies and disease: preeclampsia and beyond. Expert Rev Clin Immunol. 2011;7:659-74.

[5].       Luft FC. Activating autoantibodies and cardiovascular diseases. Physiology 2013;28:254-61.

[6].       Patel PA, Hernandez AF. Targeting anti-beta1-adrenergic receptor antibodies for dilated cardiomyopathy. Eur J Heart Fail. 2013;15:724-29.

[7].       Dandel M, Wallukat G, Englert A, Lehmkuhl HB, Knosalla C, Hetzer R. Long-term benefits of immunoadsorption in β(1)-adrenoceptor autoantibody-positive transplant candidates with dilated cardiomyopathy. Eur J Heart Fail 2012;14:1374-88.

[8].       Müller J, Wallukat G, Schimke I. Autoantibody directed therapy in cardiovascular diseases. In: Nussinovitch U. The Heart in Rheumatologic, Inflammatory and Autoimmune Diseases: Pathophysiology, Clinical Aspects and Therapeutic Approaches. Elsevier 2016 (in press)

[9].       Dandel M, Englert A, Wallukat G, Riese A, Knosalla C, Stein J, Hetzer R. Immunoadsorption can improve cardiac function in transplant candidates with non-ischemic dilated cardiomyopathy associated with diabetes mellitus. Atheroscler Suppl 2015;18:124-33.

[10].      Dandel M, Wallukat G, Englert A, Hetzer R. Immunoadsorption therapy for dilated cardiomyopathy and pulmonary arterial hypertension. Atheroscler Suppl 2013;14:203-11.

[11].      Klein-Weigel PF, Bimmler M, Hempel P, Schöpp S, Dreusicke S, Valerius J, Bohlen A, Boehnlein JM, Bestler D, Funk S, Elitok S. G-protein coupled receptor auto-antibodies in thromboangiitis obliterans (Buerger’s disease) and their removal by immunoadsorption.Vasa. 2014;43:347-52.

[12].      Ellington A. Aptamers as therapeutics. Nat Rev Drug Discov. 2010;9:537-650.

[13].      Marquis JK, Grindel JM. Toxicological evaluation of oligonucleotide therapeutics. Curr Opin Mol Ther. 2000;2:258-263.

[14].      Schimke I, Haberland A, Wallukat G (2012) Use of aptamers in therapy and/or diagnosis of autoimmune diseases. EP2497828A1, WO2012119938A2, US9234201 B2.

[15].      Wallukat G, Müller J, Haberland A, Berg S, Schulz A, Freyse EJ, Vetter R, Salzsieder E, Kreutz R, Schimke I. Aptamer BC007 for neutralization of pathogenic autoantibodies directed against G-protein coupled receptors: A vision of future treatment of patients with cardiomyopathies and positivity for those autoantibodies. Atherosclerosis 2016;244:44-7.

 

 

Multiselect Ultimate Query Plugin by InoPlugs Web Design Vienna | Webdesign Wien and Juwelier SchönmannMultiselect Ultimate Query Plugin by InoPlugs Web Design Vienna | Webdesign Wien and Juwelier Schönmann