Chronic heart failure is a complex chain of events that lead the heart to loose its ability to pump blood across all bodily blood vessels. The heartbeat is positibvely regulated by adrenaline (speeding up herat rate) or acetylcholine (slowing it down). The faster ios the heart rate, faster heart reserve undergoes exhaustion and faster it would lead to death. Though there are drugs that slow down the heart rate and relieve the cardiac load, CHF still remains a major cause of death in cardiology. In other words, current pharmacotherapy is not able to reverse the failing cellular cascade; scientists, therefore, are trying to find alternative signaling pathways to target or enhance other protecetive cellular cascades. A couople of years ago, researchers at the Cincinnati Children’s Heart Institute have focused on a protein that helps regulate the heart’s response to adrenaline. This alleviated pathological processes in murine models of human heart failure and in cardiac cells isolated from patients with heart failure, who underwent reparative surgery.
Researchers report encouraging preclinical results while pursuing elusive therapeutic strategies to repair healed and malfunctioning cardiac tissues after cardiac injury – by describing an experimental molecular treatment for heart failure. The experimental approach focuses on the role of Gβγ subunits and GRK2 proteins, which are involved in a signaling pathway activated by adrenal stimulation. Adrenaline receptora are couopled to G protein, composed of three subunits: alpha, beta and gamma. Many years ago, alpha subunit was deemed the main if not exclusive mediator of surface receptors for hormones and neuroptransmitters. Later, it was recognized that beta-gamma complex is also able to trigger or suppress other signlaing pathways or enzymes in response to an external stimulus.The adrenergic system plays a fundamental role in the maintenance of normal cardiac function and the overstimulation of the system induces ventricular hypertrophy – a thickening and enlargement of the heart muscle.
It also causes fibrosis, the formation of scar tissue. In a mouse model that closely mimics the progression of the disease in humans after an infarction, the researchers blocked the molecular Gβγ-GRK2 signaling with a small experimental molecular inhibitor called gallein. When treatment started one week after the initial cardiac injury, it retained cardiac function and reduced scarring and tissue enlargement: essentially it saved animals from heart failure. Researchers also reported a similar level of protection in a new genetically modified mouse model in which GRK2 is removed from cardiac fibroblasts. Unfortunately, there are essentially no clinical interventions that effectively target these tissue-damaging cardiac fibroblasts. These specific cells, indeed, are responsible for collagen deposition while the heart proceeds along its enlargement. Collagen is not elastic (rather, it is stiff) and basically impinges onto cardiac fibers contraction.
Cardiac fibroblasts are the most abundant non-myocyte cell type in the heart, constituting about 60% of cell number and 15% of cell mass of the tissue. They play key regulatory roles in cardiac remodeling, fibrosis and hypertrophy through regulating cell proliferation, producing and remodeling extracellular matrix (ECM). The beta-2 receptor (β2AR) is the highest expressed adrenergic subtype in cardiac fibroblasts. Stimulation of β2 receptors only promote degradation of collagen and induces autophagy. They are also able to crosstalk with the epidermal growth factor receptor (EGFR), a phenomenon called tras-activation, that allows cells to enhance mitogenic cascades (ERK1, ERK2) that are not conventionally activated by the adrenergic receptor. Indeed, beta receptors are coupled with the generation of the second messenger cyclic AMP (cAMP) and its signaling protein kinase (PKA). The cAMP-PKA axis promotes a faster heart rate, that is why beta-blockers are the hinge for the current CHF management.
Not only the study identifies the cardio-protective properties of pharmacological inhibition of the Gβγ-GRK2 in a clinically relevant laboratory model, data also demonstrated that inhibition reduced the activation of cardiac fibroblasts by human heart failure, responsible for scarring of heart tissue. Heart muscle diseases are common disorders in both the pediatric and adult populations, that are linked to heart fibrosis. Adult congenital heart disease is also a growing concern for Cincinnati Children doctors and researchers, who are developing new clinical and research strategies for adults who were pediatric cardiopaths. Despite the current promising preclinical results of the study, the authors point out that it is too early to say whether the data will result in clinically beneficial treatments for patients with human heart failure. However, the data open the door to the development of new pharmacological compounds, since it’s already fifteen years that scientists collected, numerous data about the therapeutic efficacy of the experimental compound gallein in various animal models of heart failure.
A current differente approach to identifying new therapeutics has been to identify ligands that interact with surface receptors (GPCRs), in binding modes that favour specific conformations of the receptor that activate only select downstream pathways. The emphasis has been on finding ligands that lead either to preferential activation of G proteins or to β-arrestin binding by receptors themselves. One example is the discovery of μ-opioid receptor agonists that bias it towards G protein activation over β-arrestin recruitment to improve the safety of opioid analgesics. Recent clinical trial data indicate that a new G protein-biased opioid agonist, oliceridine, is effective at relieving postoperative pain, with significantly less nausea and respiratory depression. Direct G protein targeting is an alternative approach to bias GPCRs by blocking selected post-receptor signalling pathways. Targeting specific G protein subunits can bias GPCR signals away from detrimental signalling pathways, leaving untouched pathways that are essential for normal cell functioning.
Targeting G protein subunits that are common to signalling downstream of receptor families may also improve therapeutic efficacy in complex disease such as heart failure, inflammation and asthma. Finally, targeting G proteins themselves could have efficacy in treating diseases driven by G protein dysregulation. For example, To date, the only specific bioavailable inhibitors of Gα subunits that have a good level of validation are inhibitors of Gαq family members. YM-254890 is a cyclic peptide that was isolated from a bacterium as an inhibitor of ADP-dependent platelet aggregation. These and subsequent studies demonstrated that YM-254890 inhibits Gαq/11 signalling downstream of multiple Gαq-linked receptors in platelets and other cells without affecting other signalling pathways. A highly related compound, FR900359, isolated from the ornamental primrose plant Ardisia crenata, also is a potent and specific inhibitor of Gαq/11 signaling.
Prototypical Gβγ inhibitors were discovered in a competition screen with the peptide SIGK for binding to these subunits. Gallein is one of these, along with M119. They are both fluorescein derivatives. Binding of both gallein and M119 to Gβγ is slowly reversible with an half-life of almost 2 hours. This is why sceìientista re trying to find derivatives with higher affinity and cellular stability toward the beta-gamma subunits. One concern is that Gβγ is a participant in every G protein signalling pathway, because it is required for G protein activation by GPCRs, likely because Gβγ stabilizes the conformation of Gα. As such, completely blocking Gβγ subunit function would completely decouple the G protein system. Thus, the pharmacological approach to blocking Gβγ function must prevent downstream signalling without interfering with G protein activation in general. Proof of principle that this is achievable comes from the capacity of M119 or gallein to block Gβγ signalling without affecting GPCR-dependent Gα activation.
In a recent study, inhibition of Gβγ-regulated GRK2 activation, using adenoviruses to express GRK2ct, prevented α2-receptor desensitization and reduced plasma adrenaline levels in a heart failure model. This finding suggested that systemic administration of gallein achieves high therapeutic efficacy through a dual mechanism of action: through direct effects in cardiac myocytes and by lowering circulating catecholamine levels. Indeed, treatment with gallein normalized plasma adrenaline and noradrenaline levels and restored α2-AR-mediated feedback inhibition of catecholamine release. It has been proposed that Gβγ inhibition in cardiac myocytes inhibits progression of heart failure by preventing Gβγ-dependent recruitment of GRK2, thereby preventing desensitization of the β-AR. However, treatment with Gβγ inhibitors likely blocks other pathways associated with Gβγ in the heart. For example, it was recently shown that Gβγ can directly bind to and activate ERK, leading to its phosphorylation and trans-location to the nucleus in cardiac myocytes. This will enhance cardiac fiber hypertrophy.
Another study identified Gβγ as a regulator of the phospholipase PLCε activation at the Golgi apparatus, which is also critical for endothelin 1 (ET1)-driven muscle cell hypertrophy. Gallein inhibits Gβγ-dependent regulation of PLCε in cardiac myocytes, and Gβγ inhibition has the potential to inhibit ERK activation. Thus, Gβγ inhibition has the potential to be highly efficacious by virtue of its ability to block activation of multiple targets by Gβγ linked with heart fiber overgrowth. Besides, it could also interfere with the previously mentioned cross-talk between adrenaline receptor and the mitogenic EGF receptor acting on heat fibroblast. Ot would be an additional braking effect on the deposition of collagen and, thus, over heart stiffening. There is yeat a long way ahead, but it is not to be forgotten that chronic heart failure affects approximately 6.5 million American adults and to 30 billion dollars healthcare expenditure each year. Importantly, the 1-year and 5-year mortality rate after hospitalization approaches 20% and 40%.
Therefore, understanding the fundamental mechanisms underlying the development of this condition is a must.
- edited by Dr. Gianfrancesco Cormaci, PhD, specialist in Clinical Biochemistry.
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