Rational in silico design identifies two mutations that restore UT28K SARS-CoV-2 monoclonal antibody activity against Omicron BA.1.

SARS-CoV-2 rapidly mutates and acquires resistance to neutralizing antibodies. We report an in-silico-designed antibody that restores the neutralizing activity of a neutralizing antibody. Our previously generated antibody, UT28K, exhibited broad neutralizing activity against mutant variants; however, its efficacy against Omicron BA.1 was compromised by the mutation. Using previously determined structural information, we designed a modified-UT28K (VH T28R/N57D), UT28K-RD targeting the mutation site. In vitro and in vivo experiments demonstrated the efficacy of UT28K-RD in neutralizing Omicron BA.1. Although the experimentally determined structure partially differed from the predicted model, our study serves as a successful case of antibody design, wherein the predicted amino acid substitution enhanced the recognition of the previously elusive Omicron BA.1. We anticipate that numerous similar cases will be reported, showcasing the potential of this approach for improving protein-protein interactions. Our findings will contribute to the development of novel therapeutic strategies for highly mutable viruses, such as SARS-CoV-2.

Liver lipophagy ameliorates nonalcoholic steatohepatitis through extracellular lipid secretion.

Nonalcoholic steatohepatitis (NASH) is a progressive disorder with aberrant lipid accumulation and subsequent inflammatory and profibrotic response. Therapeutic efforts at lipid reduction via increasing cytoplasmic lipolysis unfortunately worsens hepatitis due to toxicity of liberated fatty acid. An alternative approach could be lipid reduction through autophagic disposal, i.e., lipophagy. We engineered a synthetic adaptor protein to induce lipophagy, combining a lipid droplet-targeting signal with optimized LC3-interacting domain. Activating hepatocyte lipophagy in vivo strongly mitigated both steatosis and hepatitis in a diet-induced mouse NASH model. Mechanistically, activated lipophagy promoted the excretion of lipid from hepatocytes, thereby suppressing harmful intracellular accumulation of nonesterified fatty acid. A high-content compound screen identified alpelisib and digoxin, clinically-approved compounds, as effective activators of lipophagy. Administration of alpelisib or digoxin in vivo strongly inhibited the transition to steatohepatitis. These data thus identify lipophagy as a promising therapeutic approach to prevent NASH progression.

Structural delineation and computational design of SARS-CoV-2-neutralizing antibodies against Omicron subvariants.

SARS-CoV-2 Omicron subvariants have evolved to evade receptor-binding site (RBS) antibodies that exist in diverse individuals as public antibody clones. We rationally selected RBS antibodies resilient to mutations in emerging Omicron subvariants. Y489 was identified as a site of virus vulnerability and a common footprint of broadly neutralizing antibodies against the subvariants. Multiple Y489-binding antibodies were encoded by public clonotypes and additionally recognized F486, potentially accounting for the emergence of Omicron subvariants harboring the F486V mutation. However, a subclass of antibodies broadly neutralized BA.4/BA.5 variants via hydrophobic binding sites of rare clonotypes along with high mutation-resilience under escape mutation screening. A computationally designed antibody based on one of the Y489-binding antibodies, NIV-10/FD03, was able to bind XBB with any 486 mutation and neutralized XBB.1.5. The structural basis for the mutation-resilience of this Y489-binding antibody group may provide important insights into the design of therapeutics resistant to viral escape.

An engineered ACE2 decoy neutralizes the SARS-CoV-2 Omicron variant and confers protection against infection in vivo.

The Omicron (B.1.1.529) SARS-CoV-2 variant contains an unusually high number of mutations in the spike protein, raising concerns of escape from vaccines, convalescent serum, and therapeutic drugs. Here, we analyzed the degree to which Omicron pseudo-virus evades neutralization by serum or therapeutic antibodies. Serum samples obtained 3 months after two doses of BNT162b2 vaccination exhibited 18-fold lower neutralization titers against Omicron than parental virus. Convalescent serum samples from individuals infected with the Alpha and Delta variants allowed similar frequencies of Omicron breakthrough infections. Domain-wise analysis using chimeric spike proteins revealed that this efficient evasion was primarily achieved by mutations clustered in the receptor binding domain but that multiple mutations in the N-terminal domain contributed as well. Omicron escaped a therapeutic cocktail of imdevimab and casirivimab, whereas sotrovimab, which targets a conserved region to avoid viral mutation, remains effective. Angiotensin-converting enzyme 2 (ACE2) decoys are another virus-neutralizing drug modality that are free, at least in theory, from complete escape. Deep mutational analysis demonstrated that an engineered ACE2 molecule prevented escape for each single-residue mutation in the receptor binding domain, similar to immunized serum. Engineered ACE2 neutralized Omicron comparably to the Wuhan strain and also showed a therapeutic effect against Omicron infection in hamsters and human ACE2 transgenic mice. Similar to previous SARS-CoV-2 variants, some sarbecoviruses showed high sensitivity against engineered ACE2, confirming the therapeutic value against diverse variants, including those that are yet to emerge.

Energy-sparing by 2-methyl-2-thiazoline protects heart from ischaemia/reperfusion injury.

AIMS: Cardiac ischaemia/reperfusion (I/R) injury remains a critical issue in the therapeutic management of ischaemic heart failure. Although mild hypothermia has a protective effect on cardiac I/R injury, more rapid and safe methods that can obtain similar results to hypothermia therapy are required. 2-Methyl-2-thiazoline (2MT), an innate fear inducer, causes mild hypothermia resulting in resistance to critical hypoxia in cutaneous or cerebral I/R injury. The aim of this study is to demonstrate the protective effect of systemically administered 2MT on cardiac I/R injury and to elucidate the mechanism underlying this effect. METHODS AND RESULTS: A single subcutaneous injection of 2MT (50 mg/kg) was given prior to reperfusion of the I/R injured 10 week-old male mouse heart and its efficacy was evaluated 24 h after the ligation of the left anterior descending coronary artery. 2MT preserved left ventricular systolic function following I/R injury (ejection fraction, %: control 37.9 ± 6.7, 2MT 54.1 ± 6.4, P < 0.01). 2MT also decreased infarct size (infarct size/ischaemic area at risk, %: control 48.3 ± 12.1, 2MT 25.6 ± 4.2, P < 0.05) and serum cardiac troponin levels (ng/mL: control 8.9 ± 1.1, 2MT 1.9 ± 0.1, P < 0.01) after I/R. Moreover, 2MT reduced the oxidative stress-exposed area within the heart (%: control 25.3 ± 4.7, 2MT 10.8 ± 1.4, P < 0.01). These results were supported by microarray analysis of the mouse hearts. 2MT induced a transient, mild decrease in core body temperature (°C: -2.4 ± 1.4), which gradually recovered over several hours. Metabolome analysis of the mouse hearts suggested that 2MT minimized energy metabolism towards suppressing oxidative stress. Furthermore, 18F-fluorodeoxyglucose-positron emission tomography/computed tomography imaging revealed that 2MT reduced the activity of brown adipose tissue (standardized uptake value: control 24.3 ± 6.4, 2MT 18.4 ± 5.8, P < 0.05). 2MT also inhibited mitochondrial respiration and glycolysis in rat cardiomyoblasts. CONCLUSIONS: We identified the cardioprotective effect of systemically administered 2MT on cardiac I/R injury by sparing energy metabolism with reversible hypothermia. Our results highlight the potential of drug-induced hypothermia therapy as an adjunct to coronary intervention in severe ischaemic heart disease.

Orotic acid protects pancreatic ß cell by p53 inactivation in diabetic mouse model.

Impairment of pancreatic ß cells is a principal driver of the development of diabetes. Restoring normal insulin release from the ß cells depends on the ATP produced by the intracellular mitochondria. In maintaining mitochondrial function, the tumor suppressor p53 has emerged as a novel regulator of metabolic homeostasis and participates in adaptations to nutritional changes. In this study, we used orotic acid, an intermediate in the pathway for de novo synthesis of the pyrimidine nucleotide, to reduce genotoxicity. Administration of orotic acid reduced p53 activation of MIN6 ß cells and subsequently reduced ß cell death in the db/db mouse. Orotic acid intake helped to maintain the islet size, number of ß cells, and protected insulin secretion in the db/db mouse. In conclusion, orotic acid treatment maintained ß cell function and reduced cell death, and may therefore, be a future therapeutic strategy for the prevention and treatment of diabetes.

A rare appearance of a large mural thrombus in left atrium detected two years after the Maze procedure.

A 77-year-old Japanese woman underwent bioprosthetic aortic valve replacement (AVR) and the Maze procedure for severe aortic valve disease and paroxysmal atrial fibrillation (AF), and one year after the AVR, she also underwent a permanent pacemaker implantation for sick sinus syndrome. At two postoperative years, a large mural mass happened to be detected in her left atrium on routine trans-thoracic echocardiography. The cardiac rhythm records produced by the implanted pacemaker demonstrated the recurrence of AF. As anticoagulant therapy was not effective at reducing the size of the mass, surgery was performed and organized thrombus was detected on the ablation line made at the Maze procedure. .

Successful percutaneous coronary intervention with GuideLiner® catheter for subtotal occlusive lesion in the right coronary artery with anomalous origin from the left sinus of Valsalva: a case report.

INTRODUCTION: Because of the unusual anatomy of an anomalous origin of the right coronary artery from the left sinus of Valsalva, selective cannulation of the guiding catheter in percutaneous coronary intervention for these cases is always challenging. CASE PRESENTATION: A 58-year-old Japanese man was admitted to our hospital complaining of worsening exertional chest pain. He was suspected of having unstable angina pectoris and underwent cardiac catheterization. We found a subtotal occlusive lesion in the mid-portion of his right coronary artery that originated from the left sinus of Valsalva. On the previous percutaneous coronary intervention, we failed to cannulate the guiding catheter to the anomalous orifice of the right coronary artery. Therefore, we decided to use the GuideLiner catheter for stable back-up support from the beginning. A 6Fr GuideLiner catheter was introduced into the right coronary artery by anchoring it coaxially with a semi-compliant balloon catheter. And we successfully deployed two drug-eluting stents by crossing over the posterior-descending artery. Final angiography demonstrated favorable dilatation of the target lesion, and native blood flow in the right coronary artery was completely recovered. CONCLUSION: GuideLiner is a monorail-type "child" support catheter that facilitates coaxial guiding catheter engagement and an appropriate back-up force, achieving successful device delivery to target lesions in this kind of complex percutaneous coronary intervention.

Apical myocardial infarction with bizarre coronary images mimicking left ventricular apical ballooning syndrome: a case report.

INTRODUCTION: Although several etiopathogenetic mechanisms have been proposed, the causes of left ventricular apical ballooning syndrome are still controversial. CASE PRESENTATION: A 51-year-old Japanese woman consulted the emergency room complaining of the sudden onset of anterior chest pain while shopping. We initially suspected her disease as left ventricular apical ballooning syndrome based on her clinical background and laboratory examinations. However, the initial coronary angiogram demonstrated diffuse lesions in her distal left anterior descending coronary artery, and she was finally diagnosed with apical myocardial infarction. The blood flow in her distal left anterior descending coronary artery had markedly improved in the chronic phase. If the reduced blood flow in her distal left anterior descending coronary artery was induced by coronary vasospasm and the vasospasm was relieved before the coronary angiogram was performed, this case must be diagnosed as left ventricular apical ballooning syndrome. CONCLUSION: We think this case may promote discussion regarding the pathophysiology of left ventricular apical ballooning syndrome.

Corticosteroid therapy was effective in controlling refractory coronary vasospasms complicated by hypereosinophilia.

A 48-year-old man suffered from uncontrollable coronary vasospasms, even when taking the maximum dose of vasodilators. The patient had a history of hypereosinophilia, and as the eosinophilia worsened, more frequent and intense coronary spastic angina (CSA) attacks occurred. He was treated with 20 mg/day of oral prednisolone, and the chest symptoms of CSA completely resolved thereafter. We encountered a refractory CSA patient with an allergic predisposition for which the oral administration of corticosteroids was markedly effective. Although the priority of corticosteroid therapy is not clinically high in patients with CSA, it can be effective especially in patients with an allergic background.