Type II & III inhibitors of tropomyosin receptor kinase (Trk): a 2020-2022 patent update.

INTRODUCTION: The Trk family proteins are membrane-bound kinases predominantly expressed in neuronal tissues. Activated by neurotrophins, they regulate critical cellular processes through downstream signaling pathways. Dysregulation of Trk signaling can drive a range of diseases, making the design and study of Trk inhibitors a vital area of research. This review explores recent advances in the development of type II and III Trk inhibitors, with implications for various therapeutic applications. AREAS COVERED: Patents covering type II and III inhibitors targeting the Trk family are discussed as a complement of the previous review, Type I inhibitors of tropomyosin receptor kinase (Trk): a 2020-2022 patent update. Relevant patents were identified using the Web of Science database, Google, and Google Patents. EXPERT OPINION: While type II and III Trk inhibitor development has advanced more gradually compared to their type I counterparts, they hold significant promise in overcoming resistance mutations and achieving enhanced subtype selectivity - a critical factor in reducing adverse effects associated with pan-Trk inhibition. Recent interdisciplinary endeavors have marked substantial progress in the design of subtype selective Trk inhibitors, with impressive success heralded by the type III inhibitors. Notably, the emergence of mutant-selective Trk inhibitors introduces an intriguing dimension to the field, offering precise treatment possibilities.

Gastrointestinal Conditions: Acute Infectious Gastroenteritis and Colitis.

Gastroenteritis is inflammation of the stomach and intestines; colitis is inflammation of the colon. Viruses are the most common cause, followed by bacteria and parasites. Incidence of the various infections varies by age, sex, location, and vaccine availability; vaccination has reduced rotavirus infections by as much as 90% in children. Postinfectious complications include irritable bowel syndrome (IBS) and lactose intolerance. Approximately 9% of patients with acute gastroenteritis or colitis develop postinfectious IBS, which accounts for more than 50% of all IBS cases. The diagnostic approach to gastroenteritis and colitis varies with symptom severity. Microbial studies are not needed with mild symptoms that resolve within a week, but longer-lasting or more severe symptoms (including bloody stool) warrant microbial studies. In addition, recent antibiotic exposure should prompt testing for Clostridioides difficile. Multiplex antimicrobial testing is preferred; stool cultures and microscopic stool examinations are no longer first-line tests. Management depends on severity. Patients with mild or moderate symptoms are treated with oral hydration if tolerated; nasogastric or intravenous hydration are used for those with more severe illness. In addition, antiemetic, antimotility, and/or antisecretory drugs can be used for symptom control. Antimicrobial therapy is indicated for C difficile infections, travel-related diarrhea, other bacterial infections with severe symptoms, and parasitic infections.

Gastrointestinal Conditions: Pancreatitis.

Acute pancreatitis is among the most common gastrointestinal disorders requiring hospitalization. The main causes are gallstones and alcohol use. Patients typically present with upper abdominal pain radiating to the back, worse with eating, plus nausea and vomiting. Diagnosis requires meeting two of three criteria: upper abdominal pain, an elevated serum lipase or amylase level greater than 3 times the normal limit, and imaging findings consistent with pancreatitis. After pancreatitis is diagnosed, the Atlanta classification and identification of the systemic inflammatory response syndrome can identify patients at high risk of complications. Management includes fluid resuscitation and hydration maintenance, pain control that may require opioids, and early feeding. Feeding recommendations have changed and "nothing by mouth" is no longer recommended. Rather, oral feeding should be initiated, as tolerated, within the first 24 hours. If it is not tolerated, enteral feeding via nasogastric or nasojejunal tubes should be initiated. Antibiotics are indicated only with radiologically confirmed infection or systemic infection symptoms. Surgical or endoscopic interventions are needed for biliary pancreatitis or obstructive pancreatitis with cholangitis. One in five patients will have recurrent episodes of pancreatitis; alcohol and smoking are major risk factors. Some develop chronic pancreatitis, associated with chronic pain plus pancreatic dysfunction, including endocrine failure (insulin insufficiency) and/or exocrine failure that requires long-term vitamin supplementation.

Gastrointestinal Conditions: Gastroesophageal Reflux Disease.

Gastroesophageal reflux disease (GERD) affects more than 20% of adults. Risk factors include older age, obesity, smoking, and sedentary lifestyle. Lower esophageal sphincter (LES) dysfunction is a primary cause. Classic symptoms include heartburn and regurgitation. With classic symptoms, proton pump inhibitors (PPIs) can be prescribed without further testing; PPIs should be taken on an empty stomach. Patients with atypical symptoms and those not benefiting from management should undergo esophagogastroduodenoscopy (EGD), and potentially pH and impedance testing to confirm GERD or identify other conditions. This is important because GERD increases risk of esophageal erosions/stricture, Barrett esophagus, and esophageal adenocarcinoma. However, a large percentage of adults taking PPIs have no clear indication for treatment, and PPIs and other antisecretory therapy should be tapered off if possible. Of note, vonoprazan, a new drug approved by the Food and Drug Administration (FDA), has shown superiority to PPIs. In addition to pharmacotherapy, lifestyle changes are indicated, including losing weight if overweight, not lying down after meals, and ceasing tobacco use. Procedural interventions, including fundoplication and magnetic sphincter augmentation, can be considered for patients wishing to discontinue drugs or with symptoms unresponsive to PPIs. Procedural interventions are effective for the first 1 to 3 years, but effectiveness decreases over time.

Gastrointestinal Conditions: Peptic Ulcer Disease.

Peptic ulcer disease (PUD) involves ulceration of the mucosa in the stomach and/or proximal duodenum. The main causes are Helicobacter pylori infection and nonsteroidal anti-inflammatory drug (NSAID) use. PUD occurs in 5% to 10% of people worldwide, but rates have decreased by more than half during the past 20 years. This reduction is thought to be because of H pylori management, more conservative use of NSAIDs, and/or widespread use of proton pump inhibitors (PPIs). Common symptoms include postprandial abdominal pain, nausea, vomiting, and weight loss. These symptoms have broad overlap with those of other conditions, making clinical diagnosis difficult. Endoscopy is the gold standard for diagnosis, especially in older patients and those with alarm symptoms, but a test-and-treat strategy (noninvasive test for H pylori and treat if positive) can be used for younger patients with no alarm symptoms. Numerous treatment regimens are available, all of which include PPIs plus antibiotics. As an alternative to PPIs, a new triple therapy with vonoprazan (which blocks acid production) plus antibiotics has been approved and appears to be superior to conventional therapy with PPIs plus antibiotics. At least 4 weeks after treatment, repeat testing for H pylori should be obtained to confirm cure. When possible, NSAIDs should be discontinued; when not possible, antisecretory cotherapy should be considered.

Convergent evolution and targeting of diverse E2 epitopes by human broadly neutralizing antibodies are associated with HCV clearance.

The early appearance of broadly neutralizing antibodies (bNAbs) in serum is associated with spontaneous hepatitis C virus (HCV) clearance, but to date, the majority of bNAbs have been isolated from chronically infected donors. Most of these bNAbs use the VH1-69 gene segment and target the envelope glycoprotein E2 front layer. Here, we performed longitudinal B cell receptor (BCR) repertoire analysis on an elite neutralizer who spontaneously cleared multiple HCV infections. We isolated 10,680 E2-reactive B cells, performed BCR sequencing, characterized monoclonal B cell cultures, and isolated bNAbs. In contrast to what has been seen in chronically infected donors, the bNAbs used a variety of VH genes and targeted at least three distinct E2 antigenic sites, including sites previously thought to be non-neutralizing. Diverse front-layer-reactive bNAb lineages evolved convergently, acquiring breadth-enhancing somatic mutations. These findings demonstrate that HCV clearance-associated bNAbs are genetically diverse and bind distinct antigenic sites that should be the target of vaccine-induced bNAbs.

Neutralizing antibodies evolve to exploit vulnerable sites in the HCV envelope glycoprotein E2 and mediate spontaneous clearance of infection.

Individuals who clear primary hepatitis C virus (HCV) infections clear subsequent reinfections more than 80% of the time, but the mechanisms are poorly defined. Here, we used HCV variants and plasma from individuals with repeated clearance to characterize longitudinal changes in envelope glycoprotein E2 sequences, function, and neutralizing antibody (NAb) resistance. Clearance of infection was associated with early selection of viruses with NAb resistance substitutions that also reduced E2 binding to CD81, the primary HCV receptor. Later, peri-clearance plasma samples regained neutralizing capacity against these variants. We identified a subset of broadly NAbs (bNAbs) for which these loss-of-fitness substitutions conferred resistance to unmutated bNAb ancestors but increased sensitivity to mature bNAbs. These data demonstrate a mechanism by which neutralizing antibodies contribute to repeated immune-mediated HCV clearance, identifying specific bNAbs that exploit fundamental vulnerabilities in E2. The induction of bNAbs with these specificities should be a goal of HCV vaccine development.

Managing Difficult Patient Encounters.

Family physicians commonly find themselves in difficult patient encounters that can result in dissatisfaction for the patient and physician. Successful navigation of these encounters includes recognizing common physician factors, such as systemic pressures, interpersonal communication, and situational issues. The practice of labeling patient types can lead to disparities in care and patient harm and should be avoided. When physicians recognize that they are in a difficult patient encounter, simple mindfulness approaches, such as the Name It to Tame It and CALMER approaches, can improve outcomes. CALMER approaches help physicians acknowledge which situations they can control, alter their thoughts about the situation, and tolerate uncertainty. Physicians working with patients to create a therapeutic bond can focus the encounter to understand the situation that the patient is experiencing and work to recognize and acknowledge strong emotions that are nonproductive. Negotiating an agenda can help manage expectations of what can reasonably be done during each visit. Supporting patients by validating their symptoms and helping them embrace uncertainty can enable them to take control of their diagnosis and focus on managing chronic conditions rather than curing them. Motivational interviewing is a useful tool to help patients take ownership of their illnesses and therapeutic goals. Self-care through reflection groups or personal coaching or counseling can help physicians feel supported and avoid burnout.

Three doses of COVID-19 mRNA vaccine induce class-switched antibody responses in inflammatory arthritis patients on immunomodulatory therapies.

Patients with inflammatory arthritis (IA) are at increased risk of severe COVID-19 due to medication-induced immunosuppression that impairs host defenses. The aim of this study was to assess antibody and B cell responses to COVID-19 mRNA vaccination in IA patients receiving immunomodulatory therapies. Adults with IA were enrolled through the Johns Hopkins Arthritis Center and compared with healthy controls (HC). Paired plasma and peripheral blood mononuclear cell (PBMC) samples were collected prior to and 30 days or 6 months following the first two doses of mRNA vaccines (D2; HC=77 and IA=31 patients), or 30 days following a third dose of mRNA vaccines (D3; HC=11 and IA=96 patients). Neutralizing antibody titers, total binding antibody titers, and B cell responses to vaccine and Omicron variants were analyzed. Anti-Spike (S) IgG and S-specific B cells developed appropriately in most IA patients following D3, with reduced responses to Omicron variants, and negligible effects of medication type or drug withholding. Neutralizing antibody responses were lower compared to healthy controls after both D2 and D3, with a small number of individuals demonstrating persistently undetectable neutralizing antibody levels. Most IA patients respond as well to mRNA COVID-19 vaccines as immunocompetent individuals by the third dose, with no evidence of improved responses following medication withholding. These data suggest that IA-associated immune impairment may not hinder immunity to COVID-19 mRNA vaccines in most individuals.

Type I inhibitors of tropomyosin receptor kinase (Trk): a 2020-2022 patent update.

INTRODUCTION: Trk inhibitors are significant in the realm of personalized medicine as they target specific genetic alterations, such as NTRK gene fusions, leading to improved treatment outcomes for cancer patients. By tailoring the treatment to the genetic characteristics of the tumor rather than the tumor type, Trk inhibitors offer the potential for more effective and precise therapies, resulting in enhanced response rates and prolonged survival for patients with NTRK fusion-positive cancers. AREAS COVERED: Patents covering type I inhibitors targeting the Trk family are discussed, building upon our prior review series on Trk inhibitors. Relevant patents were identified through the Web of Science database, Google, and Google Patents. EXPERT OPINION: The field of Trk inhibitors has evolved significantly, as reflected in the current patent literature, which emphasizes the selective structural refinement of clinical champions. Efforts now concentrate on enhancing efficacy against on-target resistance mechanisms, with modifications made to improve potency, reduce toxicity, and enhance pharmacokinetics. Combination therapies show potential to address off-target resistance mechanisms and improve treatment outcomes. Challenges remain in accurately diagnosing NTRK gene alterations and integrating screening into routine clinical practice. Trk inhibitors have surpassed their conventional role of inhibition and are now seeing new applications in radiopharmaceutical development and as molecular targeting agents.

Corrigendum: Convergent antibody responses are associated with broad neutralization of hepatitis C virus.

[This corrects the article DOI: 10.3389/fimmu.2023.1135841.].

Convergent antibody responses are associated with broad neutralization of hepatitis C virus.

Introduction: Early development of broadly neutralizing antibodies (bNAbs) targeting the hepatitis C virus (HCV) envelope glycoprotein E2 is associated with spontaneous clearance of infection, so induction of bNAbs is a major goal of HCV vaccine development. However, the molecular antibody features important for broad neutralization are not known. Methods: To identify B cell repertoire features associated with broad neutralization, we performed RNA sequencing of the B cell receptors (BCRs) of HCV E2-reactive B cells of HCV-infected individuals with either high or low plasma neutralizing breadth. We then produced a monoclonal antibody (mAb) expressed by pairing the most abundant heavy and light chains from public clonotypes identified among clearance, high neutralization subjects. Results: We found distinctive BCR features associated with broad neutralization of HCV, including long heavy chain complementarity determining region 3 (CDRH3) regions, specific VH gene usage, increased frequencies of somatic hypermutation, and particular VH gene mutations. Most intriguing, we identified many E2-reactive public BCR clonotypes (heavy and light chain clones with the same V and J-genes and identical CDR3 sequences) present only in subjects who produced highly neutralizing plasma. The majority of these public clonotypes were shared by two subjects who cleared infection. A mAb expressing the most abundant public heavy and light chains from these clearance, high neutralization subjects had features enriched in high neutralization clonotypes, such as increased somatic hypermutation frequency and usage of IGHV1-69, and was cross-neutralizing. Discussion: Together, these results demonstrate distinct BCR repertoires associated with high plasma neutralizing capacity. Further characterization of the molecular features and function of these antibodies can inform HCV vaccine development.

Persistent SARS-CoV-2-specific immune defects in kidney transplant recipients following third mRNA vaccine dose.

Kidney transplant recipients (KTRs) show poorer response to SARS-CoV-2 mRNA vaccination, yet response patterns and mechanistic drivers following third doses are ill-defined. We administered third monovalent mRNA vaccines to n = 81 KTRs with negative or low-titer anti-receptor binding domain (RBD) antibody (n = 39 anti-RBDNEG; n = 42 anti-RBDLO), compared with healthy controls (HCs, n = 19), measuring anti-RBD, Omicron neutralization, spike-specific CD8+%, and SARS-CoV-2-reactive T cell receptor (TCR) repertoires. By day 30, 44% anti-RBDNEG remained seronegative; 5% KTRs developed BA.5 neutralization (vs 68% HCs, P < .001). Day 30 spike-specific CD8+% was negative in 91% KTRs (vs 20% HCs; P = .07), without correlation to anti-RBD (rs = 0.17). Day 30 SARS-CoV-2-reactive TCR repertoires were detected in 52% KTRs vs 74% HCs (P = .11). Spike-specific CD4+ TCR expansion was similar between KTRs and HCs, yet KTR CD8+ TCR depth was 7.6-fold lower (P = .001). Global negative response was seen in 7% KTRs, associated with high-dose MMF (P = .037); 44% showed global positive response. Of the KTRs, 16% experienced breakthrough infections, with 2 hospitalizations; prebreakthrough variant neutralization was poor. Absent neutralizing and CD8+ responses in KTRs indicate vulnerability to COVID-19 despite 3-dose mRNA vaccination. Lack of neutralization despite CD4+ expansion suggests B cell dysfunction and/or ineffective T cell help. Development of more effective KTR vaccine strategies is critical. (NCT04969263).

Functional HIV-1/HCV cross-reactive antibodies isolated from a chronically co-infected donor.

Despite prolific efforts to characterize the antibody response to human immunodeficiency virus type 1 (HIV-1) and hepatitis C virus (HCV) mono-infections, the response to chronic co-infection with these two ever-evolving viruses is poorly understood. Here, we investigate the antibody repertoire of a chronically HIV-1/HCV co-infected individual using linking B cell receptor to antigen specificity through sequencing (LIBRA-seq). We identify five HIV-1/HCV cross-reactive antibodies demonstrating binding and functional cross-reactivity between HIV-1 and HCV envelope glycoproteins. All five antibodies show exceptional HCV neutralization breadth and effector functions against both HIV-1 and HCV. One antibody, mAb688, also cross-reacts with influenza and coronaviruses, including severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). We examine the development of these antibodies using next-generation sequencing analysis and lineage tracing and find that somatic hypermutation established and enhanced this reactivity. These antibodies provide a potential future direction for therapeutic and vaccine development against current and emerging infectious diseases. More broadly, chronic co-infection represents a complex immunological challenge that can provide insights into the fundamental rules that underly antibody-antigen specificity.

Impact of Seasonal Coronavirus Antibodies on Severe Acute Respiratory Syndrome Coronavirus 2 Vaccine Responses in Solid Organ Transplant Recipients.

Antibody responses to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) vaccination are reduced in solid organ transplant recipients (SOTRs). We report that increased levels of preexisting antibodies to seasonal coronaviruses are associated with decreased antibody response to SARS-CoV-2 vaccination in SOTRs, supporting that antigenic imprinting modulates vaccine responses in SOTRs.

Serum neutralization activity declines but memory B cells persist after cure of chronic hepatitis C.

The increasing incidence of hepatitis C virus (HCV) infections underscores the need for an effective vaccine. Successful vaccines to other viruses generally depend on a long-lasting humoral response. However, data on the half-life of HCV-specific responses are lacking. Here we study archived sera and mononuclear cells that were prospectively collected up to 18 years after cure of chronic HCV infection to determine the role of HCV antigen in maintaining neutralizing antibody and B cell responses. We show that HCV-neutralizing activity decreases rapidly in potency and breadth after curative treatment. In contrast, HCV-specific memory B cells persist, and display a restored resting phenotype, normalized chemokine receptor expression and preserved ability to differentiate into antibody-secreting cells. The short half-life of HCV-neutralizing activity is consistent with a lack of long-lived plasma cells. The persistence of HCV-specific memory B cells and the reduced inflammation after cure provide an opportunity for vaccination to induce protective immunity against re-infection.

Diverticular Disease: Rapid Evidence Review.

Diverticulitis should be suspected in patients with isolated left lower quadrant pain, abdominal distention or rigidity, fever, and leukocytosis. Initial laboratory workup includes a complete blood count, basic metabolic panel, urinalysis, and C-reactive protein measurement. Computed tomography with intravenous contrast is the preferred imaging modality, if needed to confirm diagnosis and assess for complications of diverticulitis. Treatment decisions are based on the categorization of disease as complicated vs. uncomplicated. Selected patients with uncomplicated diverticulitis may be treated without antibiotics. Complicated diverticulitis is treated in the hospital with modified diet or bowel rest, antibiotics, and pain control. Abscesses that are 3 cm or larger should be treated with percutaneous drainage. Emergent surgery is reserved for when percutaneous drainage fails or the patient's clinical condition worsens despite adequate therapy. Colonoscopy should not be performed during the flare-up, but should be considered six weeks after resolution of symptoms in patients with complicated diverticulitis who have not had a high-quality colonoscopy in the past year. Diverticulitis prevention measures include consuming a vegetarian diet or high-quality diet (high in fruits, vegetables, whole grains, and legumes), limiting red meat and sweets, achieving or maintaining a body mass index of 18 to 25 kg per m2, being physically active, and avoiding tobacco and long-term nonsteroidal anti-inflammatory drugs. Partial colectomy is not routinely recommended for diverticulitis prevention and should be reserved for patients with more than three recurrences or abscess formation requiring percutaneous drainage.

Toward in vivo proof of binding of 18F-labeled inhibitor [18F]TRACK to peripheral tropomyosin receptor kinases.

BACKGROUND: Tropomyosin receptor kinases (TrkA, TrkB, TrkC) are a family of tyrosine kinases primarily expressed in neuronal cells of the brain. Identification of oncogenic alterations in Trk expression as a driver in multiple tumor types has increased interest in their role in human cancers. Recently, first- and second-generation 11C and 18F-labeled Trk inhibitors, e.g., [18F]TRACK, have been developed. The goal of the present study was to analyze the direct interaction of [18F]TRACK with peripheral Trk receptors in vivo to prove its specificity for use as a functional imaging probe. METHODS: In vitro uptake and competition experiments were carried out using the colorectal cancer cell line KM12. Dynamic PET experiments were performed with [18F]TRACK, either alone or in the presence of amitriptyline, an activator of Trk, entrectinib, a Trk inhibitor, or unlabeled reference compound TRACK in KM12 tumor-bearing athymic nude mice as well as B6129SF2/J and corresponding B6;129S2-Ntrk2tm1Bbd/J mice. Western blot and immunohistochemistry experiments were done with KM12 tumors, brown adipose tissue (BAT), and brain tissue samples. RESULTS: Uptake of [18F]TRACK was increasing over time reaching 208 ± 72% radioactivity per mg protein (n = 6/2) after 60 min incubation time. Entrectinib and TRACK competitively blocked [18F]TRACK uptake in vitro (IC50 30.9 ± 3.6 and 29.4 ± 9.4 nM; both n = 6/2). [18F]TRACK showed uptake into KM12 tumors (SUVmean,60 min 0.43 ± 0.03; n = 6). Tumor-to-muscle ratio reached 0.9 (60 min) and 1.2 (120 min). In TrkB expressing BAT, [18F]TRACK uptake reached SUVmean,60 min 1.32 ± 0.08 (n = 7). Activation of Trk through amitriptyline resulted in a significant radioactivity increase of 21% in KM12 tumor (SUVmean,60 min from 0.53 ± 0.01 to 0.43 ± 0.03; n = 6; p < 0.05) and of 21% in BAT (SUVmean,60 min from 1.32 ± 0.08; n = 5 to 1.59 ± 0.07; n = 6; p < 0.05) respectively. Immunohistochemistry showed TrkB > TrkA expression on BAT fat cells, but TrkA > TrkB in whole brain. WB analysis showed sevenfold higher TrkB expression in BAT versus KM12 tumor tissue. CONCLUSION: The present data show that radiotracer [18F]TRACK can target peripheral Trk receptors in human KM12 colon cancer as well as brown adipose tissue as confirmed through in vitro and in vivo blocking experiments. Higher TrkB versus TrkA protein expression was detected in brown adipose tissue of mice confirming a peripheral functional role of brain-derived neurotrophic factor in adipose tissue.

Computational identification of HCV neutralizing antibodies with a common HCDR3 disulfide bond motif in the antibody repertoires of infected individuals.

Despite recent success in hepatitis C virus (HCV) treatment using antivirals, an HCV vaccine is still needed to prevent reinfections in treated patients, to avert the emergence of drug-resistant strains, and to provide protection for people with no access to the antiviral therapeutics. The early production of broadly neutralizing antibodies (bNAbs) associates with HCV clearance. Several potent bNAbs bind a conserved HCV glycoprotein E2 epitope using an unusual heavy chain complementarity determining region 3 (HCDR3) containing an intra-loop disulfide bond. Isolation of additional structurally-homologous bNAbs would facilitate the recognition of key determinants of such bNAbs and guide rational vaccine design. Here we report the identification of new antibodies containing an HCDR3 disulfide bond motif using computational screening with the Rosetta software. Using the newly-discovered and already-known members of this antibody family, we review the required HCDR3 amino acid composition and propose determinants for the bent versus straight HCDR3 loop conformation observed in these antibodies.