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G protein-coupled receptors (GPCRs) are a group of seven-transmembrane receptor proteins that have proven to be successful drug targets. Antibodies are becoming an increasingly promising modality to target these receptors due to their unique properties, such as exquisite specificity, long half-life, and fewer side effects, and their improved pharmacokinetic and pharmacodynamic profiles compared to peptides and small molecules, which results from their more favorable biodistribution. To date, there are only two US Food and Drug Administration-approved GPCR antibody drugs, namely erenumab and mogamulizumab, and this highlights the challenges encountered in identifying functional antibodies against GPCRs. Utilizing Twists precision DNA writing technologies, we have created a GPCR-focused phage display library with 1 × 1010 diversity. Specifically, we mined endogenous GPCR binding ligand and peptide sequences and incorporated these binding motifs into the heavy chain complementarity-determining region 3 in a synthetic antibody library. Glucagon-like peptide-1 receptor (GLP-1 R) is a class B GPCR that acts as the receptor for the incretin GLP-1, which is released to regulate insulin levels in response to food intake. GLP-1 R agonists have been widely used to increase insulin secretion to lower blood glucose levels for the treatment of type 1 and type 2 diabetes, whereas GLP-1 R antagonists have applications in the treatment of severe hypoglycemia associated with bariatric surgery and hyperinsulinomic hypoglycemia. Here we present the discovery and creation of both antagonistic and agonistic GLP-1 R antibodies by panning this GPCR-focused phage display library on a GLP-1 R-overexpressing Chinese hamster ovary cell line and demonstrate their in vitro and in vivo functional activity.

New immune checkpoints are emerging in a bid to improve response rates to immunotherapeutic drugs. The adenosine A2A receptor (A2AR) has been proposed as a target for immunotherapeutic development due to its participation in immunosuppression of the tumor microenvironment. Blockade of A2AR could restore tumor immunity and, consequently, improve patient outcomes. Here, we describe the discovery of a potent, selective, and tumor-suppressing antibody antagonist of human A2AR (hA2AR) by phage display. We constructed and screened four single-chain variable fragment (scFv) libraries—two synthetic and two immunized—against hA2AR and antagonist-stabilized hA2AR. After biopanning and ELISA screening, scFv hits were reformatted to human IgG and triaged in a series of cellular binding and functional assays to identify a lead candidate. Lead candidate TB206-001 displayed nanomolar binding of hA2AR-overexpressing HEK293 cells; cross-reactivity with mouse and cynomolgus A2AR but not human A1, A2B, or A3 receptors; functional antagonism of hA2AR in hA2AR-overexpressing HEK293 cells and peripheral blood mononuclear cells (PBMCs); and tumor-suppressing activity in colon tumor-bearing HuCD34-NCG mice. Given its therapeutic properties, TB206-001 is a good candidate for incorporation into next-generation bispecific immunotherapeutics. [ABSTRACT FROM AUTHOR]

The presence of the acyl dihydroxyacetone phosphate (acyl DHAP) pathway enzymes in Saccharomyces cerevisiae was reported. Total membrane fraction of S. cerevisiae exhibited DHAP acyltransferase, sn-glycerol 3-phosphate acyltransferase and acyl/alkyl DHAP reductase. It was shown that expression of these three enzymes is regulated by oxygen. Moreover, the results indicate that the acyl DHAP pathway may be important for non-ether glycerolipid synthesis in this yeast.

Counting intraepithelial lymphocytes (IELs) is a key part of the assessment of duodenal biopsies. Immunohistochemistry (IHC) for CD3 can aid identification of lymphocytes in this context, but it is not evident that counts on hematoxylin and eosin (H&E) and CD3 are comparable. This study aimed to compare the IEL counts in duodenal biopsies using H&E stains and CD3 IHC, and to examine the interobserver variability. Thirty-five paired H&E and CD3 sections were reviewed by 6 pathologists who counted the number of IELs per 100 enterocytes. The counts were categorized into groups: normal (<25 lymphocytes), mildly raised (25-40 lymphocytes), and markedly raised (>40 lymphocytes). CD3 IHC was associated with significantly higher IEL counts than H&E. Four cases with normal H&E counts had raised counts with CD3. There was moderate agreement between observers for both H&E and CD3. Lack of concordance between CD3 and H&E IEL counts suggests that counts derived from the 2 methods may not be comparable to each other and should not be considered equivalent. There was no significant improvement in interobserver variability with CD3 IHC. [ABSTRACT FROM AUTHOR]

Objectives: As thromboembolism (TE) continues to be one of the principal causes of death in obstetrical patients and as the postpartum period is associated with the highest risk for TE, we sought to determine the risk factors associated with TE following cesarean section (CS). Methods: A retrospective analysis of patients who had CS at a large tertiary referral center was conducted. Patients were identified through hospital medical records and were contacted approximately 1 year following their CS. Medical records and a questionnaire were used to identify features that were potentially associated with TE. Univariate analysis was used to determine the risk associated with these characteristics. Results: A total of 2206 patients had a CS, of which 1377 (62%) participated. Of the respondents, 137 patients received heparin (94% received a prophylactic dose, 6% received a therapeutic dose) and the remainder, 1233 patients, did not receive heparin. Seven patients (0.5%) developed a TE and 86% developed a TE within 7 days of CS. The odds ratio (OR) for TE for women with hypertension prior to pregnancy compared to patients who did not receive anticoagulation was 21.28 [95% confidence interval (CI) 4.64-90.13] and for patients who had varicose veins with superficial thrombophlebitis when compared to patients who had received heparin postpartum was 21.01 (95% CI 1.55-288.24). Discussion: Hypertension and the presence of varicose veins were associated with TE following CS. Larger cohort analyses are required to confirm these associations so that risk scores incorporating these characteristics may accurately predict the occurrence of TE. [ABSTRACT FROM AUTHOR]

Reactive oxygen species (ROS) activate multiple signaling pathways involved in cardiac hypertrophy. We showed that hemin reduces hypertrophy in response to pressure overload, in association with increased HO-1 and decreased ROS. Since HO-I exerts potent antioxidant effects, we hypothesized that this enzyme inhibits ROS-induced cardiomyocyte hypertrophy. HL-1 cardiomyocytes were transduced with adenovirus expressing HO-1 (AdHO-l) or treated with 25µM hemin for 24 hrs to increase basal HO-1 expression, and then exposed to 200 µM H2O2. Hypertrophy was measured using planar morphometry and ³H-leucine incorporation 48 hr after H2O2 exposure. H2O2 induced myocyte hypertrophy in a dose-dependent manner. The hypertrophic effect of H2O2 was partially reversed by N-acetyl cysteine. Hemin decreased ROS generation and reduced H2O2-induced increases in cell size and leucine incorporation. The effect of heroin was mimicked by AdHO-1 and bilirubin, and was inhibited by HO-1 siRNA. HO-1 overexpression significantly reduced pro-oxidant-induced increase in promoter and DNA binding activities of NF-κB, and inhibition of NF-κB nuclear translocation with SN50 inhibited pro-oxidant induced cardiomyocyte hypertrophy, suggesting that the anti-hypertrophic effects HO-1 may be mediated, at least in part, by inhibition of NF-κB. These results demonstrate that HO-1 inhibits pro-oxidant induced cardiomyocyte hypertrophy and suggest that HO-1 may yield therapeutic potential in treatment of cardiac hypertrophy and prevention of heart failure. [ABSTRACT FROM AUTHOR]

In May 2023, the Detroit Health Department was notified of four cases of invasive nontypeable Haemophilus influenzae (Hi) disease among students attending the same elementary school and grade, all with illness onsets within 7 days. Three patients were hospitalized, and one died. Most U.S. cases of invasive Hi disease are caused by nontypeable strains. No vaccines against nontypeable or non-type b Hi strains are currently available. Chemoprophylaxis is not typically recommended in response to nontypeable Hi cases; however, because of the high attack rate (four cases among 46 students; 8.7%), rifampin prophylaxis was recommended for household contacts of patients with confirmed cases and for all students and staff members in the school wing where confirmed cases occurred. Only 10.8% of students for whom chemoprophylaxis was recommended took it, highlighting gaps in understanding among caregivers and health care providers about persons for whom chemoprophylaxis was recommended. Public health authorities subsequently enhanced communication and education to the school community, improved coordination with health care partners, and established mass prophylaxis clinics at the school. This outbreak highlights the potential for nontypeable Hi to cause serious illness and outbreaks and the need for chemoprophylaxis guidance for nontypeable Hi disease. Achieving high chemoprophylaxis coverage requires education, communication, and coordination with community and health care partners., Competing Interests: All authors have completed and submitted the International Committee of Medical Journal Editors form for disclosure of potential conflicts of interest. No potential conflicts of interest were disclosed.

Competing Interests: Competing interests The BMJ has judged that there are no disqualifying financial ties to commercial companies. The authors declare the following other interests: none. Further details of The BMJ policy on financial interests are here: https://www.bmj.com/about-bmj/resources-authors/forms-policies-and-checklists/declaration-competing-interests

Hepatitis due to hepatitis B virus (HBV) reactivation can be severe and potentially fatal, but is preventable. HBV reactivation is most commonly reported in patients receiving cancer chemotherapy, especially rituximab-containing therapy for hematological malignancies and those receiving stem cell transplantation. All patients with hematological malignancies receiving anticancer therapy should be screened for active or resolved HBV infection by blood tests for hepatitis B surface antigen (HBsAg) and antibody to hepatitis B core antigen (anti-HBc). Patients found to be positive for HBsAg should be given prophylactic antiviral therapy to prevent HBV reactivation. For patients with resolved HBV infection, no standard strategy has yet been established to prevent HBV reactivation. There are usually two options. One is pre-emptive therapy guided by serial HBV DNA monitoring, whereby antiviral therapy is given as soon as HBV DNA becomes detectable. However, there is little evidence regarding the optimal interval and period of monitoring. An alternative approach is prophylactic antiviral therapy, especially for patients receiving high-risk therapy such as rituximab, newer generation of anti-CD20 monoclonal antibody, obinutuzumab or hematopoietic stem cell transplantation. This strategy may effectively prevent HBV reactivation and avoid the inconvenience of repeated HBV DNA monitoring. Entecavir or tenofovir are preferred over lamivudine as prophylactic therapy. Although there is no well-defined guideline on the optimal duration of prophylactic therapy, there is growing evidence to recommend continuing prophylactic antiviral therapy for at least 12 mo after cessation of chemotherapy, and even longer for those who receive rituximab or who had high serum HBV DNA levels before the start of immunosuppressive therapy. Many novel agents have recently become available for the treatment of hematological malignancies, and these agents may be associated with HBV reactivation. Although there is currently limited evidence to guide the optimal preventive measures, we recommend antiviral prophylaxis in HBsAg-positive patients receiving novel treatments, especially the Bruton tyrosine kinase inhibitors and the phosphatidylinositol 3-kinase inhibitors, which are B-cell receptor signaling modulators and reduce proliferation of malignant B-cells. Further studies are needed to clarify the risk of HBV reactivation with these agents and the best prophylactic strategy in the era of targeted therapy for hematological malignancies.