Global Interactome Mapping Reveals Pro-tumorigenic Interactions of NF-κB in Breast Cancer.

NF-κB pathway is involved in inflammation; however, recent data shows its role also in cancer development and progression, including metastasis. To understand the role of NF-κB interactome dynamics in cancer, we study the complexity of breast cancer interactome in luminal A breast cancer model and its rearrangement associated with NF-κB modulation. Liquid chromatography-mass spectrometry measurement of 160 size-exclusion chromatography fractions identifies 5460 protein groups. Seven thousand five hundred sixty eight interactions among these proteins have been reconstructed by PrInCE algorithm, of which 2564 have been validated in independent datasets. NF-κB modulation leads to rearrangement of protein complexes involved in NF-κB signaling and immune response, cell cycle regulation, and DNA replication. Central NF-κB transcription regulator RELA co-elutes with interactors of NF-κB activator PRMT5, and these complexes are confirmed by AlphaPulldown prediction. A complementary immunoprecipitation experiment recapitulates RELA interactions with other NF-κB factors, associating NF-κB inhibition with lower binding of NF-κB activators to RELA. This study describes a network of pro-tumorigenic protein interactions and their rearrangement upon NF-κB inhibition with potential therapeutic implications in tumors with high NF-κB activity.

A syndromic neurodevelopmental disorder caused by rare variants in PPFIA3.

PPFIA3 encodes the protein-tyrosine phosphatase, receptor-type, F-polypeptide-interacting-protein-alpha-3 (PPFIA3), which is a member of the LAR-protein-tyrosine phosphatase-interacting-protein (liprin) family involved in synapse formation and function, synaptic vesicle transport, and presynaptic active zone assembly. The protein structure and function are evolutionarily well conserved, but human diseases related to PPFIA3 dysfunction are not yet reported in OMIM. Here, we report 20 individuals with rare PPFIA3 variants (19 heterozygous and 1 compound heterozygous) presenting with developmental delay, intellectual disability, hypotonia, dysmorphisms, microcephaly or macrocephaly, autistic features, and epilepsy with reduced penetrance. Seventeen unique PPFIA3 variants were detected in 18 families. To determine the pathogenicity of PPFIA3 variants in vivo, we generated transgenic fruit flies producing either human wild-type (WT) PPFIA3 or five missense variants using GAL4-UAS targeted gene expression systems. In the fly overexpression assays, we found that the PPFIA3 variants in the region encoding the N-terminal coiled-coil domain exhibited stronger phenotypes compared to those affecting the C-terminal region. In the loss-of-function fly assay, we show that the homozygous loss of fly Liprin-α leads to embryonic lethality. This lethality is partially rescued by the expression of human PPFIA3 WT, suggesting human PPFIA3 function is partially conserved in the fly. However, two of the tested variants failed to rescue the lethality at the larval stage and one variant failed to rescue lethality at the adult stage. Altogether, the human and fruit fly data reveal that the rare PPFIA3 variants are dominant-negative loss-of-function alleles that perturb multiple developmental processes and synapse formation.

De novo design of high-affinity binders of bioactive helical peptides.

Many peptide hormones form an α-helix on binding their receptors1-4, and sensitive methods for their detection could contribute to better clinical management of disease5. De novo protein design can now generate binders with high affinity and specificity to structured proteins6,7. However, the design of interactions between proteins and short peptides with helical propensity is an unmet challenge. Here we describe parametric generation and deep learning-based methods for designing proteins to address this challenge. We show that by extending RFdiffusion8 to enable binder design to flexible targets, and to refining input structure models by successive noising and denoising (partial diffusion), picomolar-affinity binders can be generated to helical peptide targets by either refining designs generated with other methods, or completely de novo starting from random noise distributions without any subsequent experimental optimization. The RFdiffusion designs enable the enrichment and subsequent detection of parathyroid hormone and glucagon by mass spectrometry, and the construction of bioluminescence-based protein biosensors. The ability to design binders to conformationally variable targets, and to optimize by partial diffusion both natural and designed proteins, should be broadly useful.

Preventing new-onset heart failure: Intervening at stage A.

Heart failure (HF) prevention is an urgent public health need with national and global implications. Stage A HF patients do not show HF symptoms or structural heart disease but are at risk of HF development. There are no unique recommendations on detecting Stage A patients. Patients in Stage A are heterogeneous; many patients have different combinations of risk factors and, therefore, have markedly different absolute risks for HF. Comprehensive strategies to prevent HF at Stage A include intensive blood pressure lowering, adequate glycemic and lipid management, and heart-healthy behaviors (adopting Life's Essential 8). First and foremost, it is imperative to improve public awareness of HF risk factors and implement healthy lifestyle choices very early. In addition, recognize the HF risk-enhancing factors, which are nontraditional cardiovascular (CV) risk factors that identify individuals at high risk for HF (genetic susceptibility for HF, atrial fibrillation, chronic kidney disease, chronic liver disease, chronic inflammatory disease, sleep-disordered breathing, adverse pregnancy outcomes, radiation therapy, a history of cardiotoxic chemotherapy exposure, and COVID-19). Early use of biomarkers, imaging markers, and echocardiography (noninvasive measures of subclinical systolic and diastolic dysfunction) may enhance risk prediction among individuals without established CV disease and prevent chemotherapy-induced cardiomyopathy. Efforts are needed to address social determinants of HF risk for primordial HF prevention.Central illustrationPolicies developed by organizations such as the American Heart Association, American College of Cardiology, and the American Diabetes Association to reduce CV disease events must go beyond secondary prevention and encompass primordial and primary prevention.

Rare variants in PPFIA3 cause delayed development, intellectual disability, autism, and epilepsy.

PPFIA3 encodes the Protein-Tyrosine Phosphatase, Receptor-Type, F Polypeptide-Interacting Protein Alpha-3 (PPFIA3), which is a member of the LAR protein-tyrosine phosphatase-interacting protein (liprin) family involved in synaptic vesicle transport and presynaptic active zone assembly. The protein structure and function are well conserved in both invertebrates and vertebrates, but human diseases related to PPFIA3 dysfunction are not yet known. Here, we report 14 individuals with rare mono-allelic PPFIA3 variants presenting with features including developmental delay, intellectual disability, hypotonia, autism, and epilepsy. To determine the pathogenicity of PPFIA3 variants in vivo , we generated transgenic fruit flies expressing either human PPFIA3 wildtype (WT) or variant protein using GAL4-UAS targeted gene expression systems. Ubiquitous expression with Actin-GAL4 showed that the PPFIA3 variants had variable penetrance of pupal lethality, eclosion defects, and anatomical leg defects. Neuronal expression with elav-GAL4 showed that the PPFIA3 variants had seizure-like behaviors, motor defects, and bouton loss at the 3 rd instar larval neuromuscular junction (NMJ). Altogether, in the fly overexpression assays, we found that the PPFIA3 variants in the N-terminal coiled coil domain exhibited stronger phenotypes compared to those in the C-terminal region. In the loss-of-function fly assay, we show that the homozygous loss of fly Liprin- α leads to embryonic lethality. This lethality is partially rescued by the expression of human PPFIA3 WT, suggesting human PPFIA3 protein function is partially conserved in the fly. However, the PPFIA3 variants failed to rescue lethality. Altogether, the human and fruit fly data reveal that the rare PPFIA3 variants are dominant negative loss-of-function alleles that perturb multiple developmental processes and synapse formation.

Non-Redundant Roles of T Cell Costimulation Pathways in Inflammatory Arthritis Revealed by Dual Blockade of ICOS and CD28 with Acazicolcept (ALPN-101).

OBJECTIVE: CD28 and inducible T cell costimulator (ICOS) appear to have nonredundant roles in T cell activation and adaptive immunity. We undertook this study to characterize in vitro and in vivo the therapeutic potential of acazicolcept (ALPN-101), an Fc fusion protein of a human variant ICOS ligand (ICOSL) domain designed to inhibit both CD28 and ICOS costimulation, in inflammatory arthritis. METHODS: Acazicolcept was compared in vitro with inhibitors of either the CD28 or ICOS pathways (abatacept and belatacept [CTLA-4Ig], prezalumab [anti-ICOSL monoclonal antibody]) in receptor binding and signaling assays, and in a collagen-induced arthritis (CIA) model. Acazicolcept was also compared in cytokine and gene expression assays of peripheral blood mononuclear cells (PBMCs) from healthy donors or rheumatoid arthritis (RA) or psoriatic arthritis (PsA) patients stimulated with artificial antigen-presenting cells (APCs) expressing CD28 and ICOS ligands*. RESULTS: Acazicolcept bound CD28 and ICOS, prevented ligand binding, and inhibited human T cell functional interactions, matching or exceeding the activity of CD28 or ICOS costimulatory single-pathway inhibitors tested individually or in combination. Acazicolcept administration significantly reduced disease in the CIA model and more potently than abatacept. Acazicolcept also inhibited proinflammatory cytokine production from stimulated PBMCs in cocultures with artificial APCs and demonstrated unique effects on gene expression distinct from those induced by abatacept, prezalumab, or a combination of both. CONCLUSION: Both CD28 and ICOS signaling play critical roles in inflammatory arthritis. Therapeutic agents such as acazicolcept that coinhibit both ICOS and CD28 signaling may mitigate inflammation and/or disease progression in RA and PsA more effectively than inhibitors of either pathway alone.

A Multipronged Unbiased Strategy Guides the Development of an Anti-EGFR/EPHA2-Bispecific Antibody for Combination Cancer Therapy.

PURPOSE: Accumulating analyses of pro-oncogenic molecular mechanisms triggered a rapid development of targeted cancer therapies. Although many of these treatments produce impressive initial responses, eventual resistance onset is practically unavoidable. One of the main approaches for preventing this refractory condition relies on the implementation of combination therapies. This includes dual-specificity reagents that affect both of their targets with a high level of selectivity. Unfortunately, selection of target combinations for these treatments is often confounded by limitations in our understanding of tumor biology. Here, we describe and validate a multipronged unbiased strategy for predicting optimal co-targets for bispecific therapeutics. EXPERIMENTAL DESIGN: Our strategy integrates ex vivo genome-wide loss-of-function screening, BioID interactome profiling, and gene expression analysis of patient data to identify the best fit co-targets. Final validation of selected target combinations is done in tumorsphere cultures and xenograft models. RESULTS: Integration of our experimental approaches unambiguously pointed toward EGFR and EPHA2 tyrosine kinase receptors as molecules of choice for co-targeting in multiple tumor types. Following this lead, we generated a human bispecific anti-EGFR/EPHA2 antibody that, as predicted, very effectively suppresses tumor growth compared with its prototype anti-EGFR therapeutic antibody, cetuximab. CONCLUSIONS: Our work not only presents a new bispecific antibody with a high potential for being developed into clinically relevant biologics, but more importantly, successfully validates a novel unbiased strategy for selecting biologically optimal target combinations. This is of a significant translational relevance, as such multifaceted unbiased approaches are likely to augment the development of effective combination therapies for cancer treatment. See related commentary by Kumar, p. 2570.

Functional relationships between estradiol and paternal care in male red-bellied lemurs, Eulemur rubriventer.

Fathers contribute substantially to infant care, yet the mechanisms facilitating paternal bonding and interactions with infants are not as well understood as they are in mothers. Several hormonal changes occur as males transition into parenthood, first in response to a partner's pregnancy, and next in response to interacting with the newborn. These changes may prepare fathers for parenting and help facilitate and maintain paternal care. Experimental studies with monkeys and rodents suggest that paternal care requires elevated estradiol levels, which increase when a male's partner is pregnant and are higher in fathers than non-fathers, but its role in the expression of paternal behaviors throughout infant development is unknown. To assess estradiol's role in paternal care, we analyzed the relationship between paternal estradiol metabolites and 1) offspring age, and 2) paternal care behavior (holding, carrying, huddling, playing, grooming), in wild, red-bellied lemurs (Eulemur rubriventer). We collected 146 fecal samples and 1597 h of behavioral data on 10 adult males who had newborn infants during the study. Estradiol metabolites increased four-fold in expectant males, and in new fathers they fluctuated and gradually decreased with time. Infant age, not paternal behavior, best predicted hormone levels in new fathers. These results suggest that hormonal changes occur in expectant males with facultative paternal care, but they do not support the hypothesis that estradiol is directly associated with the day-to-day expression of paternal care. Future research should explore estradiol's role in facilitating behaviors, including infant-directed attention and responsiveness, or preparing fathers for infant care generally.

Povetacicept, an Enhanced Dual APRIL/BAFF Antagonist That Modulates B Lymphocytes and Pathogenic Autoantibodies for the Treatment of Lupus and Other B Cell-Related Autoimmune Diseases.

OBJECTIVE: Dysregulated APRIL/BAFF signaling is implicated in the pathogenesis of multiple autoimmune diseases, including systemic lupus erythematosus and lupus nephritis. We undertook this study to develop and evaluate a high-affinity APRIL/BAFF antagonist to overcome the clinical limitations of existing B cell inhibitors. METHODS: A variant of TACI-Fc generated by directed evolution showed enhanced binding for both APRIL and BAFF and was designated povetacicept (ALPN-303). Povetacicept was compared to wild-type (WT) TACI-Fc and related molecules in vitro and in vivo. RESULTS: Povetacicept inhibited APRIL and BAFF more effectively than all evaluated forms of WT TACI-Fc and selective APRIL and BAFF inhibitors in cell-based reporter assays and primary human B cell assays, mediating potent suppression of B cell proliferation, differentiation, and immunoglobulin (Ig) secretion. In mouse immunization models, povetacicept significantly reduced serum immunoglobulin titers and antibody-secreting cells more effectively than anti-CD20 monoclonal antibodies, WT TACI-Fc, or APRIL and BAFF inhibitors. In the NZB × NZW mouse lupus nephritis model, povetacicept significantly enhanced survival and suppressed proteinuria, anti-double-stranded DNA antibody titers, blood urea nitrogen, glomerulonephritis, and renal immunoglobulin deposition. In the bm12 mouse lupus model, povetacicept significantly reduced splenic plasmablasts, follicular helper T cells, and germinal center B cells. In non-human primates, povetacicept was well tolerated, exhibited high serum exposure, and significantly decreased serum IgM, IgA, and IgG levels after a single dose. CONCLUSION: Enhanced APRIL and BAFF inhibition by povetacicept led to greater inhibition of B cell populations critical for autoantibody production compared to WT TACI-Fc and CD20-, APRIL-, or BAFF-selective inhibitors. Potent, dual inhibition by povetacicept has the potential to significantly improve clinical outcomes in autoantibody-related autoimmune diseases.

The engineered CD80 variant fusion therapeutic davoceticept combines checkpoint antagonism with conditional CD28 costimulation for anti-tumor immunity.

Despite the recent clinical success of T cell checkpoint inhibition targeting the CTLA-4 and PD-1 pathways, many patients either fail to achieve objective responses or they develop resistance to therapy. In some cases, poor responses to checkpoint blockade have been linked to suboptimal CD28 costimulation and the inability to generate and maintain a productive adaptive anti-tumor immune response. To address this, here we utilize directed evolution to engineer a CD80 IgV domain with increased PD-L1 affinity and fuse this to an immunoglobulin Fc domain, creating a therapeutic (ALPN-202, davoceticept) capable of providing CD28 costimulation in a PD-L1-dependent fashion while also antagonizing PD-1 - PD-L1 and CTLA-4-CD80/CD86 interactions. We demonstrate that by combining CD28 costimulation and dual checkpoint inhibition, ALPN-202 enhances T cell activation and anti-tumor efficacy in cell-based assays and mouse tumor models more potently than checkpoint blockade alone and thus has the potential to generate potent, clinically meaningful anti-tumor immunity in humans.

FOXO Transcription Factors Are Required for Normal Somatotrope Function and Growth.

Understanding the molecular mechanisms underlying pituitary organogenesis and function is essential for improving therapeutics and molecular diagnoses for hypopituitarism. We previously found that deletion of the forkhead factor, Foxo1, in the pituitary gland early in development delays somatotrope differentiation. While these mice grow normally, they have reduced growth hormone expression and free serum insulin-like growth factor-1 (IGF1) levels, suggesting a defect in somatotrope function. FOXO factors show functional redundancy in other tissues, so we deleted both Foxo1 and its closely related family member, Foxo3, from the primordial pituitary. We find that this results in a significant reduction in growth. Consistent with this, male and female mice in which both genes have been deleted in the pituitary gland (dKO) exhibit reduced pituitary growth hormone expression and serum IGF1 levels. Expression of the somatotrope differentiation factor, Neurod4, is reduced in these mice. This suggests a mechanism underlying proper somatotrope function is the regulation of Neurod4 expression by FOXO factors. Additionally, dKO mice have reduced Lhb expression and females also have reduced Fshb and Prl expression. These studies reveal FOXO transcription factors as important regulators of pituitary gland function.

Curvivirga aplysinae gen. nov., sp. nov., a marine bacterium isolated from the sea sponge Aplysina fistularis.

A Gram-stain-negative, strictly aerobic, motile bacterium, designated strain RKSG073T, was isolated from the sea sponge Aplysina fistularis, collected off the west coast of San Salvador, The Bahamas. Cells were curved-to-spiral rods with single, bipolar (amphitrichous) flagella, oxidase- and catalase-positive, non-nitrate-reducing and required salt for growth. RKSG073T grew optimally at 30-37 °C, pH 6-7, and with 2-3 % (w/v) NaCl. The predominant fatty acids of RKSG073T were summed feature 8 (C18 : 1ω6c and/or C18 : 1ω7c) and C16 : 0. Major isoprenoid quinones were identified as Q-10 and Q-9. Phylogenetic analyses of nearly complete 16S rRNA genes and genome sequences positioned strain RKSG073T in a clade with its closest relative Aestuariispira insulae AH-MY2T (92.1 % 16S rRNA gene sequence similarity), which subsequently clustered with Hwanghaeella grinnelliae Gri0909T, Marivibrio halodurans ZC80T and type species of the genera Kiloniella, Thalassospira and Terasakiella. The DNA G+C content calculated from the genome of RKSG073T was 42.2 mol%. On the basis of phylogenetic distinctiveness and polyphasic analysis, here we propose that RKSG073T (culture deposit numbers: ATCC collection = TSD-74T, BCCM collection = LMG 29869T) represents the type strain of a novel genus and species within the family Kiloniellaceae, order Rhodospirillales and class Alphaproteobacteria, for which the name Curvivirga aplysinae gen. nov., sp. nov. is proposed.

Prospective Validation of the Preschool Confusion Assessment Method for the ICU to Screen for Delirium in Infants Less Than 6 Months Old.

OBJECTIVES: ICU delirium is a predictor of greater morbidity and higher mortality in the pediatric population. The diagnostic obstacles and validity of delirium monitoring among neonates and young infants have yet to be fully delineated. We sought to validate the Preschool Confusion Assessment Method for the ICU in neonates and young infants and determine delirium prevalence in this young population. DESIGN: Prospective cohort study to validate the Preschool Confusion Assessment Method for the ICU for the assessment of ICU delirium in neonates and young infants compared with the reference standard, Child and Adolescent Psychiatry. SETTING: Tertiary medical center PICU, including medical, surgical, and cardiac patients. PARTICIPANTS: Infants less than 6 months old admitted to the PICU regardless of admission diagnosis. MEASUREMENTS AND MAIN RESULTS: We enrolled 49 patients with a median age of 1.8 months (interquartile range, 0.7-4.1 mo), 82% requiring mechanical ventilation. Enrolled patients were assessed for delirium in blinded-fashion by the research team using the Preschool Confusion Assessment Method for the ICU and independently assessed by the psychiatry reference rater using Diagnostic and Statistical Manual of Mental Disorders-5 criteria. A total of 189 paired assessments were completed, and the Preschool Confusion Assessment Method for the ICU performed with a sensitivity of 95% (95% CI, 89-100%), specificity of 81% (68-90%), "negative and positive predictive values" of 97% (94-100%) and 69% (55-79%), respectively, compared with the reference rater. Delirium prevalence was 47%, with higher rates of 61% observed among neonates (< 1 mo old) and 39% among infants 1-6 months old. CONCLUSIONS: The Preschool Confusion Assessment Method for the ICU is a valid screening tool for delirium monitoring in infants less than 6 months old. Delirium screening was feasible in this population despite evolving neurocognition and arousal architecture. ICU delirium was prevalent among infants. The consequence of acute brain dysfunction during crucial neurocognitive development remains unclear. Future studies are necessary to determine the long-term impact of ICU delirium and strategies to reduce associated harm in critically ill infants.

An orthogonal methods assessment of topical drug concentrations in skin and the impact for risk assessment in the viable epidermis.

Systemic toxicity assessments for oral or parenteral drugs often utilize the concentration of drug in plasma to enable safety margin calculations for human risk assessment. For topical drugs, there is no standard method for measuring drug concentrations in the stratum basale of the viable epidermis. This is particularly important since the superficial part of the epidermis, the stratum corneum (SC), is nonviable and where most of a topically applied drug remains, never penetrating deeper into the skin. We investigated the relative concentrations of a prototype kinase inhibitor using punch biopsy, laser capture microdissection, and imaging mass spectrometry methods in the SC, stratum basale, and dermis of minipig skin following topical application as a cream formulation. The results highlight the value of laser capture microdissection and mass spectrometry imaging in quantifying the large difference in drug concentration across the skin and even within the epidermis, and supports use of these methods for threshold-based toxicity risk assessments in specific anatomic locations of the skin, like of the stratum basale.

APECED-Associated Hepatitis: Clinical, Biochemical, Histological and Treatment Data From a Large, Predominantly American Cohort.

BACKGROUND AND AIMS: Autoimmune polyendocrinopathy-candidiasis-ectodermal dystrophy (APECED), caused by autoimmune regulator (AIRE) mutations, manifests with chronic mucocutaneous candidiasis (CMC) and multisystem autoimmunity, most often hypoparathyroidism (HP) and adrenal insufficiency (AI). European cohorts previously reported a ~10% prevalence of APECED-associated hepatitis (APAH) with presentations ranging from asymptomatic laboratory derangements to fatal fulminant hepatic failure. Herein, we characterized APAH in a large APECED cohort from the Americas. APPROACH AND RESULTS: Forty-five consecutive patients with APECED were evaluated (2013-2015) at the National Institutes of Health (NIH; NCT01386437). Hepatology consultation assessed hepatic and autoimmune biomarkers and liver ultrasound in all patients. Liver biopsies evaluated autoimmune features and fibrosis. The 16S ribosomal RNA (rRNA) sequencing was performed in 35 patients' stools (12 with and 23 without APAH). Among 43 evaluable patients, 18 (42%) had APAH; in 33.3% of those with APAH, APAH occurred before developing classic APECED diagnostic criteria. At APAH diagnosis, the median age was 7.8 years, and patients manifested with aminotransferase elevation and/or hyperbilirubinemia. All patients with APAH were in clinical remission during their NIH evaluation while receiving immunomodulatory treatment. We found no difference in age, sex, or prevalence of CMC, AI, or HP between patients with or without APAH. Autoantibody positivity against aromatic L-amino acid decarboxylase, cytochrome P450 family 1 subfamily A member 2, histidine decarboxylase (HDC), bactericidal/permeability-increasing fold-containing B1, tryptophan hydroxlase, and 21-hydroxylase (21-OH), and the homozygous c.967_979del13 AIRE mutation were associated with APAH development. Classical serological biomarkers of autoimmune hepatitis (AIH) were only sporadically positive. AIH-like lymphoplasmacytic inflammation with mild fibrosis was the predominant histological feature. Stool microbiome analysis found Slackia and Acidaminococcus in greater abundance in patients with APAH. CONCLUSIONS: APAH is more common than previously described, may present early before classic APECED manifestations, and most often manifests with milder, treatment-responsive disease. Several APECED-associated autoantibodies, but not standard AIH-associated biomarkers, correlate with APAH.

ICOSL+ plasmacytoid dendritic cells as inducer of graft-versus-host disease, responsive to a dual ICOS/CD28 antagonist.

Acute graft-versus-host disease (aGVHD) remains a major complication of allogeneic hematopoietic cell transplantation (HCT). CD146 and CCR5 are proteins that mark activated T helper 17 (Th17) cells. The Th17 cell phenotype is promoted by the interaction of the receptor ICOS on T cells with ICOS ligand (ICOSL) on dendritic cells (DCs). We performed multiparametric flow cytometry in a cohort of 156 HCT recipients and conducted experiments with aGVHD murine models to understand the role of ICOSL+ DCs. We observed an increased frequency of ICOSL+ plasmacytoid DCs, correlating with CD146+CCR5+ T cell frequencies, in the 64 HCT recipients with gastrointestinal aGVHD. In murine models, donor bone marrow cells from ICOSL-deficient mice compared to those from wild-type mice reduced aGVHD-related mortality. Reduced aGVHD resulted from lower intestinal infiltration of pDCs and pathogenic Th17 cells. We transplanted activated human ICOSL+ pDCs along with human peripheral blood mononuclear cells into immunocompromised mice and observed infiltration of intestinal CD146+CCR5+ T cells. We found that prophylactic administration of a dual human ICOS/CD28 antagonist (ALPN-101) prevented aGVHD in this model better than did the clinically approved belatacept (CTLA-4-Fc), which binds CD80 (B7-1) and CD86 (B7-2) and interferes with the CD28 T cell costimulatory pathway. When started at onset of aGVHD signs, ALPN-101 treatment alleviated symptoms of ongoing aGVHD and improved survival while preserving antitumoral cytotoxicity. Our data identified ICOSL+-pDCs as an aGVHD biomarker and suggest that coinhibition of the ICOSL/ICOS and B7/CD28 axes with one biologic drug may represent a therapeutic opportunity to prevent or treat aGVHD.

Fulvivirga aurantia sp. nov. and Xanthovirga aplysinae gen. nov., sp. nov., marine bacteria isolated from the sponge Aplysina fistularis, and emended description of the genus Fulvivirga.

Two Gram-stain-negative, strictly aerobic, marine bacteria, designated as strains RKSG066T and RKSG123T, were isolated from a sponge Aplysina fistularis collected at a depth of 15 m off the west coast of San Salvador, The Bahamas. Investigation of nearly full-length 16S rRNA gene and whole genome-based phylogenies revealed that both strains belong to the order Cytophagales within the class Cytophagia and phylum Bacteroidetes. Strain RKSG066T formed a monophyletic clade with described members of the genus Fulvivirga, while strain RKSG123T formed a well-supported paraphyletic branch apart from this and other related genera within the family Flammeovirgaceae. For both RKSG066T and RKSG123T, optimal growth parameters were 30-37 °C, pH 7-8 and 2-3 % (w/v) NaCl; cells were catalase- and oxidase-positive, and flexirubin-type pigments were absent. The predominant fatty acids were iso-C15 : 0, C16 : 0, C18 : 0, iso-C17 : 0 3-OH, C16 : 1 ω5c, iso-C15 : 0 3-OH, C18 : 1 ω9c and iso-C15 : 1 G for RKSG066T, and iso-C17 : 0 3-OH, C16 : 1 ω5c, iso-C15 : 0, C16 : 0 3-OH and summed feature 4 (iso-C17 : 1 I and/or anteiso-C17 : 1 B) for RKSG123T. Menaquinone-7 was the major respiratory quinone for both strains. The DNA G+C contents of RKSG066T and RKSG123T were 39.5 and 36.7 mol%, respectively. On the basis of phylogenetic distinctiveness and polyphasic analysis, the type strain RKSG066T (=TSD-73T=LMG 29870T) is proposed to represent a novel species of the genus Fulvivirga, for which the name Fulvivirga aurantia sp. nov. is proposed. The type strain RKSG123T (=TSD-75T=LMG 30075T) is proposed to represent the type species of a novel genus and species with the proposed name Xanthovirga aplysinae gen. nov., sp. nov. Additionally, the genus Fulvivirga is emended to include strains of orange-pigmented colonies that contain the predominant cellular fatty acids C16 : 0, C18 : 0, C16  :  1 ω5c and C18  :  1 ω9c.

Novel Immunomodulatory Proteins Generated via Directed Evolution of Variant IgSF Domains.

Immunoglobulin superfamily member (IgSF) proteins play a significant role in regulating immune responses with surface expression on all immune cell subsets, making the IgSF an attractive family of proteins for therapeutic targeting in human diseases. We have developed a directed evolution platform capable of engineering IgSF domains to increase affinities for cognate ligands and/or introduce binding to non-cognate ligands. Using this scientific platform, ICOSL domains have been derived with enhanced binding to ICOS and with additional high-affinity binding to the non-cognate receptor, CD28. Fc-fusion proteins containing these engineered ICOSL domains significantly attenuate T cell activation in vitro and in vivo and can inhibit development of inflammatory diseases in mouse models. We also present evidence that engineered ICOSL domains can be formatted to selectively provide costimulatory signals to augment T cell responses. Our scientific platform thus provides a system for developing therapeutic protein candidates with selective biological impact for treatments of a wide array of human disorders including cancer and autoimmune/inflammatory diseases.

Clinical Genetic Testing for Familial Hypercholesterolemia: JACC Scientific Expert Panel.

Although awareness of familial hypercholesterolemia (FH) is increasing, this common, potentially fatal, treatable condition remains underdiagnosed. Despite FH being a genetic disorder, genetic testing is rarely used. The Familial Hypercholesterolemia Foundation convened an international expert panel to assess the utility of FH genetic testing. The rationale includes the following: 1) facilitation of definitive diagnosis; 2) pathogenic variants indicate higher cardiovascular risk, which indicates the potential need for more aggressive lipid lowering; 3) increase in initiation of and adherence to therapy; and 4) cascade testing of at-risk relatives. The Expert Consensus Panel recommends that FH genetic testing become the standard of care for patients with definite or probable FH, as well as for their at-risk relatives. Testing should include the genes encoding the low-density lipoprotein receptor (LDLR), apolipoprotein B (APOB), and proprotein convertase subtilisin/kexin 9 (PCSK9); other genes may also need to be considered for analysis based on patient phenotype. Expected outcomes include greater diagnoses, more effective cascade testing, initiation of therapies at earlier ages, and more accurate risk stratification.