The rockefeller university

Fibrolamellar carcinoma (FLC) is a rare, often lethal, liver cancer affecting adolescents and young adults, for which there are no approved therapeutics. The development of therapeutics is hampered by a lack of in vitro models. Organoids have shown utility as a model system for studying many diseases. In this study, tumor tissue and the adjacent non-tumor liver were obtained at the time of surgery. The tissue was dissociated and grown as organoids. We developed 21 patient-derived organoid lines: 12 from metastases, three from the liver tumor and six from adjacent non-tumor liver. These patient-derived FLC organoids recapitulate the histologic morphology, immunohistochem-istry, and transcriptome of the patient tumor. Patient-derived FLC organoids were used in a preliminary high-throughput drug screen to show proof of concept for the identification of therapeutics. This model system has the potential to improve our understanding of this rare cancer and holds significant promise for drug testing and development.

Background Autosomal recessive (AR) PKC delta deficiency is a rare inborn error of immunity (IEI) characterized by autoimmunity and susceptibility to bacterial, fungal, and viral infections. PKC delta is involved in the intracellular production of reactive oxidative species (ROS). Material and Methods We studied a 5-year old girl presenting with a history of Burkholderia cepacia infection. She had no history of autoimmunity, lymphocyte counts were normal, and no auto-antibodies were detected in her plasma. We performed a targeted panel analysis of 407 immunity-related genes and immunological investigations of the underlying genetic condition in this patient. Results Consistent with a history suggestive of chronic granulomatous disease (CGD), oxidative burst impairment was observed in the patient's circulating phagocytes in a dihydrorhodamine 123 (DHR) assay. However, targeted genetic panel analysis identified no candidate variants of known CGD-causing genes. Two heterozygous candidate variants were detected in PRKCD: c.285C > A (p.C95*) and c.376G > T (p.D126Y). The missense variant was also predicted to cause abnormal splicing, as it is located at the splice donor site of exon 5. TOPO-TA cloning confirmed that exon 5 was completely skipped, resulting in a truncated protein. No PKC delta protein was detected in the patient's neutrophils and monocyte-derived macrophages. The monocyte-derived macrophages of the patient produced abnormally low levels of ROS, as shown in an Amplex Red assay. Conclusion PKC delta deficiency should be considered in young patients with CGD-like clinical manifestations and abnormal DHR assay results, even in the absence of clinical and biological manifestations of autoimmunity.

Protein kinases play a vital role in biology and deregulation of kinases is implicated in numerous diseases ranging from cancer to neurodegenerative diseases, making them a major target class for the pharmaceutical industry. However, the high degree of conservation that exists between ATP-binding sites among kinases makes it difficult for current inhibitors to be highly specific. In the context of neurodegeneration, several groups including ours, have linked different kinases such as CK1 and Alzheimer's disease for example. Strictly CK1-isoform specific regulators do not exist and known CK1 inhibitors are inhibiting the enzymatic activity, targeting the ATP-binding site. Here we review compounds known to target CK1, as well as other inhibitory types that could benefit CK1. We introduce the DNA-encoded library (DEL) technology that might represent an interesting approach to uncover allosteric modulators instead of ATP competitors. Such a strategy, taking into account known allosteric inhibitors and mechanisms, might help designing modulators that are more specific towards a specific kinase, and in the case of CK1, toward specific isoforms.

Global population immunity to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is accumulating through heterogeneous combinations of infection and vaccination. Vaccine distribution in low- and middle-income countries has been variable and reliant on diverse vaccine platforms. We studied B-cell immunity in Mexico, a middle-income country where five different vaccines have been deployed to populations with high SARS-CoV-2 incidences. Levels of antibodies that bound a stabilized prefusion spike trimer, neutralizing antibody titers, and memory B-cell expansion correlated with each other across vaccine platforms. Nevertheless, the vaccines elicited variable levels of B-cell immunity, and the majority of recipients had undetectable neutralizing activity against the recently emergent omicron variant. SARS-CoV-2 infection, experienced before or after vaccination, potentiated B-cell immune responses and enabled the generation of neutralizing activity against omicron and SARS-CoV for all vaccines in nearly all individuals. These findings suggest that broad population immunity to SARS-CoV-2 will eventually be achieved but by heterogeneous paths. IMPORTANCE The majority of studies on SARS-CoV-2 vaccine-elicited immunity and immune evasion have focused on single vaccines corresponding to those distributed in high-income countries. However, in low- and middle-income countries, vaccine deployment has been far less uniform. It is therefore important to determine the levels of immunity elicited by vaccines that have been deployed globally. Such data should help inform policy. Thus, this paper is very much a "real-world" study that focuses on a middle-income country, Mexico, in which five different vaccines based on mRNA, adenovirus, and inactivated-virus platforms have been extensively deployed, while (as documented in our study) SARS-CoV-2 variants with increasing degrees of immune evasiveness have propagated in the Mexican population, culminating in the recent emergence of B.1.1.529 (omicron). The majority of studies on SARS-CoV-2 vaccine-elicited immunity and immune evasion have focused on single vaccines corresponding to those distributed in high-income countries. However, in low- and middle-income countries, vaccine deployment has been far less uniform.

The induction of broadly neutralizing antibodies (bNAbs) is a potential strategy for a vaccine against HIV-1. However, most bNAbs exhibit features such as unusually high somatic hypermutation, including insertions and deletions, which make their induction challenging. VRC01-class bNAbs not only exhibit extraordinary breadth and potency but also rank among the most highly somatically mutated bNAbs. Here, we describe a VRC01-class antibody isolated from a viremic controller, BG24, that is much less mutated than most relatives of its class while achieving comparable breadth and potency. A 3.8-angstrom x-ray crystal structure of a BG24-BG505 Env trimer complex revealed conserved contacts at the gp120 interface characteristic of the VRC01-class Abs, despite lacking common CDR3 sequence motifs. The existence of moderately mutated CD4-binding site (CD4bs) bNAbs such as BG24 provides a simpler blueprint for CD4bs antibody induction by a vaccine, raising the prospect that such an induction might be feasible with a germline-targeting approach.

Anti-programmed death-1 (anti-PD-1) immunotherapy reinvigorates CD8 T cell responses in patients with cancer but PD-1 is also expressed by other immune cells, including follicular helper CD4 T cells (Tfh) which are involved in germinal centre responses. Little is known, however, about the effects of anti-PD-1 immunotherapy on noncancer immune responses in humans. To investigate this question, we examined the impact of anti-PD-1 immunotherapy on the Tfh-B cell axis responding to unrelated viral antigens. Following influenza vaccination, a subset of adults receiving anti-PD-1 had more robust circulating Tfh responses than adults not receiving immunotherapy. PD-1 pathway blockade resulted in transcriptional signatures of increased cellular proliferation in circulating Tfh and responding B cells compared with controls. These latter observations suggest an underlying change in the Tfh-B cell and germinal centre axis in a subset of immunotherapy patients. Together, these results demonstrate dynamic effects of anti-PD-1 therapy on influenza vaccine responses and highlight analytical vaccination as an approach that may reveal underlying immune predisposition to adverse events. Patients with cancer undergoing anti-PD-1 immune checkpoint blockade can experience immune-related adverse effects. Wherry and colleagues examined the immunity elicited upon immunization of patients with cancer and report that anti-PD-1 immunotherapy dynamically affects influenza vaccine-induced immune responses.

Wang et al. compare antibody immune responses following different COVID-19 vaccination regimens. Their detailed immunological and structural analysis revealed significant differences that can inform improved vaccination strategies for the prevention of COVID-19 and other respiratory viral infections of pandemic potential. The SARS-CoV-2 pandemic prompted a global vaccination effort and the development of numerous COVID-19 vaccines at an unprecedented scale and pace. As a result, current COVID-19 vaccination regimens comprise diverse vaccine modalities, immunogen combinations, and dosing intervals. Here, we compare vaccine-specific antibody and memory B cell responses following two-dose mRNA, single-dose Ad26.COV.2S, and two-dose ChAdOx1, or combination ChAdOx1/mRNA vaccination. Plasma-neutralizing activity, as well as the magnitude, clonal composition, and antibody maturation of the RBD-specific memory B cell compartments, showed substantial differences between the vaccination regimens. While individual monoclonal antibodies derived from memory B cells exhibited similar binding affinities and neutralizing potency against Wuhan-Hu-1 SARS-CoV-2, there were significant differences in epitope specificity and neutralizing breadth against viral variants of concern. Although the ChAdOx1 vaccine was inferior to mRNA and Ad26.COV.2S in several respects, biochemical and structural analyses revealed enrichment in a subgroup of memory B cell neutralizing antibodies with distinct RBD-binding properties resulting in remarkable potency and breadth.

Proteogenomic identification of translated small open reading frames has revealed thousands of previously unannotated, largely uncharacterized microproteins, or polypeptides of less than 100 amino acids, and alternative proteins (alt-proteins) that are co-encoded with canonical proteins and are often larger. The subcellular localizations of microproteins and alt-proteins are generally unknown but can have significant implications for their functions. Proximity biotinylation is an attractive approach to define the protein composition of subcellular compartments in cells and in animals. Here, we developed a high-throughput technology to map unannotated microproteins and alt-proteins to subcellular localizations by proximity biotinylation with TurboID (MicroID). More than 150 microproteins and alt-proteins are associated with subnuclear organelles. One alt-protein, alt-LAMA3, localizes to the nucleolus and functions in pre-rRNA transcription. We applied MicroID in a mouse model, validating expression of a conserved nuclear microprotein, and establishing MicroID for discovery of microproteins and alt-proteins in vivo.