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Despite the availability of effective vaccines against COVID-19, booster vaccinations are needed to maintain vaccine-induced protection against variant strains and breakthrough infections. This study aimed to investigate the efficacy, safety, and immunogenicity of the Ad26.COV2.S vaccine (Janssen) as primary vaccination plus a booster dose.ENSEMBLE2 is a randomised, double-blind, placebo-controlled, phase 3 trial including crossover vaccination after emergency authorisation of COVID-19 vaccines. Adults aged at least 18 years without previous COVID-19 vaccination at public and private medical practices and hospitals in Belgium, Brazil, Colombia, France, Germany, the Philippines, South Africa, Spain, the UK, and the USA were randomly assigned 1:1 via a computer algorithm to receive intramuscularly administered Ad26.COV2.S as a primary dose plus a booster dose at 2 months or two placebo injections 2 months apart. The primary endpoint was vaccine efficacy against the first occurrence of molecularly confirmed moderate to severe-critical COVID-19 with onset at least 14 days after booster vaccination, which was assessed in participants who received two doses of vaccine or placebo, were negative for SARS-CoV-2 by PCR at baseline and on serology at baseline and day 71, had no major protocol deviations, and were at risk of COVID-19 (ie, had no PCR-positive result or discontinued the study before day 71). Safety was assessed in all participants; reactogenicity, in terms of solicited local and systemic adverse events, was assessed as a secondary endpoint in a safety subset (approximately 6000 randomly selected participants). The trial is registered with ClinicalTrials.gov, NCT04614948, and is ongoing.Enrolment began on Nov 16, 2020, and the primary analysis data cutoff was June 25, 2021. From 34 571 participants screened, the double-blind phase enrolled 31 300 participants, 14 492 of whom received two doses (7484 in the Ad26.COV2.S group and 7008 in the placebo group) and 11 639 of whom were eligible for inclusion in the assessment of the primary endpoint (6024 in the Ad26.COV2.S group and 5615 in the placebo group). The median (IQR) follow-up post-booster vaccination was 36·0 (15·0-62·0) days. Vaccine efficacy was 75·2% (adjusted 95% CI 54·6-87·3) against moderate to severe-critical COVID-19 (14 cases in the Ad26.COV2.S group and 52 cases in the placebo group). Most cases were due to the variants alpha (B.1.1.7) and mu (B.1.621); endpoints for the primary analysis accrued from Nov 16, 2020, to June 25, 2021, before the global dominance of delta (B.1.617.2) or omicron (B.1.1.529). The booster vaccine exhibited an acceptable safety profile. The overall frequencies of solicited local and systemic adverse events (evaluated in the safety subset, n=6067) were higher among vaccine recipients than placebo recipients after the primary and booster doses. The frequency of solicited adverse events in the Ad26.COV2.S group were similar following the primary and booster vaccinations (local adverse events, 1676 [55·6%] of 3015 vs 896 [57·5%] of 1559, respectively; systemic adverse events, 1764 [58·5%] of 3015 vs 821 [52·7%] of 1559, respectively). Solicited adverse events were transient and mostly grade 1-2 in severity.A homologous Ad26.COV2.S booster administered 2 months after primary single-dose vaccination in adults had an acceptable safety profile and was efficacious against moderate to severe-critical COVID-19. Studies assessing efficacy against newer variants and with longer follow-up are needed.Janssen ResearchDevelopment.

A patient with B-cell acute lymphoblastic leukemia (ALL) and severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) had persistent, progressive pneumonia with viremia after 5 months of infection despite monoclonal antibodies, intravenous (IV) remdesivir and prolonged oral steroids. Twenty days of nirmatrelvir/ritonavir and 10 days of IV remdesivir led to full recovery.

To determine the epidemiology of human parainfluenza virus in homeless shelters during the COVID-19 pandemic, we analyzed data and sequences from respiratory specimens collected in 23 shelters in Washington, USA, during 2019-2021. Two clusters in children were genetically similar by shelter of origin. Shelter-specific interventions are needed to reduce these infections.

SARS-CoV-2 mutations that cause resistance to monoclonal antibody (mAb) therapy have been reported. However, it remains unclear whether in vivo emergence of SARS-CoV-2 resistance mutations alters viral replication dynamics or therapeutic efficacy in the immune-competent population. As part of the ACTIV-2/A5401 randomized clinical trial (NCT04518410), non-hospitalized participants with symptomatic SARS-CoV-2 infection were given bamlanivimab (700 mg or 7,000 mg) or placebo treatment. Here¸ we report that treatment-emergent resistance mutations [detected through targeted Spike (S) gene next-generation sequencing] were significantly more likely to be detected after bamlanivimab 700 mg treatment compared with the placebo group (7% of 111 vs 0% of 112 participants, P = 0.003). No treatment-emergent resistance mutations among the 48 participants who received 7,000 mg bamlanivimab were recorded. Participants in which emerging mAb resistant virus mutations were identified showed significantly higher pretreatment nasopharyngeal and anterior nasal viral loads. Daily respiratory tract viral sampling through study day 14 showed the dynamic nature of in vivo SARS-CoV-2 infection and indicated a rapid and sustained viral rebound after the emergence of resistance mutations. Participants with emerging bamlanivimab resistance often accumulated additional polymorphisms found in current variants of concern/interest that are associated with immune escape. These results highlight the potential for rapid emergence of resistance during mAb monotherapy treatment that results in prolonged high-level respiratory tract viral loads. Assessment of viral resistance should be prioritized during the development and clinical implementation of antiviral treatments for COVID-19.

Two randomized controlled trials demonstrated no clinical benefit of hydroxychloroquine (HCQ) for either postexposure prophylaxis or early treatment of SARS-CoV-2 infection. Using data from these studies, we calculated the time-weighted average change from baseline SARS-CoV-2 viral load and demonstrated that HCQ did not affect viral clearance.

Background Rhinovirus (RV) is a common cause of respiratory illness in all people, including those experiencing homelessness. RV epidemiology in homeless shelters is unknown. Methods We analyzed data from a cross-sectional homeless shelter study in King County, Washington, October 2019–May 2021. Shelter residents or guardians aged ≥3 months reporting acute respiratory illness completed questionnaires and submitted nasal swabs. After 1 April 2020, enrollment expanded to residents and staff regardless of symptoms. Samples were tested by multiplex RT-PCR for respiratory viruses. A subset of RV-positive samples was sequenced. Results There were 1066 RV-positive samples with RV present every month of the study period. RV was the most common virus before and during the coronavirus disease 2019 (COVID-19) pandemic (43% and 77% of virus-positive samples, respectively). Participants from family shelters had the highest prevalence of RV. Among 131 sequenced samples, 33 RV serotypes were identified with each serotype detected for ≤4 months. Conclusions RV infections persisted through community mitigation measures and were most prevalent in shelters housing families. Sequencing showed a diversity of circulating RV serotypes, each detected over short periods of time. Community-based surveillance in congregate settings is important to characterize respiratory viral infections during and after the COVID-19 pandemic. Clinical Trials Registration NCT04141917.

It is of interest to pinpoint SARS-CoV-2 sequence features defining vaccine resistance. In the ENSEMBLE randomized, placebo-controlled phase 3 trial, estimated single-dose Ad26.COV2.S vaccine efficacy (VE) was 56% against moderate to severe–critical COVID-19. SARS-CoV-2 Spike sequences were measured from 484 vaccine and 1,067 placebo recipients who acquired COVID-19 during the trial. In Latin America, where Spike diversity was greatest, VE was significantly lower against Lambda than against Reference and against all non-Lambda variants [family-wise error rate (FWER) p

BackgroundSyphilis incidence continues to increase dramatically in the United States and yet little is known aboutTreponema pallidum(TP) genomic epidemiology within American metropolitan areas.MethodsWe performed whole genome sequencing andtprKdeep sequencing of 28 TP-containing specimens collected mostly from remnant Aptima swabs from 24 individuals from Seattle Sexual Health Clinic during 2021-2022.ResultsAll 12 individuals infected with Nichols lineage strains were MSM, while a specific SS14 cluster (average 0.33 SNPs) included 1 MSW and five women. All TP strains sequenced were azithromycin resistant via 23S rRNA A2058G mutation. Identical TP genomic sequences were found in pharyngeal and rectal swab specimens taken from the same individuals concurrently.tprKsequences were less variable between patient-matched specimens and between epidemiologically-linked clusters. We detected a 528 bp deletion in thetprKdonor site locus, eliminating ninetprKdonor sites, in TP genomes of three individuals with secondary syphilis, associated with diminution of overalltprKsequence diversity.ConclusionsWe developed an end-to-end workflow for public health genomic surveillance of TP from remnant Aptima swab specimens. With its high rate of gene conversion,tprKsequencing may assist in linking cases beyond routine TP genome sequencing. TP strains with deletions intprKdonor sites currently circulate and are associated with diminished antigenic diversity of the TprK putative outer membrane protein.

BackgroundHuman rhinovirus (HRV) infections can progress from the upper (URT) to lower (LRT) respiratory tract in immunocompromised individuals, causing high rates of fatal pneumonia. Little is known about how HRV evolves within hosts during infection.MethodsWe sequenced HRV complete genomes from 12 hematopoietic cell transplant patients with prolonged infection for up to 190 days from both URT (nasal wash, NW) and LRT (bronchoalveolar lavage, BAL) specimens. Metagenomic (mNGS) and amplicon-based NGS were used to study the emergence and evolution of intra-host single nucleotide variants (iSNVs).ResultsIdentical HRV intra-host populations in matched NW and BAL specimens indicated no genetic adaptation is required for HRV to progress from URT to LRT. Microbial composition between matched NW and BAL confirmed no cross-contamination during sampling procedure. Coding iSNVs were 2.3-fold more prevalent in capsid over non-structural genes, adjusted for length. iSNVs modeled onto HRV capsid structures were significantly more likely to be found in surface residues, but were not preferentially located in known HRV neutralizing antibody epitopes. Newly emergent, serotype-matched iSNV haplotypes from immunocompromised individuals from 2008-2010 could be detected in Seattle-area community HRV sequences from 2020-2021.ConclusionHRV infections in immunocompromised hosts can progress from URT to LRT with no specific evolutionary requirement. Capsid proteins carry the highest variability and emergent mutations can be detected in other, including future, HRV sequences.

Do rapid antigen test results correlate with replication-competent severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) (i.e., infectious) virus? In this cross-sectional diagnostic study of 626 adults, the sensitivity of the antigen test to identify replication-competent SARS-CoV-2 was 63.6% in asymptomatic and 91.0% in symptomatic participants. Viral culture titers were the highest at the onset of symptoms and rapidly declined by 7 days after symptom onset.

Merkel cell carcinoma (MCC) is often caused by persistent expression of Merkel cell polyomavirus (MCPyV) T-antigen (T-Ag). These non-self proteins comprise about 400 amino acids (AA). Clinical responses to immune checkpoint inhibitors, seen in about half of patients, may relate to T-Ag–specific T cells. Strategies to increase CD8+ T-cell number, breadth, or function could augment checkpoint inhibition, but vaccines to augment immunity must avoid delivery of oncogenic T-antigen domains. We probed MCC tumor-infiltrating lymphocytes (TIL) with an artificial antigen-presenting cell (aAPC) system and confirmed T-Ag recognition with synthetic peptides, HLA-peptide tetramers, and dendritic cells (DC). TILs from 9 of 12 (75%) subjects contained CD8+ T cells recognizing 1–8 MCPyV epitopes per person. Analysis of 16 MCPyV CD8+ TIL epitopes and prior TIL data indicated that 97% of patients with MCPyV+ MCC had HLA alleles with the genetic potential that restrict CD8+ T-cell responses to MCPyV T-Ag. The LT AA 70–110 region was epitope rich, whereas the oncogenic domains of T-Ag were not commonly recognized. Specific recognition of T-Ag–expressing DCs was documented. Recovery of MCPyV oncoprotein–specific CD8+ TILs from most tumors indicated that antigen indifference was unlikely to be a major cause of checkpoint inhibition failure. The myriad of epitopes restricted by diverse HLA alleles indicates that vaccination can be a rational component of immunotherapy if tumor immune suppression can be overcome, and the oncogenic regions of T-Ag can be modified without impacting immunogenicity.

Biological importance of Gab1 in HNSCC cells

Supplementary Table from EGFR Inhibition Potentiates FGFR Inhibitor Therapy and Overcomes Resistance in FGFR2 Fusion–Positive Cholangiocarcinoma

Sup Tables 1-6

Sup Figures 1-13

Preclinical and clinical studies have evidenced that effective targeted therapy treatment designed against receptor tyrosine kinases (RTKs) in different solid tumor paradigms is predicated on simultaneous inhibition of both the PI3K and MEK intracellular signaling pathways. Indeed, reactivation of either pathway results in resistance to these therapies. Recently, oncogenic phosphatase SHP2 inhibitors have been developed with some now reaching clinical trials. To expand on possible indications for SHP099, we screened over 800 cancer cell lines covering over 25 subsets of cancer. We found head and neck squamous cell carcinoma (HNSCC) was the most sensitive adult subtype of cancer to SHP099. We found that, in addition to the MEK pathway, SHP2 inhibition blocks the PI3K pathway in sensitive HNSCCs, resulting in downregulation of mTORC signaling and antitumor effects across several HNSCC mouse models, including an human papillomavirus (HPV+) patient-derived xenograft. Importantly, we found low levels of the RTK ligand epiregulin identified HNSCCs that were sensitive to SHP2 inhibitor, and, adding exogenous epiregulin mitigated SHP099 efficacy. Mechanistically, epiregulin maintained SHP2–GAB1 complexes in the presence of SHP2 inhibition, preventing downregulation of the MEK and PI3K pathways. In the presence of SHP2 inhibitor, HNSCCs are highly dependent on GAB1 for their survival and knockdown of GAB1 is sufficient to block the ability of epiregulin to rescue MEK and PI3K signaling. These data connect the sensitivity of HNSCC to SHP2 inhibitors and to a broad reliance on GAB1-SHP2, revealing an important and druggable signaling axis. Overall, SHP2 inhibitors are being heavily developed and may have activity in HNSCCs, and in particular those with low levels of epiregulin.Significance:This work identifies a novel role of SHP2 inhibitor by dual downregulation of PI3K and MEK pathways, through loss of GAB1 activation and disruption of GAB1 complexes in low-epiregulin HNSCC.

Effects of knockdown of Gab1 in SHP099-resistant HNSCC cell line

RTK ligand expression in HNSCC cell lines

Neuregulin-1 does not rescue PI3K and MEK signaling in HNSCC treated with SHP2 inhibitor

Cell line information and SHP099 screen data

Three- and five-day western blot analysis of SHP099-sensitive and - resistant HNSCC cell lines

HNSCC tumor cell lines are effective to SHP099 through dual PI3K and MEK inhibition

Cell viability of SHP099-sensitive HNSCC cell lines

The mechanism of action of SHP099 inhibition

Supplementary Figure from EGFR Inhibition Potentiates FGFR Inhibitor Therapy and Overcomes Resistance in FGFR2 Fusion–Positive Cholangiocarcinoma

Western blot analysis of ErbB members

FGFR inhibitors are approved for the treatment of advanced cholangiocarcinoma harboring FGFR2 fusions. However, the response rate is moderate, and resistance emerges rapidly due to acquired secondary FGFR2 mutations or due to other less-defined mechanisms. Here, we conducted high-throughput combination drug screens, biochemical analysis, and therapeutic studies using patient-derived models of FGFR2 fusion–positive cholangiocarcinoma to gain insight into these clinical profiles and uncover improved treatment strategies. We found that feedback activation of EGFR signaling limits FGFR inhibitor efficacy, restricting cell death induction in sensitive models and causing resistance in insensitive models lacking secondary FGFR2 mutations. Inhibition of wild-type EGFR potentiated responses to FGFR inhibitors in both contexts, durably suppressing MEK/ERK and mTOR signaling, increasing apoptosis, and causing marked tumor regressions in vivo. Our findings reveal EGFR-dependent adaptive signaling as an important mechanism limiting FGFR inhibitor efficacy and driving resistance and support clinical testing of FGFR/EGFR inhibitor therapy for FGFR2 fusion–positive cholangiocarcinoma.Significance:We demonstrate that feedback activation of EGFR signaling limits the effectiveness of FGFR inhibitor therapy and drives adaptive resistance in patient-derived models of FGFR2 fusion–positive cholangiocarcinoma. These studies support the potential of combination treatment with FGFR and EGFR inhibitors as an improved treatment for patients with FGFR2-driven cholangiocarcinoma.This article is highlighted in the In This Issue feature, p. 1171

Supplementary Figure Legends PDF file 97K, Complete legends for all 5 Supplementary Figures

Source and cell culture information for cell lines.

Supplementary Figure S1. IDHm ICC are insensitive to IDH inhibition. Supplementary Figure S2. Similarity matrix for drug response of BTC cell lines. Supplementary Figure S3. Hypersensitivity of ICC cells harboring endogenous IDH mutations to dasatinib. Supplementary Figure S4. Torin 1 inhibits growth of ICC cell lines. Supplementary Figure S5. Critical role for SRC in IDHm ICC cells. Supplementary Figure S6. Ectopic expression of mutant IDH has only a modest effect on dasatinib sensitivity.

Supplemental Figure 3. Chemotherapy and venetoclax combination cell viability assays. Cell viability assays were performed for 72h treatment of cyclophosphamide and topotecan, venetoclax, or combination at indicated concentrations, with bliss synergy scores in (A) MYCN-amplified, and (B) MYCN-wild-type cell lines. (C) Plot of Bliss sum of cell lines in A-B, calculated by the summation of the bliss synergy scores across the doses presented.

Table S1: Crystallographic data collection and refinement statistics Table S2: Inhibitory activity against other BET bromodomains Table S3: Bromodomain profiling data Fig. S1: Detailed binding interactions of compounds 1 - 5 in BRD4-1 Fig. S2: Comparison of compound 2 and JQ1 in Brd4-1 Fig. S3: Chemical structures of other compounds used in this work. Fig. S4: Statistical evaluation of kinome profiling data using the Gini coefficient Fig. S5: Original flow cytometry data of MM1.S cells Fig. S6: MM1.S cells require stimulation with IL-6 to detect phospho-STAT3 levels Fig. S7: Effect of dual BRD4-kinase inhibitors on MV-4-11 cells. Fig. S8: Time dependent alteration of c-Myc and p21Cip1 levels in MM1.S cells. Fig. S9: Effect of dual BET-kinase inhibitors on ruxolitinib sensitive MPN cell lines Fig. S10: Dose-effect analysis of UKE-1 cells treated with drug combinations Fig. S11: Cell line sensitivity by tissue type

Supplemental Figure 2. PDX ex vivo model demonstrated cleavage of PARP with corresponding NOXA upregulation (A) Western blot of COG-N-561 ex vivo cell line treated overnight with antibodies as indicated.

Supplementary Table 2. Lung cell lines homozygotes for KDM4A SNP-A482 are not enriched for mutations that could be linked to sensitivity to PI3K/mTOR inhibitors.

Median centered Ln(IC50) values for 17 BTC cell lines screened against 122 drugs.

Synergistic action of kinase and BET bromodomain inhibitors in cell killing has been reported for a variety of cancers. Using the chemical scaffold of the JAK2 inhibitor TG101348, we developed and characterized single agents which potently and simultaneously inhibit BRD4 and a specific set of oncogenic tyrosine kinases including JAK2, FLT3, RET, and ROS1. Lead compounds showed on-target inhibition in several blood cancer cell lines and were highly efficacious at inhibiting the growth of hematopoietic progenitor cells from patients with myeloproliferative neoplasm. Screening across 931 cancer cell lines revealed differential growth inhibitory potential with highest activity against bone and blood cancers and greatly enhanced activity over the single BET inhibitor JQ1. Gene drug sensitivity analyses and drug combination studies indicate synergism of BRD4 and kinase inhibition as a plausible reason for the superior potency in cell killing. Combined, our findings indicate promising potential of these agents as novel chemical probes and cancer therapeutics. Mol Cancer Ther; 16(6); 1054–67. ©2017 AACR.

PDF file - 93K, This file contains 9 supplementary tables including the full primary screening data and secondary testing of BETi in neuroblastoma, supporting tables for the expression profiling analyses, and the list of Affymetrix probes (MYCN) and shRNA sequences for BRD4 and MYCN.

Supplementary Figure 3. Rapamycin treatment reduces KDM4A protein levels.

Supplementary Figure Legends

Supplementary Figure 2 PDF file 74K, Mcl-1 is essential for cell survival in SCC cells and vorinostat induces redistribution of Bim from Mcl-1 to Bcl-2/Bcl-xl

Supplementary Table 3. List of drugs and primary targets used for the study.

Supplementary Figure 1 PDF file 140K, Vorinostat induces apoptosis and alters gene expression of Bcl-2 family members in different SCC cell lines

Supplementary Figure 1. KDM4A SNP-A482 (rs586339) correlates with worse outcome in NSCLC patients.