Distinct Hodgkin lymphoma subtypes defined by noninvasive genomic profiling.

The scarcity of malignant Hodgkin and Reed-Sternberg cells hampers tissue-based comprehensive genomic profiling of classic Hodgkin lymphoma (cHL). By contrast, liquid biopsies show promise for molecular profiling of cHL due to relatively high circulating tumour DNA (ctDNA) levels1-4. Here we show that the plasma representation of mutations exceeds the bulk tumour representation in most cases, making cHL particularly amenable to noninvasive profiling. Leveraging single-cell transcriptional profiles of cHL tumours, we demonstrate Hodgkin and Reed-Sternberg ctDNA shedding to be shaped by DNASE1L3, whose increased tumour microenvironment-derived expression drives high ctDNA concentrations. Using this insight, we comprehensively profile 366 patients, revealing two distinct cHL genomic subtypes with characteristic clinical and prognostic correlates, as well as distinct transcriptional and immunological profiles. Furthermore, we identify a novel class of truncating IL4R mutations that are dependent on IL-13 signalling and therapeutically targetable with IL-4Rα-blocking antibodies. Finally, using PhasED-seq5, we demonstrate the clinical value of pretreatment and on-treatment ctDNA levels for longitudinally refining cHL risk prediction and for detection of radiographically occult minimal residual disease. Collectively, these results support the utility of noninvasive strategies for genotyping and dynamic monitoring of cHL, as well as capturing molecularly distinct subtypes with diagnostic, prognostic and therapeutic potential.

Plasma cells in human pancreatic ductal adenocarcinoma secrete antibodies against self-antigens.

Intratumoral B cell responses are associated with more favorable clinical outcomes in human pancreatic ductal adenocarcinoma (PDAC). However, the antigens driving these B cell responses are largely unknown. We sought to discover these antigens by using single-cell RNA sequencing (scRNA-Seq) and immunoglobulin (Ig) sequencing of tumor-infiltrating immune cells from 7 primary PDAC samples. We identified activated T and B cell responses and evidence of germinal center reactions. Ig sequencing identified plasma cell (PC) clones expressing isotype-switched and hypermutated Igs, suggesting the occurrence of T cell-dependent B cell responses. We assessed the reactivity of 41 recombinant antibodies that represented the products of 235 PCs and 12 B cells toward multiple cell lines and PDAC tissues and observed frequent staining of intracellular self-antigens. Three of these antigens were identified: the filamentous actin (F-actin), the nucleic protein RuvB like AAA ATPase 2 (RUVBL2), and the mitochondrial protein heat shock protein family D (Hsp60) member 1 (HSPD1). Antibody titers against F-actin and HSPD1 were substantially elevated in the plasma of patients with PDAC compared with healthy donors. Thus, PCs in PDAC produce autoantibodies reacting with intracellular self-antigens, which may result from promotion of preexisting, autoreactive B cell responses. These observations indicate the chronic inflammatory microenvironment of PDAC can support the adaptive immune response.

Immunotherapy and Targeted Therapy for Advanced Gastroesophageal Cancer: ASCO Guideline.

PURPOSE: To develop recommendations involving targeted therapies for patients with advanced gastroesophageal cancer. METHODS: The American Society of Clinical Oncology convened an Expert Panel to conduct a systematic review of relevant studies and develop recommendations for clinical practice. RESULTS: Eighteen randomized controlled trials met the inclusion criteria for the systematic review. RECOMMENDATIONS: For human epidermal growth factor receptor 2 (HER2)-negative patients with gastric adenocarcinoma (AC) and programmed death-ligand 1 (PD-L1) combined positive score (CPS) ≥ 5, first-line therapy with nivolumab and chemotherapy (CT) is recommended. For HER2-negative patients with esophageal or gastroesophageal junction (GEJ) AC and PD-L1 CPS ≥ 5, first-line therapy with nivolumab and CT is recommended. First-line therapy with pembrolizumab and CT is recommended for HER2-negative patients with esophageal or GEJ AC and PD-L1 CPS ≥ 10. For patients with esophageal squamous cell carcinoma and PD-L1 tumor proportion score ≥ 1%, nivolumab plus CT, or nivolumab plus ipilimumab is recommended; for patients with esophageal squamous cell carcinoma and PD-L1 CPS ≥ 10, pembrolizumab plus CT is recommended. For patients with HER2-positive gastric or GEJ previously untreated, unresectable or metastatic AC, trastuzumab plus pembrolizumab is recommended, in combination with CT. For patients with advanced gastroesophageal or GEJ AC whose disease has progressed after first-line therapy, ramucirumab plus paclitaxel is recommended. For HER2-positive patients with gastric or GEJ AC who have progressed after first-line therapy, trastuzumab deruxtecan is recommended. In all cases, participation in a clinical trial is recommended as it is the panel's expectation that targeted treatment options for gastroesophageal cancer will continue to evolve.Additional information is available at www.asco.org/gastrointestinal-cancer-guidelines.

Attention-deficit/hyperactivity disorder and ischemic stroke: A Mendelian randomization study.

BACKGROUND: Observational studies have found an association between attention-deficit/hyperactivity disorder (ADHD) and ischemic stroke. AIMS: The purpose of this study was to investigate whether genetic liability to ADHD has a causal effect on ischemic stroke and its subtypes. METHODS: In this two-sample Mendelian randomization (MR) study, genetic variants (nine single-nucleotide polymorphisms; P < 5 × 10-8) using as instrumental variables for the analysis was obtained from a genome-wide association study of ADHD with 19,099 cases and 34,194 controls. The outcome datasets for stroke and its subtypes were obtained from the MEGASTROKE consortium, with 40,585 cases and 406,111 controls. MR inverse variance-weighted method was conducted to investigate the effect of genetic liability to ADHD on ischemic stroke and its subtypes. Sensitivity analyses (median-based methods, MR-Egger, MR-robust adjusted profile scores, MR-pleiotropy residual sum and outlier) were also utilized to assess horizontal pleiotropy and remove outliers. Multivariable MR (MVMR) analyses were conducted to explore potential mediators. RESULTS: Genetically determined ADHD (per 1 SD) was significantly associated with a higher risk of any ischemic stroke (AIS) (odds ratio (OR) = 1.15, 95% confidence interval (CI) = 1.05-1.25, P = 0.002) and large-artery atherosclerotic stroke (LAS) (OR = 1.40, 95% CI = 1.10-1.76, P = 0.005). The significant association was also found in sensitivity analyses and MVMR analyses. CONCLUSIONS: Genetic liability to ADHD was significantly associated with an increased risk of AIS, especially LAS. The association between ADHD and LAS was independent of age of smoking initiation but mediated by coronary artery disease.

Immunotherapy in the Management of Esophagogastric Cancer: A Practical Review.

Recent data support incorporation of immune checkpoint inhibitors into the treatment armamentarium for esophageal, gastroesophageal junction, and gastric (esophagogastric) cancer. This practical review focuses on clinical trials that influenced US Food and Drug Administration approvals and treatment guidelines in esophagogastric cancer, including the impact of location, stage, histology, human epidermal growth factor receptor 2 status, and PD-(L)1 expression on these guidelines. The role of immunotherapy in the locally advanced and metastatic setting is constantly expanding. Over the next few years, the many ongoing trials exploring immunotherapy are anticipated to bring new treatment regimens into the frontline setting with the potential to improve survival in patients with advanced disease.

Multiomic analysis for optimization of combined focal and immunotherapy protocols in murine pancreatic cancer.

Background: Although combination immunotherapies incorporating local and systemic components have shown promising results in treating solid tumors, varied tumor microenvironments (TMEs) can impact immunotherapeutic efficacy. Method: We designed and evaluated treatment strategies for breast and pancreatic cancer combining magnetic resonance-guided focused ultrasound (MRgFUS) ablation and antibody therapies. With a combination of single-cell sequencing, spectral flow cytometry, and histological analyses, we profiled an immune-suppressed KPC (Kras+/LSL-G12D; Trp53+/LSL-R172H; Pdx1-Cre) pancreatic adenocarcinoma (MT4) model and a dense epithelial neu deletion (NDL) HER2+ mammary adenocarcinoma model with a greater fraction of lymphocytes, natural killer cells and activated dendritic cells. We then performed gene ontology analysis, spectral and digital cytometry to assess the immune response to combination immunotherapies and correlation with survival studies. Result: Based on gene ontology analysis, adding ablation to immunotherapy enriched immune cell migration pathways in the pancreatic cancer model and extensively enriched wound healing pathways in the breast cancer model. With CIBERSORTx digital cytometry, aCD40 + aPD-1 immunotherapy combinations enhanced dendritic cell activation in both models. In the MT4 TME, adding the combination of aCD40 antibody and checkpoint inhibitors (aPD-1 and aCTLA-4) with ablation was synergistic, increasing activated natural killer cells and T cells in distant tumors. Furthermore, ablation with immunotherapy upregulated critical Ly6c myeloid remodeling phenotypes that enhance T-cell effector function and increased granzyme and protease encoding genes by as much as 100-fold. Ablation combined with immunotherapy then extended survival in the MT4 model to a greater extent than immunotherapy alone. Conclusion: In summary, TME profiling informed a successful multicomponent treatment protocol incorporating ablation and facilitated differentiation of TMEs in which ablation is most effective.

Pancreatic Cancer Patient-derived Organoids Can Predict Response to Neoadjuvant Chemotherapy.

OBJECTIVE: To evaluate if patient-derived organoids (PDOs) may predict response to neoadjuvant (NAT) chemotherapy in patients with pancreatic adenocarcinoma. BACKGROUND: PDOs have been explored as a biomarker of therapy response and for personalized therapeutics in patients with pancreatic cancer. METHODS: During 2017-2021, patients were enrolled into an IRB-approved protocol and PDO cultures were established. PDOs of interest were analyzed through a translational pipeline incorporating molecular profiling and drug sensitivity testing. RESULTS: One hundred thirty-six samples, including both surgical resections and fine needle aspiration/biopsy from 117 patients with pancreatic cancer were collected. This biobank included diversity in stage, sex, age, and race, with minority populations representing 1/3 of collected cases (16% Black, 9% Asian, 7% Hispanic/Latino). Among surgical specimens, PDO generation was successful in 71% (15 of 21) of patients who had received NAT prior to sample collection and in 76% (39 of 51) of patients who were untreated with chemotherapy or radiation at the time of collection. Pathological response to NAT correlated with PDO chemotherapy response, particularly oxaliplatin. We demonstrated the feasibility of a rapid PDO drug screen and generated data within 7 days of tissue resection. CONCLUSION: Herein we report a large single-institution organoid biobank, including ethnic minority samples. The ability to establish PDOs from chemotherapy-naive and post-NAT tissue enables longitudinal PDO generation to assess dynamic chemotherapy sensitivity profiling. PDOs can be rapidly screened and further development of rapid screening may aid in the initial stratification of patients to the most active NAT regimen.

Structure of the MRAS-SHOC2-PP1C phosphatase complex.

RAS-MAPK signalling is fundamental for cell proliferation and is altered in most human cancers1-3. However, our mechanistic understanding of how RAS signals through RAF is still incomplete. Although studies revealed snapshots for autoinhibited and active RAF-MEK1-14-3-3 complexes4, the intermediate steps that lead to RAF activation remain unclear. The MRAS-SHOC2-PP1C holophosphatase dephosphorylates RAF at serine 259, resulting in the partial displacement of 14-3-3 and RAF-RAS association3,5,6. MRAS, SHOC2 and PP1C are mutated in rasopathies-developmental syndromes caused by aberrant MAPK pathway activation6-14-and SHOC2 itself has emerged as potential target in receptor tyrosine kinase (RTK)-RAS-driven tumours15-18. Despite its importance, structural understanding of the SHOC2 holophosphatase is lacking. Here we determine, using X-ray crystallography, the structure of the MRAS-SHOC2-PP1C complex. SHOC2 bridges PP1C and MRAS through its concave surface and enables reciprocal interactions between all three subunits. Biophysical characterization indicates a cooperative assembly driven by the MRAS GTP-bound active state, an observation that is extendible to other RAS isoforms. Our findings support the concept of a RAS-driven and multi-molecular model for RAF activation in which individual RAS-GTP molecules recruit RAF-14-3-3 and SHOC2-PP1C to produce downstream pathway activation. Importantly, we find that rasopathy and cancer mutations reside at protein-protein interfaces within the holophosphatase, resulting in enhanced affinities and function. Collectively, our findings shed light on a fundamental mechanism of RAS biology and on mechanisms of clinically observed enhanced RAS-MAPK signalling, therefore providing the structural basis for therapeutic interventions.

Inferring gene expression from cell-free DNA fragmentation profiles.

Profiling of circulating tumor DNA (ctDNA) in the bloodstream shows promise for noninvasive cancer detection. Chromatin fragmentation features have previously been explored to infer gene expression profiles from cell-free DNA (cfDNA), but current fragmentomic methods require high concentrations of tumor-derived DNA and provide limited resolution. Here we describe promoter fragmentation entropy as an epigenomic cfDNA feature that predicts RNA expression levels at individual genes. We developed 'epigenetic expression inference from cell-free DNA-sequencing' (EPIC-seq), a method that uses targeted sequencing of promoters of genes of interest. Profiling 329 blood samples from 201 patients with cancer and 87 healthy adults, we demonstrate classification of subtypes of lung carcinoma and diffuse large B cell lymphoma. Applying EPIC-seq to serial blood samples from patients treated with PD-(L)1 immune-checkpoint inhibitors, we show that gene expression profiles inferred by EPIC-seq are correlated with clinical response. Our results indicate that EPIC-seq could enable noninvasive, high-throughput tissue-of-origin characterization with diagnostic, prognostic and therapeutic potential.

PAGEANT: personal access to genome and analysis of natural traits.

GWASs have identified numerous genetic variants associated with a wide variety of diseases, yet despite the wide availability of genetic testing the insights that would enhance the interpretability of these results are not widely available to members of the public. As a proof of concept and demonstration of technological feasibility, we developed PAGEANT (Personal Access to Genome & Analysis of Natural Traits), usable through Graphical User Interface or command line-based version, aiming to serve as a protocol and prototype that guides the overarching design of genetic reporting tools. PAGEANT is structured across five core modules, summarized by five Qs: (i) quality assurance of the genetic data; (ii) qualitative assessment of genetic characteristics; (iii) quantitative assessment of health risk susceptibility based on polygenic risk scores and population reference; (iv) query of third-party variant databases (e.g. ClinVAR and PharmGKB) and (v) quick Response code of genetic variants of interest. Literature review was conducted to compare PAGEANT with academic and industry tools. For 2504 genomes made publicly available through the 1000 Genomes Project, we derived their genomic characteristics for a suite of qualitative and quantitative traits. One exemplary trait is susceptibility to COVID-19, based on the most up-to-date scientific findings reported.

Mechanical ventilation for older medical patients in a large tertiary medical care center.

BACKGROUND: The development of technologies for the prolongation of life has resulted in an increase in the number of older ventilated patients in internal medicine and chronic care wards. Our study aimed to determine the factors influencing the outcomes of older ventilated medical patients in a large tertiary medical center. METHODS: We performed a prospective observational cohort study including all newly ventilated medical patients aged 65 years and older over a period of 18 months. Data were acquired from computerized medical records and from an interview of the medical personnel initiating mechanical ventilation. RESULTS: A total of 554 patients underwent mechanical ventilation for the first time during the study period. The average age was 79 years, and 80% resided at home. Following mechanical ventilation, 8% died in the emergency room, and the majority of patients (351; 63%) were hospitalized in internal medicine wards. In-hospital mortality was 64.1%, with 48% dying during the first week of hospitalization. Overall 6-months survival was 26%. We found that a combination of age 85 years and older, functional status prior to ventilation, and associated morbidity (diabetes with target organ injury and/or oncological solid organ disease) were the strongest negative predictors of survival after discharge from the hospital. CONCLUSION: Mechanical ventilation at older age is associated with poor survival and it is possible to identify factors predicting survival. In the midst of the COVID-19 pandemic, the findings of this study may help in the decision-making process regarding mechanical ventilation for older people.

Polygenic Scores and Parental Predictors: An Adult Height Study Based on the United Kingdom Biobank and the Framingham Heart Study.

Human height is a polygenic trait, influenced by a large number of genomic loci. In the pre-genomic era, height prediction was based largely on parental height. More recent predictions of human height have made great strides by integrating genotypic data from large biobanks with improved statistical techniques. Nevertheless, recent studies have not leveraged parental height, an added feature that we hypothesized would offer complementary predictive value. In this study, we assessed the predictive power of polygenic risk scores (PRS) combined with the traditional parental height predictors. Our study analyzed genotypic data and parental height from 1,071 trios from the United Kingdom Biobank and 444 trios from the Framingham Heart Study. We explored a series of statistical models to fully evaluate the performance of several PRS constructed together with parental information and proposed a model we call PRS++ that includes gender, parental height, and PRSs of parents and proband. Our estimate of height with an R 2 of ∼0.82 is, to our knowledge, the most accurate estimate yet achieved for predicting human adult height. Without parental information, the R 2 from the best PRS-driven model is ∼0.73. In summary, using adult height prediction as an example, we demonstrated that traditional predictors still play important roles and merit integration into the current trends of intensive PRS approaches.

Interaction between genetic predisposition, smoking, and dementia risk: a population-based cohort study.

We evaluated whether the association between cigarette smoking and dementia risk is modified by genetic predisposition including apolipoprotein E (APOE) genotype and polygenic risk (excluding the APOE region). We included 193,198 UK Biobank participants aged 60-73 years without dementia at baseline. Of non-APOE-ε4 carriers, 0.89% (95% CI 0.73-1.08%) current smokers developed dementia compared with 0.49% (95% CI 0.44-0.55%) of never smokers (adjusted HR 1.78; 95% CI 1.39-2.29). In contrast, of one APOE-ε4 allele carriers, 1.69% (95% CI 1.31-2.12%) current smokers developed dementia compared with 1.40% (95% CI 1.25-1.55%) of never smokers (adjusted HR 1.06; 95% CI 0.77-1.45); of two APOE-ε4 alleles carriers, 4.90% (95% CI 2.92-7.61%) current smokers developed dementia compared with 3.87% (95% CI 3.11-4.74%) of never smokers (adjusted HR 0.94; 95% CI 0.49-1.79). Of participants with high polygenic risk, 1.77% (95% CI 1.35-2.27%) current smokers developed dementia compared with 1.05% (95% CI 0.91-1.21%) of never smokers (adjusted HR 1.63; 95% CI 1.16-2.28). A significant interaction was found between APOE genotype and smoking status (P = 0.002) while no significant interaction was identified between polygenic risk and smoking status (P = 0.25). APOE genotype but not polygenic risk modified the effect of smoking on dementia risk.

Pancreatic INI1-deficient undifferentiated rhabdoid carcinoma achieves complete clinical response on gemcitabine and nab-paclitaxel following immediate progression on FOLFIRINOX: a case report.

INTRODUCTION: INI1-deficient undifferentiated rhabdoid carcinoma is a rare pancreatic carcinoma for which the optimal treatment is unknown. Pancreatic ductal adenocarcinoma, the most common histology of pancreas cancer, is treated with combination chemotherapy in the advanced setting, a strategy supported by strong evidence in well powered studies. In patients with excellent performance status, first-line treatment usually consists of the three-drug regimen FOLFIRINOX, with the combination of gemcitabine with nab-paclitaxel, typically less toxic than the three-drug regimen, reserved for second-line therapy. Given the lack of published reports describing treatment outcomes for patients with rare forms of pancreatic cancer, the same treatment approach used for pancreatic ductal adenocarcinoma is typically employed. OBSERVATION: This case describes a patient with metastatic pancreatic INI1-deficient undifferentiated rhabdoid carcinoma who was primarily resistant to FOLFIRINOX therapy but who then achieved an immediate, marked and sustained response to gemcitabine with nab-paclitaxel. CONCLUSION: Given the lack of data informing on optimal management of INI1-deficient pancreatic undifferentiated rhabdoid carcinoma, and the exceptional response achieved by gemcitabine with nab-paclitaxel, this case report highlights a surprising and potentially informative anecdote. Additional studies are needed to confirm responses observed in this report which when taken together may strongly influence first-line therapy choice for this rare malignancy. Given the difficult in acquiring sufficient numbers of these rare histologies in any one institution, multi-institution collaboration in studying outcomes of rare pancreatic malignancies is likely essential.

PERHAPS: Paired-End short Reads-based HAPlotyping from next-generation Sequencing data.

The identification of rare haplotypes may greatly expand our knowledge in the genetic architecture of both complex and monogenic traits. To this aim, we developed PERHAPS (Paired-End short Reads-based HAPlotyping from next-generation Sequencing data), a new and simple approach to directly call haplotypes from short-read, paired-end Next Generation Sequencing (NGS) data. To benchmark this method, we considered the APOE classic polymorphism (*1/*2/*3/*4), since it represents one of the best examples of functional polymorphism arising from the haplotype combination of two Single Nucleotide Polymorphisms (SNPs). We leveraged the big Whole Exome Sequencing (WES) and SNP-array data obtained from the multi-ethnic UK BioBank (UKBB, N=48,855). By applying PERHAPS, based on piecing together the paired-end reads according to their FASTQ-labels, we extracted the haplotype data, along with their frequencies and the individual diplotype. Concordance rates between WES directly called diplotypes and the ones generated through statistical pre-phasing and imputation of SNP-array data are extremely high (>99%), either when stratifying the sample by SNP-array genotyping batch or self-reported ethnic group. Hardy-Weinberg Equilibrium tests and the comparison of obtained haplotype frequencies with the ones available from the 1000 Genome Project further supported the reliability of PERHAPS. Notably, we were able to determine the existence of the rare APOE*1 haplotype in two unrelated African subjects from UKBB, supporting its presence at appreciable frequency (approximatively 0.5%) in the African Yoruba population. Despite acknowledging some technical shortcomings, PERHAPS represents a novel and simple approach that will partly overcome the limitations in direct haplotype calling from short read-based sequencing.