APOBEC shapes tumor evolution and age at onset of lung cancer in smokers.

APOBEC enzymes are part of the innate immunity and are responsible for restricting viruses and retroelements by deaminating cytosine residues1,2. Most solid tumors harbor different levels of somatic mutations attributed to the off-target activities of APOBEC3A (A3A) and/or APOBEC3B (A3B)3-6. However, how APOBEC3A/B enzymes shape the tumor evolution in the presence of exogenous mutagenic processes is largely unknown. Here, by combining deep whole-genome sequencing with multi-omics profiling of 309 lung cancers from smokers with detailed tobacco smoking information, we identify two subtypes defined by low (LAS) and high (HAS) APOBEC mutagenesis. LAS are enriched for A3B-like mutagenesis and KRAS mutations, whereas HAS for A3A-like mutagenesis and TP53 mutations. Unlike APOBEC3A, APOBEC3B expression is strongly associated with an upregulation of the base excision repair pathway. Hypermutation by unrepaired A3A and tobacco smoking mutagenesis combined with TP53-induced genomic instability can trigger senescence7, apoptosis8, and cell regeneration9, as indicated by high expression of pulmonary healing signaling pathway, stemness markers and distal cell-of-origin in HAS. The expected association of tobacco smoking variables (e.g., time to first cigarette) with genomic/epigenomic changes are not observed in HAS, a plausible consequence of frequent cell senescence or apoptosis. HAS have more neoantigens, slower clonal expansion, and older age at onset compared to LAS, particularly in heavy smokers, consistent with high proportions of newly generated, unmutated cells and frequent immuno-editing. These findings show how heterogeneity in mutational burden across co-occurring mutational processes and cell types contributes to tumor development, with important clinical implications.

Evidence That Hypothalamic Gliosis Is Related to Impaired Glucose Homeostasis in Adults With Obesity.

OBJECTIVE: Preclinical research implicates hypothalamic glial cell responses in the pathogenesis of obesity and type 2 diabetes (T2D). In the current study we sought to translate such findings to humans by testing whether radiologic markers of gliosis in the mediobasal hypothalamus (MBH) were greater in individuals with obesity and impaired glucose homeostasis or T2D. RESEARCH DESIGN AND METHODS: Using cross-sectional and prospective cohort study designs, we applied a validated quantitative MRI approach to assess gliosis in 67 adults with obesity and normal glucose tolerance, impaired glucose tolerance (IGT), or T2D. Assessments of glucose homeostasis were conducted via oral glucose tolerance tests (OGTT) and ß-cell modeling. RESULTS: We found significantly greater T2 relaxation times (a marker of gliosis by MRI), that were independent of adiposity, in the groups with IGT and T2D as compared with the group with normal glucose tolerance. Findings were present in the MBH, but not control regions. Moreover, positive linear associations were present in the MBH but not control regions between T2 relaxation time and glucose area under the curve during an OGTT, fasting glucose concentrations, hemoglobin A1c, and visceral adipose tissue mass, whereas negative linear relationships were present in the MBH for markers of insulin sensitivity and ß-cell function. In a prospective cohort study, greater MBH T2 relaxation times predicted declining insulin sensitivity over 1 year. CONCLUSIONS: Findings support a role for hypothalamic gliosis in the progression of insulin resistance in obesity and thus T2D pathogenesis in humans.

Genomic and evolutionary classification of lung cancer in never smokers.

Lung cancer in never smokers (LCINS) is a common cause of cancer mortality but its genomic landscape is poorly characterized. Here high-coverage whole-genome sequencing of 232 LCINS showed 3 subtypes defined by copy number aberrations. The dominant subtype (piano), which is rare in lung cancer in smokers, features somatic UBA1 mutations, germline AR variants and stem cell-like properties, including low mutational burden, high intratumor heterogeneity, long telomeres, frequent KRAS mutations and slow growth, as suggested by the occurrence of cancer drivers' progenitor cells many years before tumor diagnosis. The other subtypes are characterized by specific amplifications and EGFR mutations (mezzo-forte) and whole-genome doubling (forte). No strong tobacco smoking signatures were detected, even in cases with exposure to secondhand tobacco smoke. Genes within the receptor tyrosine kinase-Ras pathway had distinct impacts on survival; five genomic alterations independently doubled mortality. These findings create avenues for personalized treatment in LCINS.

Reassessing relationships between appetite and adiposity in people at risk of obesity: A twin study using fMRI.

BACKGROUND: Neuroimaging studies suggest that appetitive drive is enhanced in obesity. OBJECTIVE: To test if appetitive drive varies in direct proportion to the level of body adiposity after accounting for genetic factors that contribute to both brain response and obesity risk. SUBJECTS/METHODS: Participants were adult monozygotic (n = 54) and dizygotic (n = 30) twins with at least one member of the pair with obesity. Body composition was assessed by dual-energy X-ray absorptiometry. Hormonal and appetite measures were obtained in response to a standardized meal that provided 20% of estimated daily caloric needs and to an ad libitum buffet meal. Pre- and post-meal functional magnetic resonance imaging (fMRI) assessed brain response to visual food cues in a set of a priori appetite-regulating regions. Exploratory voxelwise analyses outside a priori regions were performed with correction for multiple comparisons. RESULTS: In a group of 84 adults, the majority with obesity (75%), body fat mass was not associated with hormonal responses to a meal (glucose, insulin, glucagon-like peptide-1 and ghrelin, all P>0.40), subjective feelings of hunger (ß=-0.01 mm [95% CI -0.35, 0.34] P = 0.97) and fullness (ß=0.15 mm [-0.15, 0.44] P = 0.33), or buffet meal intake in relation to estimated daily caloric needs (ß=0.28% [-0.05, 0.60] P = 0.10). Body fat mass was also not associated with brain response to high-calorie food cues in appetite-regulating regions (Pre-meal ß=-0.12 [-0.32, 0.09] P = 0.26; Post-meal ß=0.18 [-0.02, 0.37] P = 0.09; Change by a meal ß=0.29 [-0.02, 0.61] P = 0.07). Conversely, lower fat mass was associated with being weight reduced (ß=-0.05% [-0.07, -0.03] P<0.001) and greater pre-meal activation to high-calorie food cues in the dorsolateral prefrontal cortex (Z = 3.63 P = 0.017). CONCLUSIONS: In a large study of adult twins, the majority with overweight or obesity, the level of adiposity was not associated with excess appetitive drive as assessed by behavioral, hormonal, or fMRI measures.

Upgrades to intensive care: The effects of COVID-19 on decision-making in the emergency department.

INTRODUCTION: The initial surge of critically ill patients in the COVID-19 pandemic severely disrupted processes at acute care hospitals. This study examines the frequency and causes for patients upgraded to intensive care unit (ICU) level care following admission from the emergency department (ED) to non-critical care units. METHODS: The number of ICU upgrades per month was determined, including the percentage of upgrades noted to have non-concordant diagnoses. Charts with non-concordant diagnoses were examined in detail as to the ED medical decision-making, clinical circumstances surrounding the upgrade, and presence of a diagnosis of COVID-19. For each case, a cognitive bias was assigned. RESULTS: The percentage of upgraded cases with non-concordant diagnoses increased from a baseline range of 14-20% to 41.3%. The majority of upgrades were due to premature closure (72.2%), anchoring (61.1%), and confirmation bias (55.6%). CONCLUSION: Consistent with the behavioral literature, this suggests that stressful ambient conditions affect cognitive reasoning processes.

Rate of Decompensation of Normoxic Emergency Department Patients with SARS-CoV-2.

INTRODUCTION: As of October 30, 2020, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has infected over 44 million people worldwide and killed over 1.1 million people. In the emergency department (ED), patients who need supplemental oxygen or respiratory support are admitted to the hospital, but the course of normoxic patients with SARS-CoV-2 infection is unknown. In our health system, the policy during the coronavirus 2019 (COVID-19) pandemic was to admit all patients with abnormal chest imaging (CXR) regardless of their oxygen level. We also admitted febrile patients with respiratory complaints who resided in congregate living. We describe the rate of decompensation among patients admitted with suspected SARS-CoV-2 infection but who were not hypoxemic in the ED. METHODS: This is a retrospective observational study of patients admitted to our health system between March 1-May 5, 2020 with suspected SARS-CoV-2 infection. We queried our registry to find patients who were admitted to the hospital but had no recorded oxygen saturation of <92% in the ED and received no supplemental oxygen prior to admission. Our primary outcome was decompensation at 72 hours, defined by the need for respiratory support (oxygen, high-flow nasal cannula, non-invasive ventilation, or intubation). RESULTS: A total of 840 patients met our inclusion criteria. Of those patients, 376 (45%) tested positive for SARS-CoV-2. Sixty patients (7.1%) with suspected COVID-19 required respiratory support at 72 hours including 27 (3%) of confirmed SARS-CoV-2 positive patients. Among the 376 patients who tested positive for SARS-CoV-2, 54 patients (14%) had normal CXR in the ED. One-third of patients with normal CXRs decompensated at 72 hours. Seven SARS-CoV-2 positive patients in our cohort died during their hospitalization, of whom five had normal CXRs on admission. CONCLUSION: Sixty (7.1%) of suspected COVID-19 patients hospitalized at 72 hours required respiratory support despite being normoxic in the ED. Further research should look to identify the normoxic SARS-CoV-2 patients at risk for decompensation.

Tracing Lung Cancer Risk Factors Through Mutational Signatures in Never-Smokers.

Epidemiologic studies often rely on questionnaire data, exposure measurement tools, and/or biomarkers to identify risk factors and the underlying carcinogenic processes. An emerging and promising complementary approach to investigate cancer etiology is the study of somatic "mutational signatures" that endogenous and exogenous processes imprint on the cellular genome. These signatures can be identified from a complex web of somatic mutations thanks to advances in DNA sequencing technology and analytical algorithms. This approach is at the core of the Sherlock-Lung study (2018-ongoing), a retrospective case-only study of over 2,000 lung cancers in never-smokers (LCINS), using different patterns of mutations observed within LCINS tumors to trace back possible exposures or endogenous processes. Whole genome and transcriptome sequencing, genome-wide methylation, microbiome, and other analyses are integrated with data from histological and radiological imaging, lifestyle, demographic characteristics, environmental and occupational exposures, and medical records to classify LCINS into subtypes that could reveal distinct risk factors. To date, we have received samples and data from 1,370 LCINS cases from 17 study sites worldwide and whole-genome sequencing has been completed on 1,257 samples. Here, we present the Sherlock-Lung study design and analytical strategy, also illustrating some empirical challenges and the potential for this approach in future epidemiologic studies.

Impact of a shelter-in-place order during the COVID-19 pandemic on the incidence of opioid overdoses.

INTRODUCTION: Since the beginning of the novel coronavirus (COVID-19) pandemic in the United States, there have been concerns about the potential impact of the pandemic on persons with opioid use disorder. Shelter-in-place (SIP) orders, which aimed to reduce the spread and scope of the virus, likely also impacted this patient population. This study aims to assess the role of the COVID-19 pandemic on the incidence of opioid overdose before and after a SIP order. METHODS: A retrospective review of the incidence of opioid overdoses in an urban three-hospital system was conducted. Comparisons were made between the first 100 days of a city-wide SIP order during the COVID-19 pandemic and the 100 days during the COVID-19 pandemic preceding the SIP order (Pre-SIP). Differences in observed incidence and expected incidence during the SIP period were evaluated using a Fisher's Exact test. RESULTS: Total patient visits decreased 22% from 46,078 during the Pre-SIP period to 35,971 during the SIP period. A total of 1551 opioid overdoses were evaluated during the SIP period, compared to 1665 opioid overdoses during the Pre-SIP period, consistent with a 6.8% decline. A Fisher's Exact Test demonstrated a p < 0.0001, with a corresponding Odds Ratio of 1.20 with a 95% confidence interval (1.12;1.29). CONCLUSION: The COVID-19 pandemic and the associated SIP order were associated with a statistically and clinically significant increase in the proportion of opioid overdoses in relation to the overall change in total ED visits.

Ridesharing as an Alternative to Ambulance Transport for Voluntary Psychiatric Patients in the Emergency Department.

INTRODUCTION: Emergency department (ED) crowding is a growing problem. Psychiatric patients have long ED lengths of stay awaiting placement and transportation to a psychiatric facility after disposition. METHODS: Retrospective analysis of length of ED stay after disposition for voluntary psychiatric patients before and after the use of Lyft ridesharing services for inter-facility transport. RESULTS: Using Lyft transport to an outside crisis center shortens time to discharge both statistically and clinically from 113 minutes to 91 minutes (p = 0.028) for voluntary psychiatric patients. Discharge time also decreased for involuntary patients from 146 minutes to 127 minutes (p = 0.0053). CONCLUSION: Ridesharing services may be a useful alternative to medical transportation for voluntary psychiatric patients.

Effects of multiple cycles of weight loss and regain on the body weight regulatory system in rats.

We studied the effects of multiple cycles of weight loss and regain on the defended body weight in rats. Thirty-six male Wistar rats were divided into three weight-matched groups: weight cyclers (n = 18), ad libitum-fed controls (n = 9), and maturity controls (n = 9). Cyclers underwent four rounds of 20% weight loss from 50% caloric restriction, each cycle followed by recovery to stable plateau weight on ad libitum feeding. Controls ate ad libitum. Maturity controls ate ad libitum and then weight cycled the final two rounds to evaluate the effect of age in later cycles. Cyclers' postdiet plateau weight became progressively lower than that of controls. With each weight loss, ghrelin increased, while insulin and leptin decreased; the magnitude of these changes did not differ across cycles. After four rounds, cyclers' weight (504 ± 7 vs. 540 ± 22 g; P < 0.05) and percent body fat (11.7 vs. 15.2%; P < 0.05) were lower than in controls. After a 4-mo follow-up period of ad libitum feeding, cyclers maintained a lower total fat-pad mass versus controls (8.6 ± 0.5 vs. 15.9 ± 3.6 g; P < 0.01) and a lower glucose area-under-the-curve on oral glucose tolerance tests (P < 0.05). Repeated weight-loss cycles exerted positive effects, durably lowering defended levels of body adiposity and improving glucose tolerance.

Evaluation of Oral Cavity DNA Extraction Methods on Bacterial and Fungal Microbiota.

The objective of this study was to evaluate the most effective method of DNA extraction of oral mouthwash samples for use in microbiome studies that utilize next generation sequencing (NGS). Eight enzymatic and mechanical DNA extraction methods were tested. Extracted DNA was amplified using barcoded primers targeting the V6 variable region of the bacterial 16S rRNA gene and the ITS1 region of the fungal ribosomal gene cluster and sequenced using the Illumina NGS platform. Sequenced reads were analyzed using QIIME and R. The eight methods yielded significantly different quantities of DNA (p < 0.001), with the phenol-chloroform extraction method producing the highest total yield. There were no significant differences in observed bacterial or fungal Shannon diversity (p = 0.64, p = 0.93 respectively) by extraction method. Bray-Curtis beta-diversity did not demonstrate statistically significant differences between the eight extraction methods based on bacterial (R2 = 0.086, p = 1.00) and fungal (R2 = 0.039, p = 1.00) assays. No differences were seen between methods with or without bead-beating. These data indicate that choice of DNA extraction method affect total DNA recovery without significantly affecting the observed microbiome.

Association of Cognitive Impairment in Patients on 3-Hydroxy-3-Methyl-Glutaryl-CoA Reductase Inhibitors.

BACKGROUND: Atherosclerotic cardiovascular diseases are the leading cause of death in the United States. A reduction in cholesterol with 3-hydroxy-3-methyl-glutaryl-CoA reductase inhibitors (statin) significantly reduces mortality and morbidity. Statins may be associated with cognitive impairment or dementia. Our aim was to study the association of cognitive impairment or dementia in patients who were on a statin. METHODS: Electronic medical records of 3,500 adult patients in our suburban internal medicine office were reviewed. RESULTS: There were 720 (20.6%) patients in the statin treatment group. Dementia or cognitive impairment was an associated comorbid condition in 7.9% patients in the statin treatment group compared to 3.1% patients in the non-statin group (P < 0.001). Analysis of all of the patients with cognitive impairment or dementia showed that among the age ranges of 51 years through 100 years, the patients in the statin treatment group had a higher prevalence of cognitive impairment or dementia compared to the non-statin group. In the statin treatment group, we found significantly higher prevalence of hyperlipidemia (86.3%), hypertension (69.6%), diabetes mellitus (36.0%), osteoarthritis (31.5%), coronary artery disease (26.1%), hypothyroidism (21.5%) and depression (19.3%) compared to the non-statin group (P < 0.001). About 39.9% of the patients with dementia or cognitive impairment were on statin therapy compared to 18.9% patients who had no dementia or cognitive impairment and were on statin therapy (P < 0.001). Among the patients with cognitive deficit or dementia in the statin treatment group, the majority of the patients were either on atorvastatin (43.9%) or simvastatin (35.1%), followed by rosuvastatin (12.2%) and pravastatin (8.8%). We found greater odds of dementia or cognitive impairment with each year increase in age (1.3 times), in women (2.2 times), African American race (2.7 times), non-consumption of moderate amount of alcohol (two times), diabetes mellitus (1.6 times), hypothyroidism (1.7 times), cerebrovascular accident (3.2 times), and other rheumatological diseases (1.8 times). CONCLUSIONS: The association of dementia or cognitive impairment was significantly higher in the patients who were on statin therapy compared to the patients who were not on a statin.

Did death certificates and a death review process agree on lung cancer cause of death in the National Lung Screening Trial?

BACKGROUND/AIMS: Randomized controlled trials frequently use death review committees to assign a cause of death rather than relying on cause of death information from death certificates. The National Lung Screening Trial, a randomized controlled trial of lung cancer screening with low-dose computed tomography versus chest X-ray for heavy and/or long-term smokers ages 55-74 years at enrollment, used a committee blinded to arm assignment for a subset of deaths to determine whether cause of death was due to lung cancer. METHODS: Deaths were selected for review using a pre-determined computerized algorithm. The algorithm, which considered cancers diagnosed during the trial, causes and significant conditions listed on the death certificate, and the underlying cause of death derived from death certificate information by trained nosologists, selected deaths that were most likely to represent a death due to lung cancer (either directly or indirectly) and deaths that might have been erroneously assigned lung cancer as the cause of death. The algorithm also selected deaths that might be due to adverse events of diagnostic evaluation for lung cancer. Using the review cause of death as the gold standard and lung cancer cause of death as the outcome of interest (dichotomized as lung cancer versus not lung cancer), we calculated performance measures of the death certificate cause of death. We also recalculated the trial primary endpoint using the death certificate cause of death. RESULTS: In all, 1642 deaths were reviewed and assigned a cause of death (42% of the 3877 National Lung Screening Trial deaths). Sensitivity of death certificate cause of death was 91%; specificity, 97%; positive predictive value, 98%; and negative predictive value, 89%. About 40% of the deaths reclassified to lung cancer cause of death had a death certificate cause of death of a neoplasm other than lung. Using the death certificate cause of death, the lung cancer mortality reduction was 18% (95% confidence interval: 4.2-25.0), as compared with the published finding of 20% (95% confidence interval: 6.7-26.7). CONCLUSION: Death review may not be necessary for primary-outcome analyses in lung cancer screening trials. If deemed necessary, researchers should strive to streamline the death review process as much as possible.

The National Lung Screening Trial's Endpoint Verification Process: determining the cause of death.

BACKGROUND: Randomized controlled trials (RCTs) evaluating cancer screening modalities usually employ cause-specific mortality as their primary endpoint. Because death certificate cause of death can be inaccurate, RCTs frequently use review committees to assign an underlying cause of death. We describe the National Lung Screening Trial's (NLST's) death review approach, the Endpoint Verification Process (EVP), which strives to minimize errors in assignment of cause of death due to lung cancer. METHODS: Deaths selected for review include those with a notation of lung cancer on the death certificate and those occurring among participants ever diagnosed with lung cancer. Other criteria that trigger death review include, but are not limited to, death within 6 months of a screen suspicious for lung cancer and death within 60 days of certain diagnostic evaluation procedures associated with a screen suspicious for lung cancer or a lung cancer diagnosis. EVP requires concordance on whether death was due to lung cancer. Deaths are first reviewed by the EVP chair. If concordance is not achieved, the death is next reviewed by an Endpoint Verification Team (EVT) member. If concordance between the chair- and member-assigned cause of death is not achieved, the death is next reviewed by a group of at least three EVT members. Cause of death is assigned at the step in which concordance was achieved, or if necessary, at the team review. CONCLUSIONS: NLST's EVP is designed to produce a highly accurate count of lung cancer deaths.

Cutting edge: vasoactive intestinal peptide (VIP) induces differentiation of Th17 cells with a distinctive cytokine profile.

Immune cellular effects of vasoactive intestinal peptide (VIP) are transduced by VIP G protein-coupled receptors type 1 (VPAC1) and type 2 (VPAC2). We now show that VIP with TGFbeta stimulates the transformation of CD4 T cells to a distinctive type of Th17 cell that generates IL-17 but not IL-6 or IL-21. VIP induction of Th17 cells was higher in VPAC2 knockout mice than wild-type mice, suggesting that VPAC1 is the principal transducer. Compared with Th17 cells elicited by IL-6, those evoked by VIP were similar in the secretion of IL-17 and IL-22, but lacked IL-21 secretion. Suppression of VIP induction of Th17 cells by protein kinase A inhibitors and enhancement by pharmacologically increased cAMP supports a role for this signal. The ability of VIP-VPAC1 axis signals to evoke development of a novel type of Th17 cells demonstrates the unique specificity of neuroregulatory mechanisms in the immunological environment.

Plan and operation of Cycle 6 of the National Survey of Family Growth.

OBJECTIVES: This report describes how Cycle 6 of the National Survey of Family Growth (NSFG) was designed, planned, and implemented. The NSFG is a national survey of women and men 15-44 years of age designed to provide national estimates of factors affecting pregnancy and birth rates; men's and women's health; and parenting. Cycle 6, conducted in 2002, was the first time the NSFG included a sample of males. METHODS: The survey used in-person, face-to-face interviews conducted by trained female interviewers. One person per household was interviewed from a national area probability sample in about 120 sample areas, with oversamples of teenagers, African Americans, and Hispanics. The data collection used computer-assisted personal interviewing (CAPI). Separate questionnaires were used for female and male respondents. The last section of the questionnaires used a technique called audio computer-assisted self-interviewing (ACASI). In order to control costs and nonresponse errors, survey managers statistically analyzed results from interviewers' visits to sampled households each day, and used those results to allocate interviewer labor and other resources more efficiently. This management improved response rates and made the sample more representative. RESULTS: Over 12,500 interviews were completed, about 7,600 with females and about 4,900 with males. The response rate was about 80 percent for females and about 78 percent for males. The survey procedures were adapted during the fieldwork to achieve the desired response rates and to control costs.