Liquid biopsy assay for lung carcinoma using centrifuged supernatants from fine-needle aspiration specimens.

INTRODUCTION: Tumor mutation profiling is standard-of-care in lung carcinoma patients. However, comprehensive molecular profiling of small specimens, including core needle biopsy (CNB) and fine-needle aspiration (FNA) specimens, may often be inadequate due to limited tissue. Centrifuged FNA supernatants, which are typically discarded, have emerged recently as a novel liquid-based biopsy for molecular testing. In this study, we evaluate the use of lung carcinoma FNA supernatants for detecting clinically relevant mutations. METHODS: Supernatants from lung carcinoma FNA samples (n = 150) were evaluated. Samples were further analyzed using next-generation sequencing (NGS) and ultrasensitive droplet digital PCR (ddPCR). Mutation profiles in a subset of samples were compared with results derived from paired tissue samples from the same patient (n = 67) and available plasma liquid biopsy assay (n = 45). RESULTS: All 150 samples yielded adequate DNA and NGS were carried out successfully on 104 (90%) of 116 selected samples. Somatic mutations were detected in 82% of the samples and in 50% of these patients a clinically relevant mutation was identified that would qualify them for targeted therapy or a clinical trial. There was high overall concordance between the mutation profiles of supernatants and the corresponding tissue samples, with 100% concordance with concurrent FNA and 96% with concurrent CNB samples. Comparison of actionable driver mutations detected in supernatant versus plasma samples showed 84% concordance. CONCLUSIONS: FNA supernatants can provide a valuable specimen source for genotyping lung carcinoma especially in patients with insufficient tumor tissue, thereby reducing multigene mutation profiling failure rates, improving turnaround times, and avoiding repeat biopsies.

Suppressed immune microenvironment and repertoire in brain metastases from patients with resected non-small-cell lung cancer.

BACKGROUND: The tumor immune microenvironment (TIME) of lung cancer brain metastasis is largely unexplored. We carried out immune profiling and sequencing analysis of paired resected primary tumors and brain metastases of non-small-cell lung carcinoma (NSCLC). PATIENTS AND METHODS: TIME profiling of archival formalin-fixed and paraffin-embedded specimens of paired primary tumors and brain metastases from 39 patients with surgically resected NSCLCs was carried out using a 770 immune gene expression panel and by T-cell receptor beta repertoire (TCRß) sequencing. Immunohistochemistry was carried out for validation. Targeted sequencing was carried out to catalog hot spot mutations in cancer genes. RESULTS: Somatic hot spot mutations were mostly shared between both tumor sites (28/39 patients; 71%). We identified 161 differentially expressed genes, indicating inhibition of dendritic cell maturation, Th1, and leukocyte extravasation signaling pathways, in brain metastases compared with primary tumors (P < 0.01). The proinflammatory cell adhesion molecule vascular cell adhesion protein 1 was significantly suppressed in brain metastases compared with primary tumors. Brain metastases exhibited lower T cell and elevated macrophage infiltration compared with primary tumors (P < 0.001). T-cell clones were expanded in 64% of brain metastases compared with their corresponding primary tumors. Furthermore, while TCR repertoires were largely shared between paired brain metastases and primary tumors, T-cell densities were sparse in the metastases. CONCLUSION: We present findings that suggest that the TIME in brain metastases from NSCLC is immunosuppressed and comprises immune phenotypes (e.g. immunosuppressive tumor-associated macrophages) that may help guide immunotherapeutic strategies for NSCLC brain metastases.

Hypercapnia-induced vasodilation in the cerebral circulation is reduced in older adults with sleep-disordered breathing.

The prevalence of sleep-disordered breathing (SDB) is higher in older adults compared with younger individuals. The increased propensity for ventilatory control instability in older adults may contribute to the increased prevalence of central apneas. Reductions in the cerebral vascular response to CO2 may exacerbate ventilatory overshoots and undershoots during sleep. Thus, we hypothesized that hypercapnia-induced cerebral vasodilation (HCVD) will be reduced in older compared with younger adults. In 11 older and 10 younger adults with SDB, blood flow velocity in the middle cerebral artery (MCAV) was measured using Doppler transcranial ultrasonography during multiple steady-state hyperoxic hypercapnic breathing trials while awake, interspersed with room air breathing. Changes in ventilation, MCAV, and mean arterial pressure (MAP) via finger plethysmography during the trials were compared with baseline eupneic values. For each hyperoxic hypercapnic trial, the change (Δ) in MCAV for a corresponding change in end-tidal CO2 and the HCVD or the change in cerebral vascular conductance (MCAV divided by MAP) for a corresponding change in end-tidal CO2 was determined. The hypercapnic ventilatory response was similar between the age groups, as was ΔMCAV/Δ[Formula: see text]. However, compared with young, older adults had a significantly smaller HCVD (1.3 ± 0.7 vs. 2.1 ± 0.6 units/mmHg, P = 0.004). Multivariable analyses demonstrated that age and nadir oxygen saturation during nocturnal polysomnography were significant predictors of HCVD. Thus, our data indicate that older age and SDB-related hypoxia are associated with diminished HCVD. We hypothesize that this impairment in vascular function may contribute to breathing instability during sleep in these individuals.NEW & NOTEWORTHY This study demonstrates, for the first time, in individuals with sleep-disordered breathing (SDB) that aging is associated with decreased hypercapnia-induced cerebral vasodilation (HCVD). In addition to advanced age, the magnitude of nocturnal oxygen desaturation due to SDB is an equal independent predictor of HCVD.

Amelioration of sleep-disordered breathing with supplemental oxygen in older adults.

Elderly adults demonstrate increased propensity for breathing instability during sleep compared with younger adults, and this may contribute to increased prevalence of sleep-disordered breathing (SDB) in this population. Hence, in older adults with SDB, we examined whether addition of supplemental oxygen (O2) will stabilize breathing during sleep and alleviate SDB. We hypothesized that exposure to supplemental O2 during non-rapid eye movement (NREM) sleep will stabilize breathing and will alleviate SDB by reducing ventilatory chemoresponsiveness and by widening the carbon dioxide (CO2) reserve. We studied 10 older adults with mild-to-moderate SDB who were randomized to undergo noninvasive bilevel mechanical ventilation with exposure to room air or supplemental O2 (Oxy) to determine the CO2 reserve, apneic threshold (AT), and controller and plant gains. Supplemental O2 was introduced during sleep to achieve a steady-state O2 saturation ≥95% and fraction of inspired O2 at 40%-50%. The CO2 reserve increased significantly during Oxy versus room air (-4.2 ± 0.5 mmHg vs. -3.2 ± 0.5 mmHg, P = 0.03). Compared with room air, Oxy was associated with a significant decline in the controller gain (1.9 ± 0.4 L/min/mmHg vs. 2.5 ± 0.5 L/min/mmHg, P = 0.04), with reductions in the apnea-hypopnea index (11.8 ± 2.0/h vs. 24.4 ± 5.6/h, P = 0.006) and central apnea-hypopnea index (1.7 ± 0.6/h vs. 6.9 ± 3.9/h, P = 0.03). The AT and plant gain were unchanged. Thus, a reduced slope of CO2 response resulted in an increased CO2 reserve. In conclusion, supplemental O2 reduced SDB in older adults during NREM sleep via reduction in chemoresponsiveness and central respiratory events.NEW & NOTEWORTHY This study demonstrates for the first time in elderly adults without heart disease that intervention with supplemental oxygen in the clinical range will ameliorate central apneas and hypopneas by decreasing the propensity to central apnea through decreased chemoreflex sensitivity, even in the absence of a reduction in the plant gain. Thus, the study provides physiological evidence for use of supplemental oxygen as therapy for mild-to-moderate SDB in this vulnerable population.

Adrenal tumor presenting as precocious puberty.

We present a case report of a two and a half-year-old boy who presented with precocious puberty. A clinical diagnosis of congenital adrenal hyperplasia was made. Patient was investigated and found to have an adrenocortical tumor. The tumor was about 7 cms in diameter. The tumor was secreting androgens, 17OHP and cortisol. This is an unusual array of hormones to be secreted by an adrenal tumor.

Frequent homozygous deletion of the LKB1/STK11 gene in non-small cell lung cancer.

LKB1/STK11 is a tumor suppressor and a negative regulator of mammalian target of rapamycin signaling. It is inactivated in 30% of lung cancer cell lines but only 5-15% of primary lung adenocarcinomas. There is evidence that homozygous deletion (HD) of chromosome 19p at the LKB locus contributes to the inactivation of the gene in primary human lung cancers. Here, we used several complementary genetic approaches to assess the LKB1 locus in primary non-small cell lung cancers (NSCLCs). We first analyzed 124 NSCLC cases for allelic imbalance using eight microsatellite markers on chromosome 19p, which revealed an overall rate of 65% (80 of 124) loss of heterozygosity (LOH). We next used chromogenic in situ hybridization (CISH) to directly examine the chromosomal status of the LKB1 locus. In all, 65 of 124 LOH tested samples were available for CISH and 58 of those (89%) showed either loss of one copy of chromosome 19p (LOH, 40 of 65 cases, 62%) or both copies (HD 18 of 65 cases, 28%). The occurrence of HD was significantly more frequent in Caucasian (35%) than in African-American patients (6%) (P=0.04). A total of 62 of 124 samples with LOH at one or both markers immediately flanking the LKB1 gene were further analyzed by directly sequencing the complete coding region, which identified 7 of 62 (11%) tumors with somatic mutations in the gene. Jointly, our data identified total inactivation of the LKB1 gene by either HD or LOH with somatic mutation in 39% of tested samples, whereas loss of chromosome 19p region by HD or LOH at the LKB1 region occured in 90% of NSCLC.

Sleep and daytime sleepiness in upper airway resistance syndrome compared to obstructive sleep apnoea syndrome.

This study has investigated differences in the nocturnal sleep and daytime sleepiness among patients with obstructive sleep apnoea syndrome (OSAS), upper airway resistance (UARS), sleep hypopnoea syndrome, and normal control subjects, using sleep scoring and spectral activity analysis of the electroencephalogram (EEG). Twelve nonobese males with UARS aged 30-60 yrs were recruited. These subjects were strictly matched for age and body mass index with twelve OSAS patients, 12 sleep hypopnoea syndrome patients, and 12 normal controls, all male. Daytime sleepiness was evaluated using the Epworth Sleepiness Scale (ESS) and the Multiple Sleep Latency Test (MSLT). The macrostructure of sleep was determined using international criteria and spectral analysis of the sleep EEG was obtained from a central lead. The sleep macrostructure of OSAS and UARS patients was significantly different from that of controls. These patients were also sleepier during the daytime than controls. Complaints of tiredness and daytime sleepiness, ESS and MSLT scores were similar in the different patient groups. Mild dysmorphia was present in all three patient groups. However, nocturnal sleep was significantly different among the different groups. OSAS patients had significantly more awake time during sleep than the UARS patients. The spectral activity of the total sleep time of the patient groups also differed significantly from that of controls. When the sleep spectral activity of UARS and OSAS patients were compared, OSAS patients had less slow wave sleep activity than UARS patients. UARS patients had a significantly higher absolute power in the 7-9 Hz bandwidth than OSAS patients. The absolute delta power over the different sleep cycles was also different between controls and patients, and between UARS and OSAS patients. There are clear differences in the macrostructure and spectral activity of sleep between upper airway resistance and obstructive sleep apnoea syndrome patients, demonstrated by differences in the cortical activity recorded in the central lead during sleep. Despite these nocturnal sleep differences, the tests of subjective daytime sleepiness are not significantly different.

Normal pregnancy, daytime sleeping, snoring and blood pressure.

Objective: Investigation of daytime sleepiness, blood pressure changes and presence of sleep disordered breathing, in healthy young women during pregnancy.Methods: Young, healthy pregnant women between 18 and 32 years of age, seen in three different prenatal care clinics, were enlisted in a prospective study divided in two parts: part 1 of the study consisted of completing a standardized questionnaire on past and present sleep disorders. It also included filling out visual analog scales (VAS) for daytime sleepiness and snoring by the subject and bed partner. Blood pressure measurement was performed at 9 AM as per the WHO protocol. Similar data were collected again at the 6-month prenatal visit and at the 3-month post-delivery visit. At the 6-month visit, ambulatory monitoring of nocturnal sleep using a portable six-channel recorder (Edentrace((R))) was performed at home. Part 2 involved a subgroup of subjects that were randomly selected after stratification based on results of VAS and ambulatory monitoring. It included 1 night of nocturnal polysomnography with esophageal manometry and 24 h of ambulatory BP monitoring with portable equipment with cuff inflation every 30 min.Results: Of the 267 women who participated in part 1 of the study, only 128 consented to enroll in part 2, from which 26 were selected to undergo polysomnography. At the 6-week prenatal visit 37.45% of the subjects reported daytime sleepiness of variable severity. At the 6-month visit, this was noted in 52% of the subjects. Bed-partners reported chronic, loud snoring prior to pregnancy in 3.7% of the study population, but this increased to 11.8% at the 6-month visit. Blood pressure (BP) remained below the pathological range, i.e. less than 150/95 mm Hg, during the entire pregnancy. However, ambulatory monitoring indicated that 37 women, including the loud chronic snorers, had some minor SaO(2) drops during sleep and this same group presented the largest increase in BP between the 6th week and the 6th month prenatal visits. Part 2 included 26 women, 13 from the above identified 37 women and 13 from the rest of the group, chosen randomly, age and body mass index (BMI) matched. Polysomnography did identify two abnormal breathing patterns during sleep: (1) esophageal pressure 'crescendos' associated predominantly with stage 1 and 2 NREM sleep, and (2) 'abnormal sustained efforts' seen predominantly with delta sleep. These abnormal breathing patterns were noted during a significantly longer time during sleep. This group of women with the abnormal breathing patterns were not only chronic snorers but also had significantly higher systolic and diastolic BP increases when compared to the 13 other non-snorers. Six out of the 13 snorers were 'non-dippers' at the 24-h BP recording.Conclusion: Abnormal breathing during sleep (that is frequently, but not always, associated with loud, chronic snoring, and may be a consequence of edema induced by hormonal changes associated with pregnancy), can be seen in otherwise healthy young pregnant women. It may contribute to the symptom of daytime sleepiness. The changes in blood pressure noted were of no pathological significance in our population but could be an added risk factor in high-risk pregnancies.

Optimization of Cd1-xZnxTe Detectors for Digital Radiography.

In this study, measurements of the electrical and detection parameters of the Cd1-xZnxTe detectors, within the x-ray diagnostic energy range, have been performed with the aim of optimizing the image quality parameters of these solid-state-ionization detectors. Namely, the leakage current and system capacitance of the x-ray imaging system have been measured as they relate to signal parameters. Similarly, the detected signal and noise contributions were measured and related to the radiation exposure and tube current setting. Furthermore, the detector contrast has been experimentally determined. The experimental results indicate that Cd1-xZnxTe detectors have low leakage current, high resistivity, and high detector contrast resolution. Therefore, they appear to be very attractive for imaging applications with applications in x-ray digital radiography.

Electric Field Dependence on Charge Collection of CdZnTe X-Ray Detectors.

In this study, the electric field dependence on the charge collection process of CdZnTe detectors, at different x-ray tube settings, within the x-ray diagnostic energy range, is investigated. In addition, the detector contrast at different applied bias voltages and x-ray tube settings have been experimentally determined. The experimental results suggest that an efficient charge collection process is obtained by increasing the applied bias voltage. Once the applied bias voltage is sufficiently high, charge collection becomes complete and the detector operates in the saturation region. This is a prerequisite for high contrast and spatial resolution. As a result, the detector contrast is enhanced significantly. Therefore, CdZnTe detectors appear to be potential candidates for digital radiographic applications.