Noninvasive Blood Test Research Articles

Clinical validation of a spectroscopic liquid biopsy for earlier detection of brain cancer.

BackgroundDiagnostic delays impact the quality of life and survival of patients with brain tumors. Earlier and expeditious diagnoses in these patients are crucial to reduce the morbidities and mortalities associated with brain tumors. A simple, rapid blood test that can be administered easily in a primary care setting to efficiently identify symptomatic patients who are most likely to have a brain tumor would enable quicker referral to brain imaging for those who need it most.MethodsBlood serum samples from 603 patients were prospectively collected and analyzed. Patients either had non-specific symptoms that could be indicative of a brain tumor on presentation to the Emergency Department, or a new brain tumor diagnosis and referral to the neurosurgical unit, NHS Lothian, Scotland. Patient blood serum samples were analyzed using the Dxcover® Brain Cancer liquid biopsy. This technology utilizes infrared spectroscopy combined with a diagnostic algorithm to predict the presence of intracranial disease.ResultsOur liquid biopsy approach reported an area under the receiver operating characteristic curve of 0.8. The sensitivity-tuned model achieves a 96% sensitivity with 45% specificity (NPV 99.3%) and identified 100% of glioblastoma multiforme patients. When tuned for a higher specificity, the model yields a sensitivity of 47% with 90% specificity (PPV 28.4%).ConclusionsThis simple, non-invasive blood test facilitates the triage and radiographic diagnosis of brain tumor patients while providing reassurance to healthy patients. Minimizing time to diagnosis would facilitate the identification of brain tumor patients at an earlier stage, enabling more effective, less morbid surgical and adjuvant care.

Novel blood test for early biomarkers of preeclampsia and Alzheimer\u2019s disease

A non-invasive and sensitive blood test has long been a goal for early stage disease diagnosis and treatment for Alzheimer’s disease (AD) and other proteinopathy diseases. We previously reported that preeclampsia (PE), a severe pregnancy complication, is another proteinopathy disorder with impaired autophagy. We hypothesized that induced autophagy deficiency would promote accumulation of pathologic protein aggregates. Here, we describe a novel, sensitive assay that detects serum protein aggregates from patients with PE (n = 33 early onset and 33 late onset) and gestational age-matched controls (n = 77) as well as AD in both dementia and prodromal mild cognitive impairment (MCI, n = 24) stages with age-matched controls (n = 19). The assay employs exposure of genetically engineered, autophagy-deficient human trophoblasts (ADTs) to serum from patients. The aggregated protein complexes and their individual components, including transthyretin, amyloid β-42, α-synuclein, and phosphorylated tau231, can be detected and quantified by co-staining with ProteoStat, a rotor dye with affinity to aggregated proteins, and respective antibodies. Detection of protein aggregates in ADTs was not dependent on transcriptional upregulation of these biomarkers. The ROC curve analysis validated the robustness of the assay for its specificity and sensitivity (PE; AUC: 1, CI: 0.949–1.00; AD; AUC: 0.986, CI: 0.832–1.00). In conclusion, we have developed a novel, noninvasive diagnostic and predictive assay for AD, MCI and PE.

Donor-derived cell-free DNA accurately detects acute rejection in lung transplant patients, a multicenter cohort study

Acute rejection, which includes antibody-mediated rejection and acute cellular rejection, is a risk factor for lung allograft loss. Lung transplant patients often undergo surveillance transbronchial biopsies to detect and treat acute rejection before irreversible chronic rejection develops. Limitations of this approach include its invasiveness and high interobserver variability. We tested the performance of percent donor-derived cell-free DNA (%ddcfDNA), a non-invasive blood test, to detect acute rejection. This multicenter cohort study monitored 148 lung transplant subjects over a median of 19.6 months. We collected serial plasma samples contemporaneously with TBBx to measure %ddcfDNA. Clinical data was collected to adjudicate for acute rejection. The primary analysis consisted of computing the area-under-the-receiver-operating-characteristic-curve of %ddcfDNA to detect acute rejection. Secondary analysis determined %ddcfDNA rule-out thresholds for acute rejection. ddcfDNA levels were high after transplant surgery and decayed logarithmically. With acute rejection, ddcfDNA levels rose six-fold higher than controls. ddcfDNA levels also correlated with severity of lung function decline and histological grading of rejection. %ddcfDNA area-under-the-receiver-operating-characteristic-curve for acute rejection, AMR, and ACR were 0.89, 0.93, and 0.83, respectively. ddcfDNA levels of <0.5% and <1.0% showed a negative predictive value of 96% and 90% for acute rejection, respectively. Histopathology detected one-third of episodes with ddcfDNA levels ≥1.0%, even though >90% of these events were coincident to clinical complications missed by histopathology. This study demonstrates that %ddcfDNA reliably detects acute rejection and other clinical complications potentially missed by histopathology, lending support to its use as a non-invasive marker of allograft injury.

Colorectal cancer screening using a stool DNA-based SDC2 methylation test: a multicenter, prospective trial

BackgroundPrevention and early detection of colorectal cancer (CRC) is a global priority, with many countries conducting population-based CRC screening programs. Although colonoscopy is the most accurate diagnostic method for early CRC detection, adherence remains low because of its invasiveness and the need for extensive bowel preparation. Non-invasive fecal occult blood tests or fecal immunochemical tests are available; however, their sensitivity is relatively low. Syndecan-2 (SDC2) is a stool-based DNA methylation marker used for early detection of CRC. Using the EarlyTect™-Colon Cancer test, the sensitivity and specificity of SDC2 methylation in stool DNA for detecting CRC were previously demonstrated to be greater than 90%. Therefore, a larger trial to validate its use for CRC screening in asymptomatic populations is now required.MethodsAll participants will collect their stool (at least 20 g) before undergoing screening colonoscopy. The samples will be sent to a central laboratory for analysis. Stool DNA will be isolated using a GT Stool DNA Extraction kit, according to the manufacturer’s protocol. Before performing the methylation test, stool DNA (2 µg per reaction) will be treated with bisulfite, according to manufacturer’s instructions. SDC2 and COL2A1 control reactions will be performed in a single tube. The SDC2 methylation test will be performed using an AB 7500 Fast Real-time PCR system. CT values will be calculated using the 7500 software accompanying the instrument. Results from the EarlyTect™-Colon Cancer test will be compared against those obtained from colonoscopy and any corresponding diagnostic histopathology from clinically significant biopsied or subsequently excised lesions. Based on these results, participants will be divided into three groups: CRC, polyp, and negative. The following clinical data will be recorded for the participants: sex, age, colonoscopy results, and clinical stage (for CRC cases).DiscussionThis trial investigates the clinical performance of a device that allows quantitative detection of a single DNA marker, SDC2 methylation, in human stool DNA in asymptomatic populations. The results of this trial are expected to be beneficial for CRC screening and may help make colonoscopy a selective procedure used only in populations with a high risk of CRC.Trial registration: This trial (NCT04304131) was registered at ClinicalTrials.gov on March 11, 2020 and is available at https://clinicaltrials.gov/ct2/show/NCT04304131?cond=NCT04304131&draw=2&rank=1.

Quantitative STAU2 measurement in lymphocytes for breast cancer risk assessment

Although mammograms play a key role in early breast cancer detection, the test is not applicable to all women, for example, women under the age of 40. The development of a noninvasive blood test with high sensitivity and accessibility will improve the effectiveness of breast cancer screening programmes. Secretory factors released from cancer cells can induce the expression of certain genes in a large number of white blood cells (WBCs). Therefore, cancer-dependent proteins in WBCs can be used as tumour markers with high sensitivity. Five proteins (LMAN1, AZI2, STAU2, MMP9 and PLOD1) from a systemic analysis of a variety of array data of breast cancer patients were subjected to immunofluorescence staining to evaluate the presence of fixed WBCs on 96-well plates from 363 healthy females and 358 female breast cancer patients. The results revealed that the average fluorescence intensity of anti-STAU2 and the percentage of STAU2-positive T and B lymphocytes in breast cancer patients (110.50 ± 23.38 and 61.87 ± 12.44, respectively) were significantly increased compared with those in healthy females (56.47 ± 32.03 and 33.02 ± 18.10, respectively) (p = 3.56 × 10–71, odds ratio = 24.59, 95% CI = 16.64–36.34). The effect of secreted molecules from breast cancer cells was proven by the increase in STAU2 intensity in PBMCs cocultured with MCF-7 and T47D cells at 48 h (p = 0.0289). The test demonstrated 98.32%, 82.96%, and 48.32% sensitivity and 56.47%, 83.47%, and 98.62% specificity in correlation with the percentage of STAU2-positive cells at 40, 53.34 and 63.38, respectively. We also demonstrated how to use the STAU2 test for the assessment of risk in women under the age of 40. STAU2 is a novel breast cancer marker that can be assessed by quantitative immunofluorescence staining of fixed WBCs that are transportable at room temperature via mail, representing a useful risk assessment tool for women without access to mammograms.

Evaluation of serum lidocaine/monoethylglycylxylidide concentration to assess shunt closure in dogs with extrahepatic portosystemic shunts.

BackgroundLiver function tests do not always normalize despite successful attenuation of extrahepatic portosystemic shunts (EHPSS).ObjectivesAssess the lidocaine/monoethylglycylxylidide (MEGX) test to determine liver perfusion after EHPSS closure.AnimalsTwenty dogs with EHPSS.MethodsA prospective cohort study was performed and all dogs were tested at diagnosis, 1, 3, and 6 months postoperatively. After collecting a baseline blood sample (T0), 1 mg/kg body weight of lidocaine was injected intravenously. Fifteen (T15) and 30 minutes (T30) later, blood was collected. Plasma concentrations of lidocaine and its metabolites MEGX and glycylxylidide (GX) were determined, using a high‐performance liquid chromatography with electrospray ionization tandem mass spectrometry method. Three months postoperatively, transsplenic portal scintigraphy was performed to determine EHPSS closure.ResultsAt T15, median MEGX concentrations were higher in dogs with closed EHPSS compared to diagnosis (33.73 ng/mL [21.11‐66.44 ng/mL] vs 13.74 ng/mL [7.25‐21.93 ng/mL]; P < .001), but were not different (12.28 ng/mL [10.62‐23.17 ng/mL] vs 13.74 ng/mL [7.25‐21.93 ng/mL]) in dogs with persistent shunting. Sensitivity to determine shunt closure for MEGX at T15 was 96.2% (95% confidence interval [CI]: 78.4‐99.8) and specificity 82.8% (95% CI: 63.5‐93.5).Conclusions and Clinical ImportanceThe lidocaine/MEGX test is a promising, rapid, and noninvasive blood test that seems helpful to differentiate dogs with closed EHPSS and dogs with persistent shunting after gradual attenuation.

Prevalence of liver fibrosis and cirrhosis in 699 Moroccan patients with chronic hepatitis C.

Introductionchronic hepatitis C (CHC) can cause severe complications, including fibrosis and cirrhosis. Very little is known about the prevalence of these complications in the Moroccan population.Methodsthe prevalence of liver fibrosis and cirrhosis using a non-invasive blood test (FibroTest and ActiTest) was studied in 699 Moroccan patients with CHC for 4 years (from January 2014 to December 2017). The serum immunological markers: α2-macroglobulin, haptoglobin, apolipoprotein A1 were analyzed nephelometrically on BN ProSpec® System. The serum biochemical markers: γ-glutamyltransferase, alanine aminotransferase, and bilirubin were performed using the VITROS® Chemistry System Ortho Clinical Diagnostic. A 699 patients with CHC were identified.Resultsthe overall prevalence of cirrhosis (F4) was estimated at 31.8%. Thirteen point nine percent (13.9%) of patients with cirrhosis had a risk of developing esophageal varices and a 3.3% risk of developing primary liver cancer. The association between cirrhosis and age showed an increase in prevalence after age 55 years old [OR=7.68(95%CI=4.9-12.2); p<0.0001]. No significant association for cirrhosis was found for sex.Conclusionaccording to the results of FibroTest, 32% of patients with CHC had cirrhosis. The older age was independently associated with liver cirrhosis.

Extracellular Vesicles in Hematological Malignancies: From Biomarkers to Therapeutic Tools.

Small extracellular vesicles (EVs) are a heterogenous group of lipid particles released by all cell types in physiological and pathological states. In hematological malignancies, tumor-derived EVs are critical players in mediating intercellular communications through the transfer of genetic materials and proteins between neoplastic cells themselves and to several components of the bone marrow microenvironment, rendering the latter a “stronger” niche supporting cancer cell proliferation, drug resistance, and escape from immune surveillance. In this context, the molecular cargoes of tumor-derived EVs reflect the nature and status of the cells of origin, making them specific therapeutic targets. Another important characteristic of EVs in hematological malignancies is their use as a potential “liquid biopsy” because of their high abundance in biofluids and their ability to protect their molecular cargoes from nuclease and protease degradation. Liquid biopsies are non-invasive blood tests that provide a molecular profiling clinical tool as an alternative method of disease stratification, especially in cancer patients where solid biopsies have limited accessibility. They offer accurate diagnoses and identify specific biomarkers for monitoring of disease progression and response to treatment. In this review, we will focus on the role of EVs in the most prevalent hematological malignancies, particularly on their prospective use as biomarkers in the context of liquid biopsies, as well as their molecular signature that identifies them as specific therapeutic targets for inhibiting cancer progression. We will also highlight their roles in modulating the immune response by acting as both immunosuppressors and activators of anti-tumor immunity.

Abstract 787: Comparable clinical benefits between low and high variant allelic frequency in NSCLC patients treated with targeted therapies based on molecular alterations from ctDNA NGS

Abstract Background: Tumor genomic information from tissue has been the standard of practice for identifying actionable molecular alterations. The same genomic profiling is also widely available by a non-invasive blood test from circulating tumor DNA (ctDNA). We hypothesized that patients with advanced non-small cell lung cancer (NSCLC) with actionable oncogenic driver mutations with either low or high variant allelic frequency (%VAF) identified by ctDNA would have similar clinical benefits after treatment with targeted therapies. Methods: In this retrospective study (UTMDACC IRB-approved protocol PA16-0061), patients with a targetable mutation identified by ctDNA NGS assay who received an FDA-approved therapy were identified in our GEMINI database. All potential patients were individually reviewed and information confirmed with review of their electronic medical records. Blood based genomic profiling was completed by NGS of ctDNA with a panel of up to 70 cancer-related genes at a CLIA-certified lab (Guardant360, Guardant Health, Redwood City, CA). The %VAF was reported for each mutation identified. We defined low group with VAF of 0.01% to 0.99% and high group with VAF &amp;gt; 1%. Tumor measurements were assessed using RECIST V1.1. For comparisons analysis, Kaplan-Meier estimator was used to estimate progression free survival (PFS) with Mantel-Cox and un-paired t-test was used for tumor responses. Results: Eighty-nine patients with advanced NSCLC treated with an FDA approved therapy based on ctDNA genomic profiling were identified and the results are summarized in table below: LOW %VAFHIGH %VAFP valuegroup (n=39)group (n=50)Gender (Female/Male)20/1928/22Age (median, range)54(27-85)63(29-89)Targetable mutations:EGFR2246ALK114MET exon 1420ROS120BRAF20Line of therapy (median, range)2(1-7)2(1-7)ALL PatientsTumor response (mean, range)-37.23% (-100 to +38)-31.08% (-100 to +59)0.42PFS (median, range)215 (27 – 848) days280 (43 – 894) days0.70EGFR onlyTumor response (mean, range)-24.86% (-100 to +16)-23.54% (-100 to +59)0.88PFS (median, range)189 (57 – 848) days293 (43-894) days0.23 Conclusions: In this analysis of patients who were treated with an FDA approved therapy based on genomic results from ctDNA profiling, there were no differences in either tumor response (P value = 0.42) or PFS (P value = 0.70) between those patients with low and high ctDNA as assessed by %VAF of identified targetable mutation. Similar findings were also observed in the analysis of the EGFR only subgroup. This study reveals patients with low %VAF received comparable clinical benefits as those with high %VAF and no deleterious harm was observed from treating patients with low %VAF with appropriate targeted therapy. Citation Format: Hai T. Tran, Vincent K. Lam, Mayra Vasquez, Yasir Elamin, Victoria Raymond, Lingzhi Hong, George Blumenschein, Brett Carter, Richard Lanman, Mehmet Altan, Ann Tsao, Lauren Byers, Don Gibbons, Jianjun Zhang, John V. Heymach. Comparable clinical benefits between low and high variant allelic frequency in NSCLC patients treated with targeted therapies based on molecular alterations from ctDNA NGS [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 787.

A simple, refined approach to diagnosing renovascular hypertension in children: A 10-year study.

Despite advances in non-invasive vascular imaging, detection of renal artery stenosis via catheter angiography is the criterion standard for the diagnosis of renovascular hypertension (RVH). However, because of lack of evidence, the utility of various blood tests and imaging modalities remains unclear. We retrospectively analyzed the utility of blood tests (plasma renin activity [PRA], aldosterone, and renal vein renin[RVR] values) and imaging studies (computed tomography angiography [CTA], kidney ultrasonography[US]) by comparing them with catheter angiography. Ten pediatric patients with RVH at two institutions from January 2008 to December 2017 were recruited. The sensitivities for diagnosing RVH via imaging and blood tests (kidney [US], PRA, and aldosterone) were derived by examining patient records. Furthermore, the sensitivity and specificity of CT angiography were calculated by considering both the affected and non-affected renal arteries of the patients. A high sensitivity for diagnosing RVH via kidney US(89%) and PRA (80%) was observed. The sensitivity and specificity of CTA were 100%, each. RVR sampling did not aid in the diagnosis of RVH; only two of six patients with unilateral RVH showed significant laterality of RVR boundary ratios. Renal scintigraphy facilitated detection of a non-functional kidney (split renal function <5%). RVH in children could be diagnosed utilizing non-invasive blood and imaging tests, without catheter angiography. We recommend kidney length measurement along with measurement of PRA level, as a simple and highly useful screening test, followed by CTA as a diagnostic test.

Non-invasive early detection of cancer four years before conventional diagnosis using a blood test

Early detection has the potential to reduce cancer mortality, but an effective screening test must demonstrate asymptomatic cancer detection years before conventional diagnosis in a longitudinal study. In the Taizhou Longitudinal Study (TZL), 123,115 healthy subjects provided plasma samples for long-term storage and were then monitored for cancer occurrence. Here we report the preliminary results of PanSeer, a noninvasive blood test based on circulating tumor DNA methylation, on TZL plasma samples from 605 asymptomatic individuals, 191 of whom were later diagnosed with stomach, esophageal, colorectal, lung or liver cancer within four years of blood draw. We also assay plasma samples from an additional 223 cancer patients, plus 200 primary tumor and normal tissues. We show that PanSeer detects five common types of cancer in 88% (95% CI: 80–93%) of post-diagnosis patients with a specificity of 96% (95% CI: 93–98%), We also demonstrate that PanSeer detects cancer in 95% (95% CI: 89–98%) of asymptomatic individuals who were later diagnosed, though future longitudinal studies are required to confirm this result. These results demonstrate that cancer can be non-invasively detected up to four years before current standard of care.

Fecal Occult Blood Tests in Colorectal Cancer Screening: Systematic Review and Meta-analysis of Traditional and New-generation Fecal Immunochemical Tests.

Noninvasive fecal occult blood tests (FOBTs) are recommended by current guidelines for colorectal cancer (CRC) screening. Our aim was to assess the diagnostic performance of traditional guaiac-based FOBTs (gFOBT) and new-generation immunochemical FOBTs (iFOBT) in CRC screening by carrying out a systematic review and meta-analysis. PubMed, Embase, Cochrane Library, and Web of Science were searched for eligible articles published before February 17, 2020. Three independent investigators conducted study assessment and data extraction. Diagnosis-related indicators for use of FOBTs in the detection of CRC (as the endpoint) in a screening setting were summarized, and further stratified by the type of FOBT (gFOBT vs. iFOBT). STATA software was used to conduct the meta-analysis. Pooled sensitivities and specificities were calculated using a random-effects model. Hierarchical summary receiver operating characteristic curves were plotted and area under the curves (AUC) were calculated. The electronic search identified 573 records after duplicates were removed, of which 75 full-text articles were assessed for eligibility. Finally, a total of 31 studies were eligible for the meta-analysis. In the ROC comparison test, there was a statistically significant difference in the performance of gFOBT and iFOBT tests, with AUC=0.77 (95% confidence intervaI=0.75-0.79) and AUC=0.87 (95% confidence intervaI=0.85-0.88), respectively (p=0.0017). In formal meta-regression, test brand did not prove to be a significant study-level covariate that would explain the observed heterogeneity between the studies. New-generation iFOBTs were found to have a significantly higher diagnostic performance as compared with gFOBTs, advocating the use of only fecal immunochemical tests in all newly implemented CRC screening programs.

Use of peripheral blood transcriptomic biomarkers to distinguish high-grade cervical squamous intraepithelial lesions from low-grade lesions.

It is crucial to classify cervical lesions into high-grade squamous intraepithelial lesions (HSILs) and low-grade SILs (LSILs), as LSILs are conservatively treated by observation, based on an expectation of natural regression, whereas HSILs usually require electrosurgical excision. In the present study, peripheral blood gene expression profiles were analyzed to identify transcriptomic biomarkers distinguishing HSILs from LSILs. A total of 102 blood samples were collected from women with cervical SILs (66 HSIL and 36 LSIL) for microarray hybridization. Candidate gene signatures were identified using AdaBoost algorithms, and a predictive model was constructed using logistic regression to differentiate HSILs from LSILs. To correct for possible bias as a result of the limited sample size and to verify the stability of the predictive model, a two-fold cross validation and null set analysis was conducted over 1,000 iterations. The functions of the transcriptomic biomarkers were then analyzed to elucidate the pathogenesis of cervical SIL. A total of 10 transcriptomic genes (STMN3, TRPC4AP, DYRK2, AGK, KIAA0319L, GRPEL1, ZFC3H1, LYL1, ITGB1 and ARHGAP18) were identified. The predictive model based on the 10-gene panel exhibited well-discriminated power. A cross validation process using known disease status exhibited almost the same performance as that of the predictive model, whereas null-set analysis with randomly reassigned disease status exhibited much lower predictive performance for distinguishing HSILs from LSILs. These biomarkers were involved in the ‘Rho GTPase cycle’, ‘mitochondrial protein import’, ‘oncogenic MAPK signaling’, ‘integrin cell surface interaction’ and ‘signaling by BRAF and RAF fusions’. In conclusion, peripheral blood gene expression analysis is a promising method for distinguishing HSILs from LSILs. The present study proposes 10 candidate genes that could be used in the future as diagnostic biomarkers and potential therapeutic targets for cervical SILs. A simple, non-invasive blood test would be clinically useful in the diagnosis and classification of patients with cervical SILs.

1448-P: Enhanced Liver Fibrosis (ELF) Test Can Accurately Predict Advanced Fibrosis in Patients with Type 2 Diabetes and Nonalcoholic Fatty Liver Disease (NAFLD)

Background: The ELF test (score calculated from markers of fibrosis: hyaluronic acid, procollagen III amino-terminal peptide, and tissue inhibitor of matrix metalloproteinase 1) is a non-invasive blood test used to estimate fibrosis stage in chronic liver disease. Since NAFLD patients with advanced fibrosis are at the highest risk for adverse long-term outcomes, the aim is to assess performance of ELF for identification of advanced fibrosis in patients with NAFLD. Methods: Patients with biopsy-proven NAFLD were included. ELF scores were calculated using an ADVIA Centaur XP analyzer. Results: There were 409 patients with NAFLD: 47 ± 12.9 years, 30% male, 70% white, 34% type 2 diabetes (T2DM), 56% hyperlipidemia, 54% hypertension, mean (SD) BMI 41.0 ± 10.0 kg/m2, ALT 48.4 ± 42.0 U/L, mean ELF score was 9.0 ± 1.3. Of the study cohort, 23% patients (N=93) had advanced fibrosis. Patients with advanced fibrosis had significantly higher ELF: 10.2 ± 1.3 vs. 8.6 ± 1.0, p&amp;lt;0.0001. There was a correlation between ELF and liver enzymes: rho=0.23 for ALT, 0.34 for AST (p&amp;lt;0.0001). ELF negatively correlated with BMI: rho = -0.28 (p&amp;lt;0.0001). Across all NAFLD patients, ELF had good performance for identifying patients with advanced fibrosis: area under curve (AUC) = 0.83 (0.79 - 0.87). The performance of ELF was similar in NAFLD patients with [AUC = 0.81 (0.73 - 0.87)] and without [AUC = 0.83 (0.77 - 0.87)] T2DM. With the commonly used cut-off of ELF ≥9.8 for prediction of advanced fibrosis, observed accuracy was as follows: sensitivity 62.4% (51.7% - 72.2%), specificity 89.6% (85.6% - 92.7%) positive predictive value (PPV) 63.7% (55.1% - 71.6%), negative predictive value (NPV) 89.0% (86.1% - 91.3%). Cut-off of 11.3 returned almost perfect specificity 99.0% (97.3% - 99.8%) with a sensitivity of 22.6% (14.6% - 32.4%), PPV 87.5% (68.1% - 95.8%), NPV 81.3% (79.6% - 82.9%). Conclusions: ELF test performs well in identifying high risk NAFLD patients with or without T2DM. Disclosure Z. Younossi: Consultant; Self; AbbVie Inc., Bristol-Myers Squibb, Gilead Sciences, Inc., Intercept Pharmaceuticals, Inc., Novo Nordisk Inc., Terns and Viking. S. Felix: None. T. Jeffers: None. E. Younossi: None. F. Nader: None. H. Pham: None. A. Afendy: None. R. Cable: None. A. Racila: None. Z. Younoszai: None. B.P. Lam: None. P. Golabi: None. L. Henry: None. S. Clement: None. M. Stepanova: None.

Enhanced liver fibrosis test for the non-invasive diagnosis of fibrosis in patients with NAFLD: A systematic review and meta-analysis

The enhanced liver fibrosis (ELF) test has been proposed for the non-invasive assessment of advanced fibrosis in patients with non-alcoholic fatty liver disease (NAFLD). We performed a systematic review to estimate the accuracy of this test against biopsy. In this systematic review, we searched MEDLINE, Embase, Web of Science and the Cochrane Library for studies that included patients with NAFLD and that used both liver biopsy (as the reference standard) and the ELF test. Two authors independently screened the references, extracted the data and assessed the quality of included studies. Due to the variation in reported thresholds, we used a multiple thresholds random effects model for meta-analysis (diagmeta R-package). The meta-analysis of 11 studies reporting advanced fibrosis and 5 studies reporting significant fibrosis showed that the ELF test had a sensitivity of >0.90 for excluding fibrosis at a threshold of 7.7. However, as a diagnostic test at high thresholds, the test only achieved specificity and positive predictive value >0.80 in very high prevalence settings (>50%). To achieve a specificity of 0.90 for advanced and significant fibrosis, thresholds of 10.18 (sensitivity: 0.57) and 9.86 (sensitivity: 0.55) were required, respectively. The ELF test showed high sensitivity but limited specificity to exclude advanced and significant fibrosis at low cut-offs. The diagnostic performance of the test at higher thresholds was found to be more limited in low-prevalence settings. We conclude that clinicians should carefully consider the likely disease prevalence in their practice setting and adopt suitable test thresholds to achieve the desired performance. The enhanced liver fibrosis test has been suggested as a non-invasive blood test to aid the diagnosis of severe liver fibrosis in patients with non-alcoholic fatty liver disease (NAFLD). Our study results showed that the test has a high negative predictive value, especially in populations with low disease prevalence (likely encountered in primary care); so, it can exclude advanced fibrosis in patients with NAFLD. However, when prevalence is low, the positive predictive value of the enhanced liver fibrosis test is low, suggesting that additional strategies may be needed to make a positive diagnosis in such settings.

Non-invasive Assessment of Liver Fibrosis

In the USA, there is a growing prevalence of liver disease, particularly from alcohol and fatty liver, as well as improvements in hepatitis C therapy that should be adjusted for level of fibrosis. There is accordingly a great need for rapid non-invasive fibrosis testing (NIT) to determine who should be referred for specialty care, started on specific therapies, screened for varices and malignancy, and selected for clinical trials. It is not only the accuracy of NIT that matters, but the order in which they are applied and the population selected for screening. All NIT to date have high negative predictive value for ruling out advanced fibrosis, and despite high specificity, have limited positive predictive value due to the relative infrequency of advanced fibrosis in screened populations. A testing algorithm based on primary care screening (e.g., with FIB-4) followed by referral for specialty confirmatory testing (e.g., vibration-controlled transient elastography) would best fit most practice models. This focused review will characterize the most validated non-invasive blood tests and imaging and discuss their practical applications for real-world use.

Combined AlloSure and AlloMap Testing in Multi-Organ Heart Transplantation Rejection Surveillance

AlloSure test is a non-invasive blood test designed to detect the presence of donor cell-free DNA in the recipients blood as a measure of the probability of rejection in heart transplant recipients. The result is displayed as a percent of donor-derived cell-free DNA (dd-cfDNA) in the totalcell-free DNA (cfDNA). Recommended threshold of 0.2% dd-cfDNA has been predictive of low likelihood of rejection. There is no data looking at its value and validation in multi-organ transplant. We aimed to study this area by reviewing our transplant database. All patients who received more than one solid organ transplant at our center and had AlloSure testing were included. Study outcomes included one year survival, rejection grade 2R or more, and hemodynamically significant graft dysfunction. There were (7) patients included in the study, average AlloSure score was 0.93% ±0.61% with an average AlloMap score of 33.8±2.14. Four patients (57.1%) had heart and kidney transplant, 3 patients (42.9%) had heart and liver transplantation. Average AlloSure score for heart/kidney transplant recipients was 0.52±0.17% with an average AlloMap score of 33.6±2.28. The while average AlloSure score for heart/liver transplant recipients was 1.21±0.55% mean AlloMap score of 33.93±2.32. None of our patients had graft dysfunction or biopsy-proven rejection >=2R. Survival rate was 100% for the first year in our patients CONCLUSION: AlloSure levels were higher in heart-liver vs. heart-kidney recipients, and in both cohorts higher than the 0.2% threshold recommended for heart transplant prediction of rejection, however did not predict rejection. In our cohort, while AlloMap scores were consistent in both groups, (33.93±2.32 vs. 33.6±2.28 respectively). Elevated AlloSure was not predictive of rejection in multiorgan heart transplant recipients.

Performance of a blood-based test for the detection of multiple cancer types.

283 Background: Cancers of the esophagus, stomach, pancreas, gallbladder, liver, bile duct, colon and rectum will account for 17% of incident cancer diagnoses and 26% of cancer-related deaths in the US in 2019. We developed a methylation-based cfDNA early multi-cancer detection test that also can predict the tissue of origin (TOO) of these and other cancers types; performance of this test for gastrointestinal (GI) tract cancers is reported here. Methods: The Circulating Cell-free Genome Atlas (CCGA; NCT02889978) study is a prospective, multi-center, observational, case-control study with longitudinal follow-up, enrolling individuals with cancer ( &gt; 20 cancers, all stages, newly diagnosed) and without cancer. Plasma cfDNA was subjected to a cross-validated targeted methylation (TM) sequencing assay. Methylation fragments were combined across targeted genomic regions and assigned a probability of cancer and a predicted TOO. GI cancer classes were upper GI (esophagus/stomach, n = 67), pancreas/gallbladder/extrahepatic bile duct (n = 95), liver/intrahepatic bile duct (n = 29), and colon/rectum (n = 121). Results: Detection across all GI cancers was 82% (95% CI 77-86) at a &gt; 99% pre-set specificity. Overall predicted TOO accuracy was 92% (88-95) among the samples for which TOO was predicted (6/255 had indeterminate predicted TOO). The table shows performance by GI cancer type. Conclusions: Simultaneous detection at high specificity ( &gt; 99%) of multiple cancer types, including GI cancers across stages at high sensitivity (82%), was shown using TM analysis of cfDNA. Accurate (92%) localization of cancers to specific regions of the GI tract was also achieved. Detection of multiple GI cancers from a single noninvasive blood test could be a practical method for detecting GI and other cancers, and may facilitate diagnostic work-ups. Clinical trial information: NCT02889978. [Table: see text]

Simultaneous multi-cancer detection and tissue of origin (TOO) localization using targeted bisulfite sequencing of plasma cell-free DNA (cfDNA).

44 Background: A noninvasive cfDNA blood test detecting multiple cancers at earlier stages could decrease cancer mortality. In earlier discovery work, whole-genome bisulfite sequencing outperformed whole-genome and targeted sequencing approaches for multi-cancer detection across stages at high specificity. Here, multi-cancer detection and TOO localization using bisulfite sequencing of plasma cfDNA to identify methylomic signatures was evaluated in preparation for clinical validation, utility, and implementation studies. Methods: 2301 analyzable participants (1422 cancer [ &gt; 20 tumor types, all stages], 879 non-cancer) were included in this prespecified substudy from the Circulating Cell-free Genome Atlas (CCGA) (NCT02889978) study - a prospective, multi-center, observational, case-control study with longitudinal follow-up. Plasma cfDNA was subjected to a targeted methylation sequencing assay using high-efficiency methylation chemistry to enrich for methylation targets, and a machine learning classifier determined cancer status and tissue of origin (TOO). Observed methylation fragments characteristic of cancer and TOO were combined across targeted regions and assigned a relative probability of cancer and of a specific TOO. Results: Performance is reported at 99% specificity (ie, a combined false positive rate across all cancer types of 1%), a level required for population-level screening. Across cancer types, sensitivity ranged from 59 to 86%. Combined cancer detection (sensitivity [95% CI]) was 34% (27-43%) in stage I (n = 151), 77% (70-83%) in stage II (n = 171), 84% (79-89%) in stage III (n = 204), and 92% (88-95%) in stage IV (n = 281). TOO was provided for 94% of all cancers detected; of these, TOO was correct in &gt; 90% of cases. Conclusions: Detection of multiple deadly cancers across stages using methylation signatures in plasma cfDNA was achieved with a single, fixed, low false positive rate, and simultaneously provided accurate TOO localization. This targeted methylation assay is undergoing validation in preparation for prospective clinical investigation as a cancer detection diagnostic. Clinical trial information: NCT02889978.

LBA83 - Final efficacy results from B-F1RST, a prospective phase II trial evaluating blood-based tumour mutational burden (bTMB) as a predictive biomarker for atezolizumab (atezo) in 1L non-small cell lung cancer (NSCLC)

BackgroundbTMB assays determine TMB using a noninvasive blood test. B-F1RST (ITT, n=152) is the first prospective trial to evaluate bTMB as a biomarker to predict benefit of 1L atezo monotherapy in advanced NSCLC. bTMB high (score of≥16; ≥ 14.5 mut/Mb) predicted better ORR with atezo vs bTMB low (< 16; 28.6% vs 4.4%) in the biomarker evaluable population (BEP) with ≥ 6mo follow up in the primary analysis. Numerical benefit for bTMB high was seen in median (m)PFS and mOS. Here we report the B-F1RST final analysis. MethodsEligibility criteria included untreated stage IIIB-IVB NSCLC and ECOG PS 0/1. Pts received atezo 1200mg IV q3w until PD, intolerance or loss of benefit. Co-primary endpoints were investigator assessed ORR for efficacy (ITT) and PFS for biomarker analysis (BEP) at a prespecified bTMB cutoff of 16 for high (≥ 16) vs low (< 16). PFS and OS, a secondary endpoint, were further evaluated at various bTMB cutoffs. Serum C-reactive protein (CRP), an inflammation marker in cancer, was evaluated as a surrogate biomarker, ratio of CRP at C3D1 to CRP at screening, to predict PFS and OS. ResultsWith ≥ 18mo follow up (data cutoff, 14 May 2019) in ITT pts, ORR was 17% (95% CI: 12, 24), mPFS was 4.1mo (95% CI: 2.8, 4.9) and mOS was 14.8mo (95% CI: 12.7, 21.3). In bTMB ≥ 16 vs<16, mPFS was 5.0 vs 3.5mo and mOS was 23.9 vs 13.4mo (Table). For CRP ratio < 0.5 vs≥0.5, mPFS was 14.1 vs 4.6mo (HR, 0.43 [90% CI: 0.24, 0.77]), and mOS was NE vs 15.9mo (HR, 0.30 [90% CI: 0.13, 0.72]). 14% of pts had treatment-related (TR) serious AEs, and 20% had Gr 3-4 TRAEs. 18% of pts had AEs that led to discontinuation. ConclusionsB-F1RST shows the clinical utility of bTMB as a predictive biomarker for pts receiving 1L atezo monotherapy. The final analysis confirmed that pts with bTMB ≥ 16 had numerical benefit for PFS and OS. Decrease in serum CRP over 6 wk predicted PFS and OS benefit. No new safety signals were seen.TableLBA83TablePFS resultsMedian (95% CI), moITT (N=152)4.1 (2.8, 4.9)BEP (n=119)bTMB High (≥ bTMB cutoff)bTMB Low (< bTMB cutoff)HR90% CIbTMB cutoffMedian, mo (n)Median, mo (n)122.6 (44)4.1 (75)1.160.83, 1.64165.0 (28)3.5 (91)0.800.54, 1.18206.9 (19)2.9 (100)0.590.37, 0.93OS resultsMedian (95% CI), moITT (N=152)14.8 (12.7, 21.3)BEP (n=119)bTMB High (≥ bTMB cutoff)bTMB Low (< bTMB cutoff)HR90% CIbTMB cutoffMedian, mo (n)Median, mo (n)1215.7 (44)13.8 (75)0.950.62, 1.461623.9 (28)13.4 (91)0.660.40, 1.102023.9 (19)13.1(100)0.440.23, 0.85 Clinical trial identificationNCT02848651. Editorial acknowledgementMedical writing assistance was provided by Chris Lum, PhD, of Health Interactions, and funded by F. Hoffmann-La Roche, Ltd. Legal entity responsible for the studyF. Hoffmann-La Roche, Ltd. FundingF. Hoffmann-La Roche, Ltd. DisclosureM. Socinski: Research grant / Funding (institution), Non-remunerated activity/ies: F. Hoffmann-La Roche. V. Velcheti: Advisory / Consultancy, Speaker Bureau / Expert testimony: Genentech, Inc.; Advisory / Consultancy, Speaker Bureau / Expert testimony: BMS; Advisory / Consultancy, Speaker Bureau / Expert testimony: AstraZeneca; Advisory / Consultancy, Speaker Bureau / Expert testimony: Takeda; Advisory / Consultancy, Speaker Bureau / Expert testimony: Foundation Medicine. T. Mekhail: Speaker Bureau / Expert testimony: Roche/Genentech. Y.K. Chae: Advisory / Consultancy: Foundation Medicine; Advisory / Consultancy: Guardant Health; Advisory / Consultancy: Biodesix; Honoraria (self): F. Hoffmann La Roche; Honoraria (self): AstraZeneca; Honoraria (self): Merck; Research grant / Funding (institution): BMS. T.A. Leal: Advisory / Consultancy: Genentech, Inc.; Advisory / Consultancy: AstraZeneca; Advisory / Consultancy: BMS; Advisory / Consultancy: Bayer; Travel / Accommodation / Expenses, Non-remunerated activity/ies: Mirati Therapeutics. J.E. Dowell: Research grant / Funding (institution): Genentech; Research grant / Funding (institution): Trizell. V. Shen: Travel / Accommodation / Expenses, Shareholder / Stockholder / Stock options, Full / Part-time employment: Roche/Genentech. S. Hu: Full / Part-time employment: Genentech, Inc.. S.M. Paul: Full / Part-time employment: Genentech, Inc.. D.S. Shames: Full / Part-time employment: Genentech, Inc.. E. Schleifman: Full / Part-time employment: Genentech, Inc.. D.A. Fabrizio: Shareholder / Stockholder / Stock options, Full / Part-time employment: Foundation Medicine, Inc. M. Nowicki: Shareholder / Stockholder / Stock options, Full / Part-time employment: F. Hoffmann-La Roche. C. Yun: Full / Part-time employment: Genentech, Inc.. S. Phan: Full / Part-time employment: Genentech, Inc.. E.S. Kim: Honoraria (self), Advisory / Consultancy, Travel / Accommodation / Expenses: Roche/Genentech; Honoraria (self), Advisory / Consultancy, Travel / Accommodation / Expenses: AstraZeneca; Honoraria (self), Advisory / Consultancy, Travel / Accommodation / Expenses: Pfizer; Honoraria (self), Advisory / Consultancy, Travel / Accommodation / Expenses: Merck; Honoraria (self), Advisory / Consultancy, Travel / Accommodation / Expenses: Boehringer Ingelheim; Honoraria (self), Advisory / Consultancy, Travel / Accommodation / Expenses: Takeda. All other authors have declared no conflicts of interest.