Diagnostic Performance Improvement with Combined use of Proteomics Biomarker Assay and Breast Ultrasound

Purpose: To investigate the combined use of blood-based 3-protein signature and breast ultrasound (US) for validating US detected lesions.Methods: From July 2011 to April 2020, women who underwent whole-breast US within at least 6 months from sampling period were retrospectively included. Blood-based 3-protein signature (Mastocheck®) value and US findings were evaluated. Following outcome measures were compared between US alone and the combination of Mastocheck® value with US: sensitivity, specificity, positive predictive value (PPV), negative predictive value (NPV), area under the receiver operating characteristic curve (AUC), and biopsy rate. Results: Among the 237 women included, 59 (24.9%) were healthy individuals and 178 (75.1%) cancer patients. Mean size of cancers was 1.2±0.8 cm. Median value of Mastocheck® was significantly different between non-malignant (-0.24, interquartile range [IQR], -0.48, -0.03) and malignant lesions (0.55, IQR, -0.03, 1.42) (P < .001). Utilizing Mastocheck® value with US increased the AUC from 0.67 (95% confidence interval [CI], 0.61, 0.73) to 0.81 (95% CI: 0.75, 0.88; P < .001), specificity from 35.6% (95% CI: 23.4, 47.8) to 64.4% (95% CI: 52.2, 76.6; P < .001) without loss in sensitivity. PPV was increased from 82.2% (95% CI: 77.1, 87.3) to 89.3% (95% CI: 85.0, 93.6; P < .001), and biopsy rate was significantly decreased from 79.3% (188/237) to 72.1% (171/237) (P < .001). Consistent improvements in specificity, PPV, and AUC were observed in asymptomatic women and in those with normal/benign mammographic findings. Conclusion: Mastocheck® is an effective tool that can be used with US to improve diagnostic specificity and reduce false-positive findings and unnecessary biopsies.

112. A Rapid Host-Protein Signature Based on TNF-related Apoptosis-Induced Ligand (TRAIL), Interferon Gamma Induced Protein-10 (IP-10) and C-Reactive Protein (CRP) Accurately Differentiates Between Bacterial and Viral Infection in Febrile Children: Apollo Sub-Study

Background Identifying infectious etiology is essential for appropriate patient management, including antibiotic use. A host-protein signature for differentiating bacterial from viral infection has exhibited robust performance (AUC of 0.9, 95% CI 0.86-0.95) in prior studies. Performance data was lacking for a broad pediatric population recruited in emergency departments (EDs) and urgent care centers (UCCs). Methods Non-immunocompromised children were recruited prospectively from 5 EDs and 3 UCCs in the U.S. and 1 ED in Israel between May 2019 and August 2020. Eligibility required physician’s clinical suspicion of acute infection and reported fever. Reference standard etiology was adjudicated by experts based on clinical, laboratory, radiological, microbiological and follow-up data. For the primary analysis, experts blinded to one another, to the host-signature results and also to procalcitonin and CRP, classified cases as bacterial or viral. For the secondary analysis, experts blinded to one another and the host signature results, were permitted to classify cases as bacterial, viral or indeterminate; indeterminates were removed from the secondary analysis. Host signature (comprising TRAIL, IP-10 and CRP; MeMed BV®) was measured using a rapid platform (MeMed Key®) generating a bacterial likelihood score (0-100) in 15 minutes. Results The study cohort comprised 162 children (median age, 5.5 yrs; interquartile range, 8.5), of whom 69 (43%) presented within 2 days of symptom onset and 37 (23%) were hospitalized for a median of 3 days. Respiratory tract infection was the predominant syndrome (11% lower and 44% upper). Host signature attained AUC 0.87 (0.74-1) and 0.92 (0.79-1) in the primary and secondary analysis, respectively. With higher the signature score, there was a significantly higher likelihood of bacterial infection (p&lt; 0.001; Table 1). The 3 bacterial infections assigned score &lt; 35 (false negative) would have been identifiable by physical examination (Table 2). Increasing host signature score is associated with increasing likelihood of bacterial infection across both the primary and secondary cohort The performance of the host signature score in differentiating between bacterial and viral infection was evaluated by allocating children to one of five score bins and within each bin according to their adjudication label and determining if there is a meaningful increase in the relative likelihood of bacterial infection across the bins based on the Cochrane-Armitage test of trend. PPV, positive predictive value. NPV, negative predictive value. *Includes patients adjudicated as non-infectious Three children assigned a bacterial adjudication label and a score of 35 or less (false negatives) have bacterial infections identifiable in physical exam Conclusion The host-protein signature measured using a rapid platform attained robust performance in differentiating bacterial vs viral infection in children with acute febrile illness, supporting its potential to enhance rational use of antibiotics in the ED and UCC. Disclosures Sheldon L. Kaplan, MD, Pfizer (Research Grant or Support) Mark Kellogg, PhD, MeMed (Scientific Research Study Investigator) Andrea T. Cruz, MD, MPH, American Academy of Pediatrics (Individual(s) Involved: Self): editorial board member Kristina G. Hulten, PhD, Pfizer (Research Grant or Support) Cesar A. Arias, M.D., MSc, Ph.D., FIDSA, Entasis Therapeutics (Grant/Research Support)MeMed Diagnostics (Grant/Research Support)Merk (Grant/Research Support) Richard Gordon, MD, MeMed (Scientific Research Study Investigator) Sergey Motov, MD, MeMed (Scientific Research Study Investigator) Theresa Jacob, PHD MPH, MeMed (Scientific Research Study Investigator) Natasha Ballard, MD, MeMed (Scientific Research Study Investigator) George Suits, MD, MeMed (Scientific Research Study Investigator) Jeffrey Harris, MD, MeMed (Scientific Research Study Investigator) Maanit Shapira, Ph.D, MeMed (Scientific Research Study Investigator) Richard E. Rothman, PhD, MD, Chem bio (Grant/Research Support) Karen C. Carroll, MD, MeMed (Scientific Research Study Investigator)Meridian Diagnostics, Inc. (Grant/Research Support)Pattern Diagnostics (Advisor or Review Panel member)Scanogen, Inc. (Advisor or Review Panel member) Karen C. Carroll, MD, Pattern Diagnostics, Inc. (Individual(s) Involved: Self): Grant/Research Support; Scanogen, Inc. (Individual(s) Involved: Self): Consultant Leticia M. Ryan, MD MPH, MeMed (Scientific Research Study Investigator) Richard Bachur, MD, MeMed (Scientific Research Study Investigator)

Detection of soluble biomarkers of pancreatic cancer in endoscopic ultrasound-guided fine-needle aspiration samples

Background Biomarkers are urgently needed for pancreatic ductal adenocarcinoma (PDAC). Endoscopic ultrasound-guided fine-needle aspiration (EUS-FNA) is the cornerstone for diagnosing PDAC. We developed a method for discovery of PDAC biomarkers using the discarded EUS-FNA liquid. Methods This retrospective study included 58 patients with suspected pancreatic lesions who underwent EUS-FNA. Protein extracts from EUS-FNA liquid were analyzed by mass spectrometry. Proteomic and clinical data were modeled by supervised statistical learning to identify protein markers and clinical variables that distinguish PDAC. Results Statistical modeling revealed a protein signature for PDAC screening that achieved high sensitivity and specificity (0.92, 95 % confidence interval [CI] 0.79–0.98, and 0.85, 95 %CI 0.67–0.93, respectively). We also developed a protein signature score (PSS) to guide PDAC diagnosis. In combination with patient age, the PSS achieved 100 % certainty in correctly identifying PDAC patients > 54 years. In addition, 3 /4 inconclusive EUS-FNA biopsies were correctly identified using PSS. Conclusions EUS-FNA-derived fluid is a rich source of PDAC proteins with biomarker potential. The PSS requires further validation and verification of the feasibility of measuring these proteins in patient sera.

Proteome profiling of human placenta reveals developmental stage-dependent alterations in protein signature

Introduction Placenta is a complex organ that plays a significant role in the maintenance of pregnancy health. It is a dynamic organ that undergoes dramatic changes in growth and development at different stages of gestation. In the first-trimester, the conceptus develops in a low oxygen environment that favors organogenesis in the embryo and cell proliferation and angiogenesis in the placenta; later in pregnancy, higher oxygen concentration is required to support the rapid growth of the fetus. This oxygen transition, which appears unique to the human placenta, must be finely tuned through successive rounds of protein signature alterations. This study compares placental proteome in normal first-trimester (FT) and term human placentas (TP). Methods Normal human first-trimester and term placental samples were collected and differentially expressed proteins were identified using two-dimensional liquid chromatography-tandem mass spectrometry. Results Despite the overall similarities, 120 proteins were differently expressed in first and term placentas. Out of these, 72 were up-regulated and 48 were down-regulated in the first when compared with the full term placentas. Twenty out of 120 differently expressed proteins were sequenced, among them seven showed increased (GRP78, PDIA3, ENOA, ECH1, PRDX4, ERP29, ECHM), eleven decreased (TRFE, ALBU, K2C1, ACTG, CSH2, PRDX2, FABP5, HBG1, FABP4, K2C8, K1C9) expression in first-trimester compared to the full-term placentas and two proteins exclusively expressed in first-trimester placentas (MESD, MYDGF). Conclusion According to Reactome and PANTHER softwares, these proteins were mostly involved in response to chemical stimulus and stress, regulation of biological quality, programmed cell death, hemostatic and catabolic processes, protein folding, cellular oxidant detoxification, coagulation and retina homeostasis. Elucidation of alteration in protein signature during placental development would provide researchers with a better understanding of the critical biological processes of placentogenesis and delineate proteins involved in regulation of placental function during development.

Anti-CD20 Therapy Alters the Protein Signature in Experimental Murine AIH, but Not Exclusively Towards Regeneration

Background: Autoimmune hepatitis (AIH) is a chronic autoimmune inflammatory disease that usually requires lifelong immunosuppression. Frequent recurrences after the discontinuation of therapy indicate that intrahepatic immune regulation is not restored by current treatments. Studies of other autoimmune diseases suggest that temporary depletion of B cells can improve disease progression in the long term. Methods: We tested a single administration of anti-CD20 antibodies to reduce B cells and the amount of IgG to induce intrahepatic immune tolerance. We used our experimental murine AIH (emAIH) model and treated the mice with anti-CD20 during the late stage of the disease. Results: After treatment, the mice showed the expected reductions in B cells and serum IgGs, but no improvements in pathology. However, all treated animals showed a highly altered serum protein expression pattern, which was a balance between inflammation and regeneration. Conclusions: In conclusion, anti-CD20 therapy did not produce clinically measurable results because it triggered inflammation, as well as regeneration, at the proteomic level. This finding suggests that anti-CD20 is ineffective as a sole treatment for AIH or emAIH.

Influencing Factors on the Oncuria™ Urinalysis Assay: An Experimental Model

Background: The Oncuria™ urine test for the detection of bladder cancer measures a multiplex protein signature. In this study, we investigated the influence of urinary cellularity, protein, and hematuria on the performance of the Oncuria™ test in an ex vivo experimental model. Materials and Methods: Pooled urine from healthy subjects was spiked with cultured benign (UROtsa) or malignant cells (T24), cellular proteins, or whole blood. The resulting samples were analyzed using the Oncuria™ test following the manufacturer’s instructions. Results: Urine samples obtained from healthy subjects were negative for bladder cancer by Oncuria™ test criteria. The majority of the manipulated conditions did not result in a false-positive test. The addition of whole blood (high concentration) did result in a false-positive result, but this was abrogated by sample centrifugation prior to analysis. The addition of cellular proteins (high concentration) resulted in a positive Oncuria™ test, and this was unaffected by pre-analysis sample centrifugation. Conclusions: The Oncuria™ multiplex test performed well in the ex vivo experimental model and shows promise for clinical application. The identification of patients who require additional clinical evaluation could reduce the need to subject patients who do not have bladder cancer to frequent, uncomfortable and expensive cystoscopic examinations, thus benefiting both patients and the healthcare system.