Diagnostic Pathology Research

Virology Testing

The core mission of Montefiore Einstein’s Diagnostic Virology Laboratory is to provide state-of-the-art routine and complex diagnostic Virology testing to the Montefiore network. Under the leadership of  Amy S. Fox, MD, MS, Director, and D. Yitzchak Goldstein, MD, Assistant Director, the Diagnostic Virology Laboratory has been transformed into one of most comprehensive molecular virology diagnostic laboratories on the East Coast, providing both diagnostic services for Montefiore patients and consultation to Montefiore physicians.

In addition, the laboratory engages in clinical and collaborative research with physician colleagues as well as industry partners. For over twenty years, the main focus of this research has been on improving diagnostics based on clinical outcomes. Using databases such as Clinical Looking Glass and others, the laboratory has repeatedly set the standard for diagnostic testing and algorithms; with industry partners, it has become a critical resource for clinical trials to test new methodologies and emerging technologies. Current areas of interest include: Hepatitis viruses with respect to streamlining clinical algorithms; Human Papilloma virus with respect to improving diagnostics and reexamination of current diagnostic algorithms; and Respiratory viral testing from a clinical utility and outcomes perspective.

The laboratory is always looking for meaningful ways to expand its portfolio, with an eye towards impact and cost effectiveness. Dr. Fox and her team welcome collaborations with both clinical and basic science colleagues.
 

Point of Care Testing

The Division of Point of Care (POC) Testing and Outreach Laboratories, directed by Amy S. Fox, MD, MS, is responsible for all near-patient testing performed throughout the Montefiore Network.

Available assays include both waived and non-waived testing at over 300 patient locations. Testing is performed by staff trained and deemed competent through a certification process overseen by the Division. All regulatory aspects of testing (Quality Control, Quality Assurance, Proficiency) are overseen by Dr. Fox and her team. Fifteen different tests are currently included in the test menu, ranging in complexity from waived urinalysis on instrumentation to non-waived coagulation testing in the Operating Room. The Point of Care team trains and certifies more than 5,000 licensed employees annually who are eligible to perform on-site testing.

In collaboration with clinical colleagues, the POC team has been instrumental in the evaluation of new Point of Care tests for the FDA and in assessing the capabilities of new methodologies in early phases of development for industry partners. In addition, current platforms have been evaluated for their accuracy, clinical outcomes and cost effectiveness.

The integration of Point of Care testing into a diverse clinical network is of particular research interest. The Montefiore network includes hospitals, clinics, practices, urgent care centers and schools, providing a diverse landscape for clinical research into optimization of healthcare delivery. Collaborations with clinical colleagues, basic scientists and industry are welcome.
 

Diagnostic Testing for Bleeding and Thrombotic Disorders

The Clinical Hematology/Coagulation Laboratory at Montefiore Medical Center (MMC), directed by Morayma Reyes Gil, MD, PhD, performs over one million clinical tests annually. These include CBCs, routine coagulation tests and highly complex, state-of-the-art tests to diagnose bleeding and thrombotic disorders. Dr. Reyes Gil is a physician-scientist experienced in basic, translational and clinical research in hematology and coagulation. She validates clinical assays to diagnose bleeding and thrombotic disorders, as well as monitoring of new anticoagulation therapies.

Current research areas include:

• Sickle cell disease (SCD): State-of-the-art tests to diagnose SCD and coinheritance with other hemoglobinopathies and thalassemias. Dr. Reyes Gil is developing a flow cytometry assay to measure expression of Hemoglobin F capable of differentiating pancellular (hereditary persistence of hemoglobin F) versus heterocellular (β thalassemia), which will improve classification of the SCD phenotype and may help predict risk of sickle cell crisis and complications. The Reyes Gil laboratory is also studying the mechanisms of hypercoagulability in SCD, namely acquired protein S deficiency in SCD.
• Platelet functional assays: State-of-the-art testing for diagnosis of platelet disorders and testing of new platelet activator and platelet inhibitor agents. Dr. Reyes Gil is developing new flow cytometry assays and whole blood assays to study platelet activation and their interaction with red and white blood cells.
• Coagulation factor assays: Daily testing for patients with bleeding conditions served at the Montefiore Hemophilia Treatment Center. Dr. Reyes Gil is developing new chromogenic factor assays to reliably monitor hemophilia patients treated with new drugs that cannot be measured with traditional clottable factor assays. The Coagulation laboratory performs a comprehensive panel for Von Willebrand Disease (VWD), the most common hereditary bleeding disorder, including an in-house VWF multimers assay. In collaboration with the MMC Molecular Laboratory, Dr. Reyes Gil is validating a VWF genetic sequencing assay to confirm most cases of VWD type II.
• Thrombophilia testing and anticoagulant monitoring testing for patients seen in the Einstein-Montefiore Thrombosis Prevention and Treatment Program: Dr. Reyes Gil is validating assays to monitor new oral anticoagulants (rivaroxaban, apixaban, edoxaban and dabigatran) and their antidotes. She is developing clinical assays to identify and measure neutrophil extracellular traps which are highly associated with infection, inflammation and thrombosis.

Next Generation Diagnosis of Infectious Disease

The Clinical Microbiology Laboratory at Montefiore, under the leadership of Wendy Szymczak, PhD, interim director, is working with the research laboratory of Dr. William Jacobs and several Montefiore gastroenterologists to implement next-generation, whole-genome sequencing for pathogen detection, identification, and antimicrobial susceptibility prediction. Sequencing of DNA extracted from clinical specimens has particular utility for the detection of pathogens not recoverable by culture or slow-growing organisms since diagnosis and determination of optimal antimicrobial therapy may not otherwise be possible or significantly delayed. Whole-genome sequencing offers advantages over targeted, PCR-based assays because a broader range of pathogens can be detected and a comprehensive analysis of genotypic resistance and virulence characteristics simultaneously performed.

The investigators are exploring the utility of sequencing for predicting the antimicrobial susceptibility of Helicobacter pylori, a gastric pathogen that can cause chronic gastritis, ulceration, or cancer. This is important because therapy is needed for symptom relief and because eradication of H. pylori has been shown to be effective in the prevention of primary gastric cancer. Susceptibility testing of this organism is necessary because H. pylori antimicrobial resistance is increasing, and eradication is improved by susceptibility-guided therapy. H. pylori is a fastidious pathogen that is difficult to isolate and sustain in culture, and the lack of phenotypic susceptibility testing guidelines and interpretation standards for most antibiotics used to treat H. pylori highlight a need for the development of new methods that accurately determine susceptibility. Future projects will also assess the feasibility of H. pylori detection and antimicrobial susceptibility determination directly from gastric-tissue specimens.

Mass Spectrometry: Amyloid Diagnostic Tests and Other Applications

Mass spectrometry (MS) is an analytical technique that accurately measures the mass-to-charge ratio of different molecules within a sample. MS is becoming an increasingly important tool in biomedical research. Clinician-scientists in the Montefiore Einstein Pathology Department are currently involved in cross-campus collaborations focused on the research and development of mass spectrometry applications in Clinical Pathology.

A major research highlight is the development of a diagnostic test for Amyloidosis sub-typing by mass spectrometry. This project is a collaborative effort between the Pathology Department, the Laboratory for Macromolecular Analysis and Proteomics, and the Weiler Clinical Mass Spectrometry Laboratory. Amyloidosis is a rare disease caused by abnormal deposition of misfolded proteins which forms “amyloid” fibrils that can accumulate in different organs and can lead to organ failure or malfunction. The correct identification of the amyloid protein subtype is critical in determining the type of treatment for this disease. There are about 30 different sub-types, depending on the specific amyloid protein, that can be successfully identified and diagnosed by mass spectrometry. The group invites inquiries from other investigators interested in potential collaborative projects aimed at the translation of basic biomarker research into diagnostic tests that can be developed and implemented at Einstein-Montefiore’s mass spectrometry laboratories. Contact Dr. Jennifer Aguilan at: jennifer.aguilan@einsteinmed.org

Clinical Cytogenetics and CytoGenomics

Started as a small laboratory in the early 1990s, the Clinical Cytogenetics and CytoGenomics Laboratory at Montefiore Einstein has grown to become a specialized, state-of-the-art Molecular Cytogenetics and CytoGenomics facilty directed by K. H. Ramesh, PhD, ABMGG, FACMGG, a board certified Cytogeneticist. It was among the first laboratories in the United States approved (by the New York State Department of Health) to offer Fluorescence in Situ Hybridization Analysis (FISH) for HER2 Amplification in Breast Cancer in 2000; for CytoGenomic Arrays in 2008, and for the rarer Cholangiocarcinoma in 2010. The lab continually develops new molecular tests, the most recent of which is ROS1 FISH testing for lung cancer as mandated by the National Comprehensive Cancer Network.

The laboratory’s technical and professional experience in diagnosing and reporting the most complex Cytogenetics and CytoGenomics results is unparalleled. The expertise and the wide array of Molecular Cytogenetics tests offered have drawn Institutions such as MD Anderson Cancer Center’s School of Health Professions to send their undergraduate students for out-of-state training here. Research interests encompass diverse malignant diseases, and the lab provides support for many Montefiore oncologists. Most notably, Dr. Ramesh and his team have collaborated with Dr. Amit Verma to confirm MDM2 (12q15), MDM4 (1q32.1) and PAK1 (11q13.5-11q14.1) mutations that indirectly impair the function of p53 in MDS, AML and ATLL by a second method such as FISH Analysis. Pilot studies have shown that mutations and amplifications (detected by FISH) of MDM4 and MDM2 indirectly impair the normal functioning of p53 in AML and ATLL. Treating patients harboring such mutations with a novel stapled peptide molecule would inhibit MDM4 and MDM2, thereby restoring the normal functions of p53.

Dr. Ramesh is also currently working on an IRB-approved project to establish the clinical significance and implications of HER2 Genetic Heterogeneity (GH) in Invasive Ductal Carcinoma (IDC) of the breast. FISH analysis data are being collected and correlated with patient demographics, histological tumor type and stage, IHC results, treatment regimen and morbidity to determine if HER2 –GH has an impact on treatment modality. The goal of this research is to standardize the accuracy and consistency of interpreting GH in breast tumors, and to determine whether a systematic study of outcomes among patients with HER2-GH will contribute toward reconsiderations of HER2 reporting guidelines as mandated by ASCO/CAP (2018). Dr. Ramesh believes that FISH analysis results that reveal a high percentage of HER2-GH in IDC of the breast should be considered in the reporting guideline, as a small percentage (~8%) of patients could benefit from Kinase Inhibitor Therapy, thereby increasing their chance of survival (unpublished data).

The laboratory’s ongoing mission is to continue to develop new tests and create comprehensive testing algorithms to bridge the gap(s) between Molecular Cytogenetics, Microarray Analysis and Testing by Next Generation Sequencing.

Molecular Genetic Pathology

The Molecular Genetic Pathology Laboratory at Montefiore, directed by D. Yitzchak Goldstein, MD is engaged in a wide variety of research endeavors, including the development of a high throughput screening methodology for Spinal Muscular Atrophy (SMA). SMA is an autosomal recessive neurodegenerative disease that occurs in children who have two deleted or mutated copies of the functional SMN1 gene.

Many conventional screening methods may leave diverse populations at increased risk since they simply assess gene copy number. Montefiore is developing additional advanced diagnostic assays to enhance screening sensitivity and to ensure providers are given all necessary information to advise their patients.

Additionally, the laboratory is investigating the nature of various T-cell receptor gene rearrangements to aid in the clinical prognostication and risk stratification of patients with Mycosis Fungoides.

Comparative Pathology: Enhancing Basic and Translational Research

Comparative Pathology data analysis provides critical information required for the validation and characterization of animal models. Amanda Beck, DVM, Scientific Director of Einstein’s Histology and Comparative Pathology Facility, is a board-certified veterinary comparative pathologist with extensive experience in the development and evaluation of non-human primate and mouse models of cancer and pre-neoplasia, infectious disease, and aging. Dr. Beck interfaces with Einstein/Montefiore researchers on a variety of basic science and translational research projects. These include, for example, models of pancreatic, intestinal, liver, breast, skeletal muscle, lung, hematopoietic, endocrine and bone cancer. She advises investigators on the influence of mouse genetic background/strain on phenotypic expression and study outcomes, and on tissue collection, preparation and fixation during the planning stages of studies.

Dr. Beck also closely interacts with Einstein’s Institute for Animal Studies, which provides vivarium housing and colony healthcare for all experimental animals in the College of Medicine’s Animal Barrier facility. She provides diagnostic veterinary pathology support to the laboratory animal veterinarians and veterinary technicians, which is particularly important when an investigator experiences unexpected illness or death in their research animals. This allows the veterinary staff to monitor for potential outbreaks of infectious disease and to detect any potential husbandry or experimental issues to be rectified.

Super Resolution Microscopy: Advancing Diagnostic Renal Pathology

The laboratory of James Pullman, MD, PhD, is exploring the use of a new method to diagnose glomerular disease, a complex area of renal pathology that requires more than the routine light microscopy (LM) that suffices for diagnosis in other organs.

Glomerular disease pathology uses electron microscopy (EM) to image structures too small for LM to resolve, as well as immunofluorescence microscopy (IFM) to identify and localize specific proteins. Super Resolution Microscopy (SRM) is a group of new technologies that combines the molecular mapping capabilities of IFM with the resolving power of EM, and can provide information similar to and likely beyond what either can offer.

Dr. Pullman is working with a team of physicists from the University of St. Andrews in Scotland, led by Professor Kishan Dholakia. They have employed Structured Illumination Microscopy (SIM), a type of SRM with the advantage of using specimen preparations similar to those of conventional IFM. Initially, the investigators examined the pathology of the podocyte, one of three cell types in the glomerulus. The podocyte is distinguished by its “foot processes,” unusual cytoplasmic extensions which retract into the cell body in diseases associated with proteinuria, or nephrotic syndrome.

Until now, EM, and not LM or IFM, provided the only way to visualize foot processes. The Pullman Laboratory has shown that SIM, with an immunofluorescent-labeled antibody bound to the protein podocin, imaged the outlines of foot processes in normal and diseased podocytes, and matched the changes seen by EM. In addition, the large field of view provided by SIM, an order of magnitude greater than that of EM, revealed novel changes in foot processes that could not be seen with EM.

Dr. Pullman and his colleagues are further analyzing these changes to understand their significance in nephrotic disease. In addition to podocyte pathology, they’re using SIM to map disease-related changes in other glomerular proteins, cells and extracellular matrices, as well as molecules that infiltrate the glomerulus and cause disease, such as immune complexes.