How New Regulation of Laboratory-Developed Antimicrobial Susceptibility Tests Will Affect Infectious Diseases Clinical Practice.

Antimicrobial resistance (AMR) affects 2.8 million Americans annually. AMR is identified through antimicrobial susceptibility testing (AST), but current and proposed regulatory policies from the United States Food and Drug Administration (FDA) jeopardize the future availability of AST for many microorganisms. Devices that perform AST must be cleared by the FDA using their susceptibility test interpretive criteria, also known as breakpoints. The FDA list of breakpoints is relatively short. Today, laboratories supplement FDA breakpoints using breakpoints published by the Clinical and Laboratory Standards Institute, using legacy devices and laboratory-developed tests (LDTs). FDA proposes to regulate LDTs, and with no FDA breakpoints for many drug-bug combinations, the risk is loss of AST for key clinical indications and stifling innovation in technology development. Effective solutions require collaboration between manufacturers, infectious diseases clinicians, pharmacists, laboratories, and the FDA.

The gaps between the new EU legislation on in vitro diagnostics and the on-the-ground reality.

The background to this debate is now well-known: an EU policy decision to tighten controls on the devices and diagnostics sector led to the adoption of a regulation in 2017 with a schedule for implementation over coming years - a timetable extended still further by last-minute legislation in early 2022, to provide the sector and regulators with more time to adapt to the changes. Discussions among experts organised in April by the European Alliance for Personalized Medicine (EAPM) exposed continuing challenges that cannot be fully resolved by the recent deferral of implementation deadlines. One salient problem is that there is little awareness of the In Vitro Diagnostic Regulation (IVDR) across Europe, and only limited awareness of the different structures of national systems involved in implementing IVDR, with consequent risks for patient and consumer access to in vitro diagnostics (IVDs). The tentative conclusion from these consultations is that despite a will across the sector to seek workable solutions, the obstacles remain formidable, and the potential solutions so far proposed remain more a matter of aspirations than of clear pathways.

ISO 15189 is a sufficient instrument to guarantee high-quality manufacture of laboratory developed tests for in-house-use conform requirements of the European In-Vitro-Diagnostics Regulation.

The EU In-Vitro Diagnostic Device Regulation (IVDR) aims for transparent risk-and purpose-based validation of diagnostic devices, traceability of results to uniquely identified devices, and post-market surveillance. The IVDR regulates design, manufacture and putting into use of devices, but not medical services using these devices. In the absence of suitable commercial devices, the laboratory can resort to laboratory-developed tests (LDT) for in-house use. Documentary obligations (IVDR Art 5.5), the performance and safety specifications of ANNEX I, and development and manufacture under an ISO 15189-equivalent quality system apply. LDTs serve specific clinical needs, often for low volume niche applications, or correspond to the translational phase of new tests and treatments, often extremely relevant for patient care. As some commercial tests may disappear with the IVDR roll-out, many will require urgent LDT replacement. The workload will also depend on which modifications to commercial tests turns them into an LDT, and on how national legislators and competent authorities (CA) will handle new competences and responsibilities. We discuss appropriate interpretation of ISO 15189 to cover IVDR requirements. Selected cases illustrate LDT implementation covering medical needs with commensurate management of risk emanating from intended use and/or design of devices. Unintended collateral damage of the IVDR comprises loss of non-profitable niche applications, increases of costs and wasted resources, and migration of innovative research to more cost-efficient environments. Taking into account local specifics, the legislative framework should reduce the burden on and associated opportunity costs for the health care system, by making diligent use of existing frameworks.

[Study on Clinical Application and Supervision of LC-MS in in Vitro Diagnosis Test].

From the perspective of technical evaluation, this study reviewed the current situation of application and clinical application of medical device products were detected by liquid chromatography-tandem mass spectrometry in the market in recent years. The regulatory requirements of these products in China, USA, EU and Japan were compared and analyzed, and the monitoring situation of adverse events after listing, the standards for reference and the domestic and foreign regulatory documents were combined, the clinical application and regulatory risks of the product were analyzed. The problems such as pre-treatment, system matching, adequacy of performance index requirements, inter-room consistency, reference interval and registration unit were discussed and suggestions for supervision were given, with a view to the field of product R&D and production, review and approval of supervision to provide technical reference.

[Ideas and Briefing about Regulatory Requirements for Laboratory Developed Tests in the US].

As a special kind of in vitro diagnostic devices（IVDs）, laboratory developed tests（LDTs） are of great significance to the development of clinical laboratories. This study aims to explore the regulatory requirements ideas of LDTs. By introducing the development of LDTs and the changing of regulatory requirements in the United States, combing the current regulatory framework and discussing relevant ideas in the regulatory requirements of LDTs.

The Panther Fusion System with Open Access Functionality for Laboratory-Developed Tests for Influenza A Virus Subtyping.

Nucleic acid amplification tests, such as PCR, are the method of choice for respiratory virus testing, due to their superior diagnostic accuracy and fast turnaround time. The Panther Fusion (Fusion; Hologic) system has an array of highly sensitive in vitro diagnostic (IVD) real-time PCR assays for respiratory viruses, including an assay for influenza A (FluA) virus, influenza B (FluB) virus, and respiratory syncytial virus (RSV) (FFABR assay). The Fusion system has Open Access functionality to perform laboratory-developed tests (LDTs) alongside IVD assays. We developed two LDTs for FluA virus strain typing on the Panther Fusion instrument, enabling side-by-side testing with the FFABR assay. The LDT-FAST assay uses proprietary primers and probes designed by Hologic for the Prodesse ProFAST+ (PFAST) assay. The exWHO-FAST assay is an expanded redesign of the WHO-recommended reverse transcriptase PCRs (RT-PCRs). To evaluate the performance of these two LDTs, 110 FluA virus-positive samples were tested. Of these, 104 had been subtyped previously; 54 were H3, 46 were 09H1, and 4 were fsH1. All were appropriately subtyped by both LDTs. Of the untyped FluA virus samples, three were subtyped as H3 by both LDTs and two were subtyped as H3 by the LDT-FAST assay only. The sample not subtyped by either LDT was retested with the FFABR assay and was now negative. Limit-of-detection (LOD) analyses were performed with five FluA virus strains. The LDT-FAST LODs were similar to the FFABR assay LODs, while the exWHO-FAST LODs were higher for two H3N2 strains, findings that were explained by analysis of primer/probe homology. In conclusion, either FluA virus typing assay would be a valuable complement to the Panther Fusion respiratory menu given the performance of these LDTs, the system's full automation, and the ability to split eluates for both IVD and LDT testing.

Variant call concordance between two laboratory-developed, solid tumor targeted genomic profiling assays using distinct workflows and sequencing instruments.

Targeted genomic profiling (TGP) using massively parallel DNA sequencing is becoming the standard methodology in clinical laboratories for detecting somatic variants in solid tumors. The variety of methodologies and sequencing platforms in the marketplace for TGP has resulted in a variety of clinical TGP laboratory developed tests (LDT). The variability of LDTs is a challenge for test-to-test and laboratory-to-laboratory reliability. At the University of Vermont Medical Center (UVMMC), we validated a TGP assay for solid tumors which utilizes DNA hybridization capture and complete exon and selected intron sequencing of 29 clinically actionable genes. The validation samples were run on the Illumina MiSeq platform. Clinical specificity and sensitivity were evaluated by testing samples harboring genomic variants previously identified in CLIA-approved, CAP accredited laboratories with clinically validated molecular assays. The Molecular Laboratory at Dartmouth Hitchcock Medical Center (DHMC) provided 11 FFPE specimens that had been analyzed on AmpliSeq Cancer Hotspot Panel version 2 (CHPv2) and run on the Ion Torrent PGM. A Venn diagram of the gene lists from the two institutions is shown. This provided an excellent opportunity to compare the inter-laboratory reliability using two different target sequencing methods and sequencing platforms. Our data demonstrated an exceptionally high level of concordance with respect to the sensitivity and specificity of the analyses. All clinically-actionable SNV and InDel variant calls in genes covered by both panels (n=17) were identified by both laboratories. This data supports the proposal that distinct gene panel designs and sequencing workflows are capable of making consistent variant calls in solid tumor FFPE-derived samples.

Maintaining Life-saving Testing for Patients With Infectious Diseases: Infectious Diseases Society of America, American Society for Microbiology, and Pan American Society for Clinical Virology Recommendations on the Regulation of Laboratory-developed Tests.

In 2014, the US Food and Drug Administration (FDA) proposed to regulate laboratory-developed tests (LDTs)-diagnostics designed, manufactured, and used within a single laboratory. The Infectious Diseases Society of America, the American Society for Microbiology, and the Pan American Society for Clinical Virology recognize that the FDA is committed to protecting patients. However, our societies are concerned that the proposed regulations will limit access to testing and negatively impact infectious diseases (ID) LDTs. In this joint commentary, our societies discuss why LDTs are critical for ID patient care, hospital infection control, and public health responses. We also highlight how the FDA's proposed regulation of LDTs could impair patient access to life-saving tests and stifle innovation in ID diagnostics. Finally, our societies make specific recommendations for the FDA's consideration to reduce the burden of the proposed new rules on clinical laboratories and protect patients' access to state-of-the art, quality LDTs.

Regulating whole exome sequencing as a diagnostic test.

In the last decade, there has been a flood of new technology in the sequencing arena. The onset of next-generation sequencing (NGS) technology has resulted in the vast increase in genetic diagnostic testing available to the ordering physician. Whole exome sequencing (WES) has become available as a diagnostic test performed in certified clinical laboratories. This has led to increased presence in the diagnostic marketplace, increased consumer awareness, and the question has been raised by various stakeholders to whether there is sufficient stringent regulation of WES and other NGS-based tests. We discuss the various WES services currently available in the marketplace, current regulation of WES as a laboratory developed test, the proposed FDA involvement in its oversight as well as the response of various laboratory groups that provide these diagnostic services. Overall, a rigorous process oversight and assessment of inter-lab reproducibility is strongly warranted for WES as it is used as a diagnostic test, but regulation should be mindful of the excessive administrative burden on academic and smaller diagnostic laboratories.

Whole-Blood PCR Preferred for Timely Diagnosis of Neuroinvasive West Nile Virus Infections: Lessons From the 2021 Arizona Outbreak.

Background: In 2021, the state of Arizona experienced the largest focal outbreak of West Nile virus (WNV) in US history. Timely and accurate diagnostic testing remains a challenge for WNV due to transient viremia and limited immunoassay specificity. Recent studies have identified whole blood (WB) and urine as more sensitive specimen types for the detection of WNV RNA. Methods: We evaluated ordering practices, test performance, and patient characteristics of probable and confirmed cases. In total, we identified 190 probable and proven cases, including 127 patients (66.8%) with neuroinvasive disease. Results: Among all cases, only 29.5% had WNV polymerase chain reaction (PCR) testing ordered on WB, of which 80.3% resulted as positive, including 7 cases in which WNV serologic testing was negative and 5 cases for which serologic testing was not ordered. In comparison, only 23.7% of cases that had cerebrospinal fluid (CSF) PCR ordered had a positive result, including 3 cases that were negative by PCR on WB. In contrast, WNV PCR on WB detected 12 neuroinvasive cases that were CSF PCR negative. WNV PCR testing in urine was only ordered on 2 patients, both of whom were positive. Crossing cycle threshold (Ct) values were not significantly different between WB and CSF specimen types, nor was there a correlation between Ct value and days from symptom onset at the time of sample collection; all specimen types and time points had Ct values, with 98% above 30. WB was positive by WNV PCR in several patients for >7 days (range, 7-25 days) after symptom onset, as was the CSF PCR. Conclusions: Taken together, these findings indicate that WNV PCR testing on WB may be the best initial test for timely diagnosis of WNV infection, irrespective of clinical manifestation; however, if negative in patients with suspected neuroinvasive disease, WNV PCR testing on CSF should be ordered.

The US FDA's proposed rule on laboratory-developed tests: Impacts on clinical laboratory testing.

Objectives: To solicit quantifiable feedback from clinical laboratorians on the U.S. Food and Drug Administration (FDA) proposed rule to regulate laboratory-developed tests (LDTs) as medical devices. Design and Methods: A ten-item questionnaire was developed and submitted to clinical laboratory customers of ARUP Laboratories, a national nonprofit clinical laboratory of the University of Utah Department of Pathology. Results: Of 503 clinical laboratory respondents, only 41 (8 %) support the FDA's proposed rule. 67 % of respondents work in laboratories that perform LDTs and were therefore asked additional questions regarding the proposed rule. 84 % of these respondents believe that the proposed rule will negatively impact their laboratories, while only 3 % believe that they have the financial resources to pay for FDA user fees. 61 % of respondents anticipate removing tests from their laboratory menus if the proposed rule is enacted, while an additional 33 % indicated that they do not yet know. Only 11 % of respondents believe that they would pursue FDA submissions for all of their existing LDTs if the final rule is enacted. The vast majority of respondents (>80 %) were either 'extremely concerned' or 'very concerned' about the impact of the proposed rule on patient access to essential testing, financial and personnel resources to comply, innovation, the FDA's ability to implement the rule, and send-out costs and test prices. Conclusions: The majority of clinical laboratorians surveyed do not support the FDA's proposed rule on LDTs and report having insufficient resources to comply with the rule if it is enacted.

Patient safety and healthcare quality of U.S. laboratory developed tests (LDTs) in the AI/ML era of precision medicine.

This policy brief summarizes current U.S. regulatory considerations for ensuring patient safety and health care quality of genetic/genomic test information for precision medicine in the era of artificial intelligence/machine learning (AI/ML). The critical role of innovative and efficient laboratory developed tests (LDTs) in providing accurate diagnostic genetic/genomic information for U.S. patient- and family-centered healthcare decision-making is significant. However, many LDTs are not fully vetted for sufficient analytic and clinical validity via current FDA and CMS regulatory oversight pathways. The U.S. Centers for Disease Control and Prevention's Policy Analytical Framework Tool was used to identify the issue, perform a high-level policy analysis, and develop overview recommendations for a bipartisan healthcare policy reform strategy acceptable to diverse precision and systems medicine stakeholders.

[Regulation (EU) 2017/746 (IVDR): practical implementation of annex I in pathology]. / Die Verordnung (EU) 2017/746 (IVDR) in der Praxis: Umsetzung von Anhang I in der Pathologie.

BACKGROUND: Regulation (EU) 2017/746 on in vitro diagnostic medical devices (IVDR) imposes several conditions on pathology institutes that develop and use in-house in vitro diagnostic medical devices (IH-IVDs). However, not all of these conditions need to be implemented immediately after the IVDR entered into force on 26 May 2022. Based on an amending regulation of the European Parliament and the Council of the European Union, the requirements for IH-IVDs will be phased in. Conformity with the essential safety and performance requirements of annex I must be ensured from May 2022. OBJECTIVES: With this article, we would like to present the practical implementation of the currently valid conditions for IH-IVDs at the Institute of Pathology at the University Hospital of Heidelberg, in order to provide possible assistance to other institutions. CONCLUSIONS: In addition to the intensive work on the requirements for IH-IVDs, several guidance documents and handouts provide orientation for the implementation and harmonisation of the requirements for healthcare institutions mentioned in Article 5 (5). Exchange in academic network structures is also of great importance for the interpretation and practical implementation of the IVDR. For university and nonuniversity institutions, ensuring conformity with the IVDR represents a further challenge in terms of personnel and time, in addition to the essential tasks of patient care, teaching and research and the further development of methods for optimal and targeted diagnostics, as well as the maintenance of the constantly evolving quality management system.

Regulation (EU) 2017/746 (IVDR): practical implementation of annex I in pathology.

BACKGROUND: Regulation (EU) 2017/746 on in vitro diagnostic medical devices (IVDR) imposes several conditions on pathology departments that develop and use in-house in vitro diagnostic medical devices (IH-IVDs). However, not all of these conditions need to be implemented immediately after the IVDR entered into force on 26 May 2022. Based on an amending regulation of the European Parliament and the Council of the European Union, the requirements for IH-IVDs will be phased in. Conformity with the essential safety and performance requirements of annex I must be ensured from May 2022. OBJECTIVES: With this article, we would like to present the practical implementation of the currently valid conditions for IH-IVDs at the Institute of Pathology at the University Hospital of Heidelberg, in order to provide possible assistance to other institutions. CONCLUSIONS: In addition to the intensive work on the requirements for IH-IVDs, several guidance documents and handouts provide orientation for the implementation and harmonisation of the requirements for healthcare institutions mentioned in Article 5 (5). Exchange in academic network structures is also of great importance for the interpretation and practical implementation of the IVDR. For university and nonuniversity institutions, ensuring conformity with the IVDR represents a further challenge in terms of personnel and time, in addition to the essential tasks of patient care, teaching and research and the further development of methods for optimal and targeted diagnostics, as well as the maintenance of the constantly evolving quality management system.

Structure and content of the EU-IVDR : Current status and implications for pathology.

BACKGROUND: Regulation (EU) 2017/746 on in vitro diagnostic medical devices (IVDR) was passed by the European Parliament and the Council of the European Union on 5 April 2017 and came into force on 26 May 2017. A new amending regulation, which introduces a phased implementation of the IVDR with new transitional provisions for certain in vitro diagnostic medical devices (IVDs) and a later date of application of some requirements for in-house devices for healthcare facilities, was adopted on 15 December 2021. The combined use of CE-certified IVDs (CE-IVDs), in-house IVDs (IH-IVDs), and research use only (RUO) devices are a cornerstone of diagnostics in pathology departments and crucial for optimal patient care. The IVDR not only regulates the manufacture and placement on the market of industrially manufactured IVDs, but also imposes conditions on the manufacture and use of IH-IVDs for internal use by healthcare facilities. OBJECTIVES: Our work provides an overview of the background and structure of the IVDR and identifies core areas that need to be interpreted and fleshed out in the context of the legal framework as well as expert knowledge. CONCLUSIONS: The gaps and ambiguities in the IVDR crucially require the expertise of professional societies, alliances, and individual stakeholders to successfully facilitate the implementation and use of the IVDR in pathology departments and to avoid aberrant developments.

Laboratory-Developed Tests Account for a Small Minority of Tests Ordered in an Academic Hospital System.

OBJECTIVES: To determine the frequency of use of laboratory-developed tests (LDTs) in an academic medical center system. METHODS: Retrospective analysis of 2021 test order data from an academic medical center (hospital, outpatient clinics, and cancer center) was done. Measures included assay type, assay methodology, regulatory status, test order volume, inpatient vs outpatient setting, and provider medical specialty. RESULTS: Of the 3,016,928 tests ordered in 2021, 2,831,489 (93.9%) were tests cleared, approved, and/or authorized by the US Food and Drug Administration (FDA); 116,583 (3.9%) were LDTs; and 68,856 (2.3%) were standard methods. These test orders were performed using a total of 1,954 distinct assays. Of these, 983 (50.3%) were FDA assays, 880 (45.0%) were LDTs, and 91 (4.7%) were standard methods. Laboratory-developed tests were more commonly ordered in the outpatient vs inpatient setting and represented a higher proportion of the test volume at the cancer center compared with the university hospital (5.6% vs 3.6%, respectively). The top 167 LDT assays accounted for 90% of the LDT volume (104,996 orders). Among the 20 most frequently ordered LDTs were mass spectrometry assays and tests used in the care of immunocompromised patients. Internal/family medicine placed the greatest number of orders (1,044,642) and ordered one of the lowest proportions of LDTs (3.2%). CONCLUSIONS: Laboratory-developed tests made up a small percentage of the total laboratory tests ordered within the academic health system studied.

Advanced technologies for molecular diagnosis of cancer: State of pre-clinical tumor-derived exosome liquid biopsies.

Exosomes are membrane-defined extracellular vesicles (EVs) approximately 40-160 â€‹nm in diameter that are found in all body fluids including blood, urine, and saliva. They act as important vehicles for intercellular communication between both local and distant cells and can serve as circulating biomarkers for disease diagnosis and prognosis. Exosomes play a key role in tumor metastasis, are abundant in biofluids, and stabilize biomarkers they carry, and thus can improve cancer detection, treatment monitoring, and cancer staging/prognosis. Despite their clinical potential, lack of sensitive/specific biomarkers and sensitive isolation/enrichment and analytical technologies has posed a barrier to clinical translation of exosomes. This review presents a critical overview of technologies now being used to detect tumor-derived exosome (TDE) biomarkers in clinical specimens that have potential for clinical translation.

Comparison of two highly sensitive benzodiazepine immunoassay lab developed tests for urine drug testing in clinical specimens.

Background: The VALID Act is a legislative effort that, if enacted, would alter the regulatory requirements of laboratory developed tests (LDTs) used for clinical testing in the United States. Benzodiazepines, which are primarily excreted into urine as glucuronidated metabolites such as lorazepam, cross-react poorly with FDA-cleared immunoassays, leading to false-negatives. This shortfall can be addressed with LDTs created by adding glucuronidase to the immunoassay reagents producing "high sensitivity" assays that detect glucuronidated metabolites. Methods: Precision and stability of two high-sensitivity (HS) benzodiazepine immunoassays from Roche and Thermo Scientific were evaluated using manufacturer-supplied quality control (QC) material and glucuronidated QC material. The immunoassays were directly compared to an LC-MS/MS LDT benzodiazepine assay to determine clinical sensitivity/specificity using urine specimens (n = 82 for Thermo Scientific; n = 265 for Roche). The clinical impact of the HS LDT immunoassay was determined by analyzing clinical testing results 60 days before and after its implementation. Results: The precision and clinical sensitivity/specificity of the HS-Thermo Scientific and HS-Roche benzodiazepine assays were acceptable. The reagent stability of the HS-Thermo Scientific immunoassay was poor, whereas the HS-Roche immunoassay was stable. After implementation of the HS-Roche benzodiazepine immunoassay as an LDT, there was a 30-fold increase (p-value: < 0.00001) in the percentage of lorazepam confirmations. Conclusions: We demonstrate the development and validation of an immunoassay LDT with improved sensitivity for glucuronidated benzodiazepines. This LDT can detect glucuronidated benzodiazepines in clinical urine specimens and is stable for 60 days. Importantly, we were able to validate the immunoassay as an LDT by utilizing an LC-MS/MS LDT.