A Review of the XXXVth International Symposium on Technical Innovations in Laboratory Hematology 2023 (ISLH 2023)

Posted on: September 8, 2023

David L. McGlasson, MS, MLS(ASCP)

The most recent ISLH meeting was held in New Orleans, LA May 11-13, 2023.  The conference had many State-of-the-Art sessions and I concentrated on the hemostasis/thrombosis presentations.

On the first day, Katrien Devreese MD, PhD, Ghent University Hospital, Belgium gave an excellent presentation on the current Diagnosis of Acquired Thrombotic Thrombocytopenia Purpura (TTP).

Thrombotic thrombocytopenia purpura (TTP) is a rare thrombotic microangiopathy blood disorder characterized by the formation of blood clots in small blood vessels, thrombocytopenia, and a lack of ADAMTS13 (a disintegrin-like and metalloproteinase with thrombospondin type 1 motif 13) activity. The ADAMTS-13 protease is responsible for cleaving von Willebrand Factor (VWF) multimers into smaller fragments in a mechanism that regulates thrombosis. When ADAMTS13 activity is decreased or absent, ultra large VWF multimers (ULVWF) accumulate in the blood which activates platelet adhesion, platelet aggregation, and microvascular thrombosis. TTP-associated tissue damage can cause multiorgan failure, stroke, and death. When left untreated, TTP carries a high mortality rate of 90%.

The speaker took attendees through the early cases describing TTP in 1924 up through the ADAMTS13 discovery and its significance in diagnosing TTP. The speaker discussed the definition, pathophysiology, and prevalence of TTP. Clinical symptoms of TTP were also covered in depth to include thrombocytopenia, anemia, and location of thrombi in the body. Non-idiopathic symptoms of underlying conditions were also discussed. Laboratory testing algorithms were presented in detail. Dr. Devreese discussed progress made throughout the last decade regarding ADAMTS13 laboratory assays and their advantages. A review of pre-analytical variables, type of specimen required, specimen handling, and levels of interference of icteric and hemolyzed citrated plasma was provided. The importance of a timely ADAMTS13 testing result so that treatment via plasma exchange, possibly along with newer therapeutics, was also presented.

On the vWF front, Kristi J Smock, MD, Professor of Pathology, University of Utah School of Medicine, Medical Director of Hemostasis/Thrombosis Laboratory, ARUP Laboratories presented on Ratios in von Willebrand Disease (VWD) Testing.

Dr. Smock provided detailed information on von Willebrand factor (VWF) testing ratios in the diagnosis and subtyping of VWD. Currently there are six recognized subtypes. Inherited VWD is an autosomal dominant disorder characterized by deficiency of VWF due to mutations in the VWF genes.  Dr. Smock also described the condition as a bleeding disorder of platelet adhesion.

For further reading, see Smock KJ. Von Willebrand factor testing ratios in the diagnosis and subtyping of von Willebrand disease. Int J Lab Hematol. 2023;45 (Supp.2) 23-29.

Robert Sidonio, MD, Associate Professor of Pediatrics at Emory University Hospital, presented on VWD Diagnosis and Significance of Low VWF Levels. He discussed the issue of getting an accurate diagnosis in individuals with VWF levels between 30-50IU/dl and the problems that may occur in such diagnosis. For further reading, see Sidonio RF, Lavin M. Diagnostic pitfalls and conundrums in type 1 von Willebrand disease. Hematology Am -Soc Hematol Educ Program. 2022(1):618-623.

One very helpful topic that is timely in today’s coagulation world, was Monitoring Alternative Anticoagulants by Dr. Eric Salazar, MD from UT Health San Antonio, TX. Dr. Salazar discussed the uses and methods of monitoring direct oral anticoagulants (DOACs). These include Factor Xa inhibitors (anti-FXa) like apixaban, rivaroxaban, and edoxaban, and direct thrombin inhibitors (DTI) like dabigatran, argatroban; and bivalirudin. He talked about real-world situations which may require specialized testing and implementation of laboratory measurement of these anticoagulants.

Measuring FXa inhibitors may be warranted in certain situations:

  • With suspected hemorrhaging
  • Patients with a history of bleeding
  • To clear a patient for a surgical procedure
  • When checking for clearance of heparin
  • Beginning a new medication or dose change of medication
  • Elevated D-dimer and PT/INR
  • ESRD/CKD and a decline in renal function
  • Post-reversal of a DOAC
  • Subjects with a BMI> 40 kg/m2 or Weight <60 kg

Dr. Catherine Hayward, MD., Professor of Pathology and Molecular Medicine, McMaster University gave a presentation on Rare Inherited Coagulation and Fibrinolytic Defects that Challenge Diagnostic Laboratories.

Dr. Hayward discussed the importance of how mutations could affect coagulation factors, anticoagulants, and fibrinolytic proteins. These mutations actions may cause the formation of rare bleeding conditions that make it difficult to identify a diagnosis.

The presenter gave the latest information on how clinicians and laboratories should consider the presence of seldom seen coagulation disorders when confronted with a difficult diagnosis.

I suggest reading the manuscript that this presentation was derived from: Mathews N, Tasneem S, Hayward C. Rare inherited coagulation and fibrinolytic defects that challenge diagnostic laboratories. Int J Lab Hematol. 2023 45Suppl2:30-43. The submission discusses Bleeding disorders caused by autosomal dominant F5 mutations that affect FV splicing and secondarily increase plasma tissue factor pathway inhibitor levels (TFPI),  combined FV and FVIII deficiency (F5F8D), inherited thrombomodulin-associated coagulopathies, familial Vitamin K-a  clotting factor deficiency (VKCFD), inherited thrombomodulin-associated (TM) coagulopathies, congenital disorders of glycosylation (CDG), inherited disorders of fibrinolysis (SERPINS).

The manuscript contains a vast amount of information on testing by procedures that may not be available except in special coagulation laboratories to make a clinical diagnosis.  The authors stress the need for detection of pathogenic mutations in these rare bleeding disorders.

The last oral presentation I wanted to mention was presented on Saturday morning at 7 am on the last day of the conference and was one of the most popular sessions.  The topic was intriguing, Update on Coagulation Mixing Studies: Looking for Harmonization, and it was presented by one of the top thought leaders in the last several decades in the field of Thrombosis/Hemostasis, Dr. Dorothy (Dot) Adcock, M.D., Laboratory Corporation of American, Retired Chief Medical Officer, Burlington, NC.

I have been dealing with some form of mixing studies since 1974.  The consensus on how to calculate and interpret a result to determine presence of a factor deficiency or an inhibitor has never really been decided by research or “professional opinion.” There was the 1:2 mix of equal parts of normal pooled plasma (NPP) and patient specimen. Then we had the 1:4 and even the 1:10 mixing testing.  One way to suggest a factor deficiency was if it corrected within 5 seconds of the original test result.  The presence of an inhibitor was determined if result was greater than 5 seconds of the reference range of the NPP limit of the assay. This was a classic example of “professional opinion.” However, I remember some interpretations such as this scenario. The APTT was 80 seconds.  The 1:1 mix then gave a 51 second result.  The normal reference range was 25-35 seconds.  I witnessed some laboratories who then ran both studies for the presence of a lupus anticoagulant (LA) and a factor deficiency because “it corrected over 20 seconds, but it didn’t get into the reference range or 5 seconds within the upper limit.”  Then then there was the issue of running incubated mixing studies and interpretating results. It was a lot of “hit or miss.”  Dr. Adcock’s presentation provides good data for standardization of the issue of how to interpret and perform coagulation mixing studies.  She reported on a field study that was performed in conjunction with the International Council for Standardization in Haematology (ICSH). They performed a multi-site study that examined 11 different methods for evaluating and calculating mixing studies using several PT and APTT reagent/instrument combinations. The study showed that using the Rosner Index, which subtracts the clotting time of the pooled normal plasma (PNP) from the clotting time of the 1:1 mix, and percentage correction calculations gave the lowest misprediction data.

While searching the literature I found an interesting paper published by Tori Seasor, MD, Karen A Moser on Mixing studies (Seasor T, Moser KA. Mixing studies. https://www.pathologyoutlines.com/topic/coagulationmixingstudies.html). They discussed differential diagnosis of prolonged PT or APTT in concurrence with mixing study data and correlated it with diagnostic situations.

A recommendation was to use commercially purchased NPP that was pooled with at least 20-30 normal donors who were screened for PT, APTT, Fibrinogen, Factors V-XII and checked for inhibitors. This could save the laboratory a lot of work and aid in the standardization of mixing study testing.

ISLH 2023 INTERESTING ABSTRACTS

One of the most interesting abstracts I observed at the meeting helped clarify what may be a common point of confusion. While use of the chromogenic FVIII assay is used to measure FVIII levels, there has been a question of how to measure emicizumab (Hemlibra) itself. An abstract by SJ McBryant, Pickering WM, Robinson MM, Poier BF. 2023 Validation of a modified Factor VIII one-stage clotting assay using the Siemens BCS XP to measure emicizumab on plasma was presented at this meeting. It discussed an adaptation for performing the one-stage clotting assay method using an ellagic acid activated APTT reagent with an assigned value calibrator, using a 7-point calibration curve that used emicizumab as the calibrator material. They were able to show data that using this method can give accurate results of the levels of emicizumab.

One of the most discussed topics at the meeting dealt with methods and performance of mixing studies used in determining whether there is the presence of an inhibitor or factor deficiency interpreting correction or non-correction of results.  As we know there are no standards or guidelines on what a correction is.  If you read the earlier discussion on the presentation by Dr. Dorothy Adcock, M.D., and the paper by Seasor T et al, we are still struggling with this issue.

An abstract was presented by Moiz B, Rizvi B, Tahir M, Rashid A. 2023, Performance evaluation of mixing studies prior to factor assay that was very informative.  The results were based on APTT corrections.  As we know there are no standards for mixing study corrections or consensus what the range would be to denote the difference between a factor deficiency or the existence of an inhibitor. They used a laboratory established upper range and then compared the results using the Rosner Index, ratio, and percentile correction methods. The Rosner index showed the results that best correlated with their in-house methods.

A question came up recently from a colleague about the interference of hemolysis on routine coagulation tests on different regent/instrument combinations such as PT, APTT, TT, fibrinogen, and D-dimer assays.  There are several manuscripts and opinions on this subject. There was one abstract presented at this meeting that looked at this topic with a very “practical eye.” The abstract authors: Tarpley G, Horner K, Bowyer A, Kitchen S. 2023, The use of paired matched samples to determine the true effect of haemolysis interference on routine coagulation tests on the Sysmex CN-series coagulation analyzer. 

The issue of whether to refuse a hemolyzed specimen for coagulation testing or to run the sample with a comment on possible interference added on to the test result has long been a subject of speculation for the laboratory.  We know about elevations of potassium and LDH, but little about the effect on assays for monitoring coagulation studies.  Hemolysis “noise” in coagulation testing can depend on the reagent/instrument combinations, whether the system uses a photo-optic or mechanical system and how the specimen is collected and processed prior to testing.  In this study, hemolyzed specimens clotting times were decreased in the PT, APTT, TT.  The APTT showed the largest decrease. Therefore, hemolyzed specimens for the APTT could impact the patient diagnosis and treatment.

As of this writing, the presentations and abstracts of all these discussions are posted on the ISLH 2023 website. 

Further reading:

Palic J, Lapic I, Rogic D. The Reliability of Hemolysis, Incterus and Lipemia Incides on Automated Coagulation Analyzers Sysmex CS-5100 and Atellica Coag 360. Nov, 2021 Poster DOI:10.1515/cclm-2021-5035.

 

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