Laboratory HaematologyServices in Sarawak – Meeting

the Needs of Modern Day Clinical Practice

Dr. Henry R Gudum

Professor of Haematology, Dept. of Pathology

FMHS, UNIMAS

Outline

• History• Haematology workload• Specialised haematology tests• Resources• Needs of modern day clinical practice

Background & History

• Haematology Lab services – Central Medical Laboratory• Central Medical Lab – funded from HQ and SGH; compiles budget for

lab services in the whole state of Sarawak

Background & History – Prior to 1985

• No Anatomic pathologist/Haematologist• Dr. Rajamani (Expat from India) – Microbiology• Dr. Kothari (Cytologist, WHO) – conducts pap smear workshops• Dr. Thomas Chung – Medical officer-in-charge• All biopsies and BMAT specimens sent to IMR or Hosp with

pathologists in Pen. Malaysia• Lab superintendent – in charge of Central Lab (Australian (1972),

Bartholomew Tan (1974)• Lab technologists – in charge of peripheral Labs

Background & History – 1985 and after

• 1985 - Qualified Pathologist (Local) – Dr. Mapphy Duncan started serving in the Central Lab

• Pathologist in Charge• Reports all biopsies, cytologies, including bone marrows and trephines

• 1987 (September) – 1. Dr. Jamil Dolkadir (Haematologist), 2. Dr. Henry R Gudum (Haematologist) – joined Central Lab after specialist training; (May 1988 – Dr. Henry R Gudum was transferred to QEH to start pathology service in Sabah); Later Dr. Jamil became Forensic specialist; Dr Mapphy Duncan left for private practice in 1991

• Dec 1994 Dr. Henry R Gudum joined UNIMAS – helped in Central Lab. In reporting of BMs, and other consultations; also visiting consultant haematologist to private hospital (lab and clinical)

• Clinical Haematology:• Dr Lau Lee Gong – came back in 2003 after clinical haematology training –

joined private hospital (Visiting Consultant to SGH)• Dr Chew Lee Ping – came to SGH as Clinical Haematologist in 2009• Dr Kuan Jew Win – Clinical Haematologist joined SGH much later – now in

UNIMAS (Assoc. Prof.)

Background & History – 1985 and after

Workload

Year Samples Tests2012 169817 419864

2013 168775 426691

2014 177825 445712

2015 190602 491886

2016 193749 502301

BMAT REQUESTS

Year No. of Requests2012 272

2013 379

2014 594

2015 372

2016 321

Specialised Haematology Tests

• Immunophenotyping – HKL• Cytogenetics – HKL, IMR• Molecular genetics – HKL, IMR• Coagulation Factor / Inhibitor Assays – SGH (FVIII, FIX); Ampang Hosp.

(Other coagulation Factors)• Thrombophilia Testing (ATIII, Protein C, Protein S, FV Leiden,

Prothrombin (G20210A) Mutation, etc) – Ampang Hosp.• Platelet Function Testing – Send patient to National Blood Centre

Specialised Haematology Tests

• Immunohistochemistry on trephine biopsies – various immuno markers for leukaemia and lymphoma are available in SGH; if referral or additional IHC required – send to HQE (Dr. Ahmad Toha)

Human Resource

• MLTs – 17 (in Haematology unit SGH)• Lab Haematologists

• SGH – 2• Miri Hosp. – 1• Sibu Hosp. – 1 (to come in 2018)

Major Equipment

• Haematology Analysers – 5• Coagulation Analysers – 2• HPLC for HB Analysis – 1• Flow cytometry equipment - 1

Space - inadequate

Future Expansion/Services

• To expand flow cytometer services beyond CD4/CD8 counting to include immunophenotyping services for leukaemia and lymphomas

• New lab building (building expected to start end of 2017)

Needs of Modern day clinical haematology practice

• New Classification of Haematological Malignancies• Targeted therapies• Disease monitoring• New anticoagulants (NOACs/DOACs)• Anti-Xa in UFH, LMWH

2BLOOD, 19 MAY 2016 x VOLUME 127, NUMBER 20

Acute Myeloid Leukaemia

• AML with recurrent genetic abnormalities• AML with myelodysplasia-related changes• Therapy-related myeloid neoplasms• AML, not otherwise specified• Myeloid sarcoma

• Myeloid proliferations related to Down syndrome• Blastic plasmacytoid dendritic cell neoplasm• Acute leukaemia of ambiguous lineage

AMLwith Recurrent GeneticAbnormalities• Recurrent genetic abnormalities: retained

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New entities addedPrognostic significance of mutations updatedNew genetic (genomic) discoveries not included

Lymphoblastic Leukaemia / Lymphoma

▪

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Acute erythroid leukaemiaNomenclature for myelodysplasia: “MDS with …..”

• New provisional entities (genetic basis):▪

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AML with BCR-ABL1; CEBPA bi-allelic; mutated RUNX1ALL with iAMP21 and B-ALL, BCR-ABL1-like; ETP-ALL

• Genetic changes and revised classification:▪

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SF3B1 in MDS and 5% ring sideroblasts2nd genetic abnormality in MDS with del(5q)

Not all genetic changes directly impact diagnosis

WHO Classification 2016 RevisionsArber D. Blood, 2016: 127 (20): 2391-2405

• New diagnostic criteria (morphology):

2008Both major and 1 minor or1st major and 2 minor

2016/17All 3 major or1st 2majorandoneminor

Major1.Hb>18.5(M)>16.5(F) orOther evidence of increased RC volume*

1.Hb>16.5(M)>16.0(F) orHct>49(M)>48(F)

2.JAK2+(either exon 14 or 12) 2.BM hypercellularity for age,Panmyelosis with pleomorphicMEG**

3.JAK2+

Minor 1.BM trilineage myeloproliferation (panmyelosis) 1.Subnormal EPO

2.SubnormalEPO

3.EndogenousErycolonygrowth

increase in fibrosis can be detected

*Hb or Hct >99th percentile of ref range for age/sex/altitudeorHb>170(M) >150(F) if sustained increase of 20g/l from baselineorelevated RCM >25% above mean normal

**BM examination not necessary forclinically overt PV, i.e. JAK2+ andHb/Hct >18.5/52 M >16.5/48 F

BUT only by performing BMB an

Polycythaemia Vera

2008All 4 major criteria:

2016/17All 4 major or1st 3 major + the minor criterion

Major Plt≥450 Plt≥450

MEG proliferation with large and maturemorphology

BMB: mainly MEG proliferation with large andMature morphologyNo significant increase or left-shift of neutrophilGranulopoiesis or erythropoiesis and very rarelyMinor increase in reticulin fibers (grade1)

Not meeting WHO criteria for CML, PV,PMF, MDS or other MN

Not meeting WHO criteria for CML, PV,PMF, MDS or other MN

JAK2 or other clonal marker or noEvidence of reactive thrombocytosis

JAK2, CALR or MPL mutation

Minor Presence of a clonal marker or noEvidence of reactive thrombocytosis

Essential Thrombocythaemia

2008All 3 major + 2 minor

2016/17All 3 major and at least one minor(confirmed in 2 consecutive determinations)

Major

1.MEG proliferation and atypia, accompanied byfibrosis; if no fibrosis, increased cellularity,Granulocytic proliferation and decreasederythropoiesis

Pre-PMF1.MEG proliferation and atypia without>1gr 1 fibrosis plusIncreased for age cellularity, granul. Proliferation and oftendecreased EryPMF1.MEG proliferation and atypia accompanied by eitherreticulin and/or collagen fibrosis grades 2 or 3

2.Not meeting criteria for CML, PV, MDS or other MN 2.Not meeting criteria for CML, PV, ET, MDS or other MN

3.JAK2 or other clonal marker or no evidence ofReactive fibrosis

3.JAK2, CALR or MPL or in the absence, presence of anotherClonal marker*or exclusion of reactive fibrosis

Minor Anaemia Anaemia not attributable to another condition

WBC≥119x10/l

Splenomegaly Palpable splenomegaly

IncreasedLDH LDH above upper normal limit

Leukoerythroblastosis Leukoerythroblastosis

is helpful to determine clonality

In the absence of JAK2/CALR/MPL finding mutations inASXL1, EZH2, TET2, IDH1,2, SRSF2, SF3B129

PMF

Targeted Therapies

• Therapeutics directed at specific molecular “lesions” responsible for carcinigenesis:

• Rituximab – monoclonal anti CD20 – Non-Hodgkin’s lymphoma• Imatinib (Gleevec) – TKI (bcr-abl oncogene) for Rx of CML• Nilotinib (Tasigna) – TKI (bcr-abl oncogene) for Rx of CML• Ruxolitinib (Jakavi) – inhibitor of JAK1 and JAK2 – for Rx of PMF and PV• Ibrutinib (Imbruvica) – targets TP53, 17p in CLL, ?Mantle zone lymphoma

Disease Monitoring

• Minimal residual disease – AML, ALL – e.g. molecular genetics, immunophenotyping

• Molecular monitoring for CML – bcr-abl quantitation – response matters as optimal response is associated with best long-term outcome; detect early resistance/treatment failure

Genetic Testing in VKA/Warfarin Therapy

Eur J Clin Pharmacol 2010;66(5):525–530Int J Clin Pharm (2013) 35:359–368

Genes affecting response to VKA – genes encoding:• Cytochrome P450 2C9 (CYP2C)• Vitamin K epoxide reductase complex 1 (VKORC1)

• Early studies - pharmacogenomic testing for warfarindosing is more accurate that other dosing schemes.

- Improves time to a therapeutic INR

- Requires fewer dosing adjustments.

• Patients who require higher or lower than usual dosesseem to benefit the most.

• BUT cost effectiveness and prevention of bleeding orthrombosis complications required further evaluation

THROMBIN

Dabigatran

RivaroxabanApixabanEdoxaban

Xa

IIa

Advances in Anticoagulation -- Direct Oral Anticoagulants (DOACs)

Prevention of stroke andsystemic embolism in atrialfibrillation(AF)

VTE prophylaxis in majororthopaedic surgery

Treatment of acute VTE andsecondary prevention ofrecurrent VTE

Prevention of cardiovasculardeaths after acute coronarysyndrome (Rivaroxaban)

DOAC - Ideal Anticoagulant ?

• Oral administration

• Good Efficacy and Safety

• Metabolic Properties with No FEW food and drug interaction

• Specific Reversal Agents NOT available for ALL DOACs

• No need for regular coagulation monitoring

DOACs - Coagulation Monitoring and Lab Testing

• DOACs marketed before Lab tests for drug levels anticoagulation function available• Lab measurement for residual drug effect:

• Before surgery or invasive procedure• Trauma

• Suspected overdose - drug interactions, renal impairment• Recurrent Thrombosis• The bleeding patient

• Major Bleeds – access anticoagulation effects of drugs• Is bleeding due to high drug levels or other reasons ?

•••

used to study plasma concentrations and PK/PD propertieshigh specificity, sensitivity, selectivity, and reproducibility,Gold standard method for the measurement of DOACs

• Not a functional assay, recommended to measure active metabolites to avoid systematical bias whenassessing the performance of functional assays.

• Supplemented by specific and sensitive functional assays (i.e., dilute thrombin time [dTT] or ecarin-basedassays for dabigatran and chromogenic anti-Xa assays for direct factor-Xa inhibitors)

• A major limitation - absence of standardization or harmonization of mass spectrometry-based assays.

• To improve inter-laboratory reproducibility, high-quality and international reference standards are urgentlyneeded

• NOT Readily available in most labs

Mass Spectrometry and the Measure of DOAC Plasma Levels

• Liquid chromatography coupled with tandem-mass spectrometry (LC-MS/MS)

Seminars in Thrombosis & Hemostasis Vol. 43 No. 3/2017

Anti-Xa for monitoring UFHeparin

• Shorter time to therapeutic target• Less dosing changes in 24 hours• Less testing

Guervil, et al Ann Pharmacother 2011; 45:861-8

Lab monitoring of LMWH and Fondaparinux

• Necessary in certain situations/patients• pregnant patients, children, obese patients, and patients with renal

impairment

• Gold Standard is Anti-Xa measurement

Acknowledgement

• Dr. Khamisah Gaus, Haematopathologist, SGH• Dr. Mapphy Duncan• Mr. Kho Hu Chiang

THANK YOU