White Blood Cell Disease

WHITE BLOOD CELL DISEASE

National Taiwan University Hospital Department of Internal Medicine Division of Hematology 陳建源

Non-malignant Disorder of Leukocyte

1.Variations of leukocytes in disease 2.Neutropenia 3.Qualitative disorder of leukocytes 4.Abnormal monocyte-macrophage

system 5.Langerhans cell histocytosis X 6.Infectious Mononucleosis 7.Hematology aspects of HIV and

AIDS 8.Disorder of spleenWintrobe’s Clinical Hematology 10th ed

Variations of leukocytes in disease

Wintrobe’s Clinical Hematology 10th ed

Neutropenia-acquired

The most common cause of acquired neutropenia is infection. HBV, EBV, and HIV.

The second most common is medica-tion exposure: beta-lactam antibiotics, anti-thyroid drug, anti-tuberculosis, ticlopidine, baktar, carbamazepine, captopril, digitals, indomethacin


Neutropenia-congenital

Kostmann syndrome Severe congenital neutropenia Cyclic neutropenia Ela2 gene HAX1 is a ubiquitously expressed mitochondrial

protein with weak homology to bcl-2 Wiskott-Aldrich syndrome protein (WASp) Gfi-1 zinc-finger protein, transcriptional repressor

Nancy Berliner Blood 2008

Qualitative disorder of leukocytes Pelger Huet Anomaly: AD, bilobed

PMN Alder-Reilly Anomaly: AR, Large

azurophil granules May-Hegglin Anomaly: AD, inclusion

in granulocytes, giant platelets with thrombocytopenia

Chediak-Higashi Anomaly: AR, lysosome-like inclusion body, children with pale hairWintrobe’s Clinical Hematology 10th ed

Abnormal monocyte-macrophage system Fabry disease: X-linked,

glycosphingolipid anemia, decreased serum iron concentration, platelet aggregation. Lipid-laden foamy macrophage in BM

Gaucher disease: AR, Glycocerebroside

Niemann-Pick disease: sphingomyelin and cholesterol accumulation Wintrobe’s Clinical Hematology 10th ed

Langerhans cell histocytosis X

Hand-Schuller-Christian Disease Clinical Triad: defect in membranous

bone, exophthalmos and polyuria Pathology: granulomatosis,

histocytes, mature eosinophil, and lymphocytes

Birbeck granules


Infectious Mononucleosis

Sorethroat and dysphagia (80-85%) Lymphadenopathy Hepatosplenomegaly Fever, malaise Lymphocytosis Abnormal liver function Hyperbilirubinemia


Infectious Mononucleosis

Polyclonal T cell, CD8 subset Hematological complications Immune hemolytic anemia Immune thrombocytopenia Granulocytopenia Marrow aplasia Virus-associated hemophagocytic syndrome Acquired immune Deficiency Non-Hematological complication Splenic rupture, Neurological complication, Cardiac

complication, Respiratory complications, Liver failure, Pancreatitis, Renal Failure

Decreased Production Drugs Zidovudine, Trimethoprim-sulfamethoxazole Amphotericin B, Ganciclovir, Dapsone,

Delavirdine Deficiencies Erythropoietin, Iron, Folate, Vitamin B12 Infection HIV, Parvovirus B19, Mycobacterium avium

complex (MAC), Mycobacterium tuberculosis, Histoplasma capsulatum

Neoplasia Non-Hodgkin’s lymphoma, Multiple myeloma, Castleman’s disease, Hodgkin’s disease Miscellaneous Anemia of chronic disease Preexisting condition (sickle cell disease,

thalassemia,

Hematology aspects of HIV and AIDS: anemia

Hematology Am Soc Hematol Educ Program. 2003:294-313

Increased LossHemolysisThrombotic thrombocytopenic purpuraGlucose-6-phosphate dehydrogenase deficiencytrimethoprim-sulfamethoxazole, dapsone, primaquine, Autoimmune hemolytic anemiaIdiopathicDrugs (ceftriaxone, indinavir, “Ecstasy”)Infection (cytomegalovirus [CMV])Gastrointestinal bleeding Kaposi’s sarcoma Non-Hodgkin’s lymphoma Infection (CMV, Candida)HypersplenismInfectionLymphomaHemophagocytosisCirrhosis (hepatitis B virus, hepatitis C virus)

Decreased ProductionDrugs: Trimethoprim-sulfamethoxazole, Pentamidine, Pyrimethamine, Ganciclovir, Fluconazole, Alpha-interferon, Rifabutin, Clarithromycin, Didanosine, Amphotericin B, Indinavir, Ritonavir, Delavirdine, NelfinavirNeoplasia Non-Hodgkin’s lymphoma

Miscellaneous

Preexisting condition

Infection HIV, Parvovirus B19,Mycobacterium avium complex (MAC), Mycobacterium tuberculosis,Histoplasma capsulatum, Bartonella henselae (bacillary angiomatosis)

Deficiencies Folate, Vitamin B12

Hematology aspects of HIV and AIDS: thrombocytopenia


Increased LossImmune thrombocytopenic purpuraThrombotic thrombocytopenic purpuraHypersplenism

Infection

Hemophagocytosis

Cirrhosis

Drugs

Saquinavir

Interferon

Hematology aspects of HIV and AIDS: neutropenia Decreased Production Drugs

Ganciclovir

Zidovudine

Trimethoprim-sulfamethoxazole

Pentamidine

Rifabutin

Antineoplastic chemotherapy

Dapsone

Amphotericin B

Ritonavir

Delavirdine

Nelfinavir

DeficienciesFolateVitamin B12 InfectionHuman immunodeficiency virus (HIV)Mycobacterium avium complex (MAC)Mycobacterium tuberculosisHistoplasma capsulatum NeoplasiaNon-Hodgkin’s lymphomaMultiple myeloma Increased LossAutoimmune neutropeniaHypersplenismInfectionHemophagocytosisCirrhosis


Disorder of spleenMechanism Causative diseaseImmune response work hypertrophy

Subacute bacterial endocarditisInfectious mononucleosisFelty’s syndrome

RBC destruction work hypertrophy

Spherocytosis

Congestive (Venous outflow obstrcution)

Thalassemia majorPyruvate kinase deficiency

Infiltrative SarcoidosisAmyloidosis

Neoplastic LymphomaChronic lymphocytic leukemiaHairy cell leukemiaMetastatic carcinoma

Myeloproliferative Chronic myeloid leukemiaMyeloid metaplasia

Miscellaneous TraumaSplenic cystHemangioma


Malignant hematopoietic disorder

Normal AML CML

Differentiation (++) (-) (+/-)

Proliferation (+/-) (++) (+++)

Stem cell

Mutation of transcriptional factor/ cofactorEx: PML/RARa, AML1, CEBPA, CBFB, NPM, MLL

Mutation of proliferation, survival geneEx: FLT3, PTPN11, RAS, Kit, JAK2, PDGF gene Acute

leukemia

MDSMPD

Mutation of transcriptional factor and proliferation gene

Tumor suppressor gene?

Interactions of Class I and Class II gene mutations

Class I mutation (No of Patients with the mutation)

Pt no KIT FLT3/ITD

FLT3/TKD

NRAS KRAS PTPN11 JAK2 Total*

Class II mutation AML1/ETO t(8;21)

33 7 2 2 4 1 0 2 16

PML/RARA t(15;17) 24 0 7 5 4 1 0 0 14

CBFB/MYH11 inv(16) 9 1 0 1 2 1 0 0 5

MLL t(11q23) 13 0 1 1 2 1 0 0 4

NUP98/HOXA9 t(7;11) 6 0 1 0 2 2 0 0 3

AML1 31 0 7 1 2 0 3 0 11

MLL/PTD 13 0 9 0 1 0 1 0 10

CEBPA 45 1 7 2 2 0 0 0 11

NPM1 63 0 33 9 9 0 6 0 49

Others** 96 1 16 4 11 2 4 1 36

Total* 324 10 78 24 39 8 14 3 155

* Some patients had more than one mutations.** Excluding patients with the Class II gene mutation shown above.

(NTUH 1995-2003)

NTUH 1995-2003

Interactions between Class I and II Gene Mutations

Class I Gene: Proliferation and Survival

Class II Gene:

Differen-tiation

FLT3/ITD FLT3/TKD NRAS KRAS PTPN11 JAK2 KIT

AML1/ETO t(8;21) P=0.011 P=0.028 P<0.001

PML/ RARα t(15;17) P=0.024

CBFβ /MYH11 inv(16) MLL 11q23NUP98-HOXA9 t(7;11)

P=0.008

NPM1 P<0.001 P=0.03 P=0.035

CEBPAAML1MLL/ PTD P<0.001

: Positive association NTUH 1995-2003: Negative association

Minimal residual disease

Flow cytometry for MRD detectionLeukemia-associated Aberrant Immunophenotypes(LAIP) Classification Cross-lineage expression of lymphoid antigensCD33+CD2+CD34+CD34+CD13+CD19+ Over-expression HLA-DR++CD33++CD34++CD64++CD4++CD45++ Lack of expression of antigen HLA-DR-CD33+CD34+ Asynchronous expression of antigens CD15+CD33+CD34+CD65+CD33+CD34+ + indicates expression; ++, over-expression; -, no expression.

Kern W et al. Cancer 2008 112(1):4-16

Myeloproliferative disorder ET, PV, MF, MPD, unclassified JAK2 mutation mpl W515L(5% MF, 1%ET)

PTPN11:JMML PDGFR: CMML, HES

Diagnosis Frequency of JAK2 V617F(%) Frequency of JAK2 V617F homozygosity(%)

MPD PV >90 24-27

ET ～ 50 3-4

MF ≧50 6-18

CML Rare ND

aCML <20 ND

HES <2 ND

mastocytosis Rare ND

MPD/MDS CMMoL <5 ND

JMMoL <20 ND

MDS ～ 5 ND

AML De novo AML 1 ND

AML from MPD ≦50 ND

ALL None ND

Hodgkin None ND

NHL None ND

The frequency of JAK2 V617 mutation in hematopoietic disorders

Human Pathology 2008(39):795–810

PV: 2001 WHO criteria and the proposed 2007 WHO criteria

2001 WHO criteriaDiagnosis requires:

Both 1 and 2 of the A criteria, plus 1 additional A criteria, or 2 B criteria

A-criteria:

1.Increased red cell mass 1. >25% above mean normal predicted value, or >18.5 g/dL in men. >16.5 g/dL in women, or >99th percentile of method-specific reference range for age, sex, altitude of residence

2. No cause of secondary erythrocytosis, including:

Absence of familial erythrocytosis, No elevation of EPO caused by: i.Hypoxia arterial PO2 ≤92%, ii. High oxygen affinity hemoglobin, iii. Truncated EPO receptor ,iv. Inappropriate EPO production by tumor

3. Splenomegaly

4. Clonal genetic abnormality other than Philadelphia chromosome or BCR-ABL fusion gene in marrow cells

5. EEC formation

B-criteria:

1. Thrombocytosis >400 × 109/L

2. Leukocytosis >12 × 109/L

3. Bone marrow biopsy showing panmyelosis with prominent erythroid and megakaryocytic proliferation

4. Low serum EPO levels

Diagnosis requires:Both major criteria and one minor criteria, or the first major criterion and 2 minor criteriaMajor criteria: 1. Hemoglobin >18.5 g/dL in men,Hemoglobin >16.5 g/dL in women, or Other evidence of increased red cell volume 2. Presence of JAK2 V617F or other functionally similarmutation such as JAK2 exon 12 mutation Minor criteria:1. Bone marrow biopsy showing hypercellularity forage with trilineage growth (panmyelosis) with prominent erythroid, granulocytic, and megakaryocytic proliferation 2. Serum EPO level below the reference range . 3. EEC growth

Proposed 2007 WHO criteria

Smith CA. Human Pathology 2008:795-810

Chronic myeloid leukemia

Philadelphia chromosome(+)

t(9;22) BCR-ABL Tyrosine kinase Glivec Dasatinib Niclotinib

Mechanism of BCR/ABL resistance Mechanisms of Imatinib

Resistance Decreased intracellular drug levels Plasma binding by -1 acid

glycoprotein Drug efflux from P-glycoprotein

(MDR-1) overexpression Increased expression of BCR-ABL

kinase from genomic amplification Clonal evolution (non-BCR-ABL–

dependent mechanism) Mutations in ABL kinase of BCR-

ABL affecting drug interaction or kinase activity

Arch Pathol Lab Med. 2006 May;130(5):669-79.

Myelodysplastic syndrome• Nuclear Budding• Nuclear fragmentation• Micro-megakaryocyte• Nuclear-cytoplasm

differentiation disassociation

• apoptosis

Myelodysplastic syndrome

Clinical manifestation: cytopenia, ineffective hematopoiesis, leukemic transformation

Cytogenetic: -5, -7, +8 , 20q, complex Classification: FAB, WHO, IPSS Treatment: Supportive, Allogeneic HSCT, ATG, CsA

Chemotherapy(Low dose Ara-C or Standard dose) , Danazole, Demethylating agent , lenalidomide

Blood. 2008 May 15;111(10):4841-51.

Myelodysplastic syndrome

Ringed sideroblasts and associated with marked thrombocytosis (RARS-T), JAK2 V617F mutation

5-Azacytidine was approved for the treatment of MDS in 2004

Azacytidine prolonged survival, delayed progression to AML, and improved quality of life. Complete response (CR) rate 7%, 16%PRs and 37% hematologic improvement also seen.

Decitabine was also evaluated in a phase 3 trial, and a 35% overall response rate was seen (9% CR, 8% PR, 18% hematologic improvement).

Blood. 2008 May 15;111(10):4841-51.

Acute Myeloid LeukemiaFavorable-Risk Group

Balanced structural rearrangements t(15;17)(q22;q12-21), t(8;21)(q22;q22)inv(16)(p13q22)/t(16;16)(p13;q22)

Intermediate-Risk Group

Normal karyotype

Balanced structural rearrangements t(9;11)(p22;q23)

Unbalanced structural rearrangements del(7q), del(9q), del(11q), del(20q)

Numerical aberrations: –Y, +8, +11, +13, +21

Unfavorable-Risk Group

Complex karyotype

Balanced structural rearrangements inv(3)(q21q26)/t(3;3)(q21;q26)t(6;9)(p23;q34), t(6;11)(q27;q23)t(11;19)(q23;p13.1)

Unbalanced structural rearrangements del(5q)

Numerical aberrations: -5,-7


Genetic alterations affecting clinical outcome of cytogenetically normal acute myeloid leukemia (AML) patientsGenetic Alteration Prognostic SignificanceFavorableNPM1 mutations Patients with NPM1 mutations who do not harbor FLT3-ITD have significantly

better CR rates, EFS, RFS, DFS, and OS than patients without NPM1 mutations and FLT3-ITD.NPM1 mutations do not have a significant effect on prognosis of patients with FLT3-ITD.

CEBPA mutations Patients with CEBPA mutations have CRD and OS significantly longer than patients with the wild-type CEBPA gene.

Unfavorable

FLT3-ITD Patients with FLT3-ITD have significantly shorter CRD, DFS and OS than patients who do not harborFLT3-ITD.Particularly poor prognosis is conferred by FLT3-ITD coupled with no expression of a FLT3 wild-type allele or a high FLT3 mutant to FLT3 wild-type allele ratio.

MLL-PTD Patients with MLL-PTD have remission duration significantly shorter than patients without MLL-PTD.


Kit mutation in patients with t(8;21) most studies revealed unfavorable outcome, in patients with inv(16) no significance related to outcomes

Acute lymphoblastic leukemia

Children

Adults

Peak incidence 5 years 50 years% of Leukemias 80-85% 15%Chromosomes Ph+ 3% 30% MLL 1-2% 7% TEL/AML1 20% 2%

Hyperdiploid 25% 5%

T-cell 10-15% 20-25%Mature B 1-2% 3-5%

Acute lymphoblastic leukemia (ALL): incidence & biological differences


Acute lymphoblastic leukemia

Subgroup Disease characteristics Cytogenetic / Molecular markers

Specific Poor prognostic factors

General Poor Prognostic factors

Pro-B-ALL (11%)(CD10 negative)

-high WBC (>100,000/μL in 70%) -CD13/CD33 coexpression (> 50%)

-70% t(4;11)/ALL1-AF4-(20% Flt3 in ALL1-AF4+)

High Risk -Late achievement of CR-Poor PRED response (?)-MRD persistence-Increasing age-In-vitro resistance (?)-MDR1 function (?)-Complex aberrant karyotype (?)

c-ALL (49%)pre-B-ALL (12%)

-incidence increasing with age(75% if > 55 yrs)-partly CD20+ (45%)

-4% t(1;19)/PBX-E2A(pre-B only)

-WBC > 30-50,000/μL-t(9;22)/BCR-ABL-t(1;19)/PBX-E2A (?)

Mature B-ALL (4%)(L3-ALL, Burkitt leukemia)

-large tumor mass(LDH increased in > 90%)-organ involvement (32%)-CNS involvement (13%)-CD20+ (> 80%)

-t(8;14)/c-myc-IgH

T-ALL (25%) -mediastinal tumor (60%)-CNS involvement (8%)-high WBC (> 50,000) (46%)Subtypes:Early T (6%)Thymic T (12%)Mature T (6%)

20% t(10;14)/HOX11-TCR< 20% t(11;14/LMO/TCR8% SIL-TAL14% NUP213-ABL133% HOX11**5% HOX11L2**50% Notch1**

-Early, mature T-ALL-WBC > 100,000/μL (?)-HOX11L2

Chronic lymphoblastic leukemia

Prognostic factor: Traditional staging according to Rai or Binet IgHV mutation status,lymphocyte doubling time morphology,Immunophenotype characteristics, CD23 expression, Beta-2-microglobulin, Cytogenetics, ZAP-70, Pattern of bone marrow involvement, Cytogenetic: 13q14 in about 50% of cases (by FISH) followed by del

11q22–q23 (20%), trisomy 12 (15%), del 6q21 (10%) and del 17p13 (5–10%).

BJH 2007 139(5):630-634Blood 2005 105(5):1839-1840

Surface marker of B-Cell Lymphomas

36

Surface Marker MCL FL SLL/CLL MZL

CD5 ++ - ++ -

Surface Ig ++ +++ + +

CD19 ++ ++ ++ ++

CD20 +++ ++ + (weak) ++

CD10 - + (80%) - -

CD23 - +/- + -

Cyclin D1 +++ - - -

Cytogenetics t(11:14) t(14:18) - t(11:18)

EBV associated lymphoproliferative disorder

Blood. 2006 Feb 1;107(3):862-9.

EBV associated lymphoproliferative disorder

Rituximab Plus Chemotherapy in First-Line Therapy of Advanced Stage Follicular Lymphoma

Lymphoma Follicular

Author Regimen P

Hiddemann et al CHOP (n = 205) R-CHOP (n = 223) Response rate 90% 96% .011 Median time to treatment failure

31 months Not reached < .0001

Marcus et al CVP (n = 159) R-CVP (n = 162) Response rate 57% 81% < .0001 Median time to treatment failure

7 months 27 months < .0001

Herold et al MCP (n = 96) R-MCP (n = 105) Response rate 75% 92% < .001 Median event-free survival 19 months Not reached < .0001

Salles et al CHVP/IFN- (n = 175) R-CHVP/IFN- (n = 184) Response rate 85% 94% < .0001 Median event-free survival Not reached Not reached

Hochster et al CVP (n = 157) CVP + R (n = 148) Median event-free survival 17 months* 50 months*

J Clin Oncol. 2005 Sep 10;23(26):6394-9

40

t(11:14)Cytogenetics

+++Cyclin D1

-CD23

-CD10

+++CD20

++CD19

++Surface Ig

++CD5

MCLSurface Marker • Small-medium sized cells, indented nuclei; occasional small-cell variant

• Expresses B cell antigens (CD20); CD5• Over-express cyclin D1

(immunohistochemistry)• Genetic hallmark: t(11;14) (q13;q32)

translocation• Molecular: BCL1 translocation, mutated

IGH ~20-30%

Mantle Cell Lymphoma

NCCN Mantle Cell Lymphoma Guidelines 2006

Rituximab + HyperCVAD Rituximab + CHOP Rituximab + EPOCH

ASCTAllogeneic transplant in the context of a clinical trial

BortezomibCladribineFCMR FC PCR Thalidomide + Rituximab

41

1st Line

• HyperCVAD (cyclophosphamide, vincristine, doxorubicin, dexamethasone) • CHOP (cyclophosphamide, doxorubicin, vincristine, prednisone)• EPOCH (etoposide, prednisone, vincristine, cyclophosphamide, doxorubicin)• FCMR (fludarabine, cyclophosphamide, mitoxantrone, rituximab)• FC (fludarabine, cyclophosphamide) ± rituximab• PCR (pentostatin, cyclophosphamide, rituximab)• Thalidomide + rituximab

1st Line Consolidation

Second-line Therapy

Lymphoma MALToma

J Clin Oncol. 2005 Sep 10;23(26):6370-8

Lymphoma Follicular

No. of risk

factors*

FLIPI score Proportion of patients,

%

Overall survival

at 5 y, % at 10 y, %

0-1 Low 36% 91 71

2 Intermediate 37% 78 51

3-5 high 27% 53 36

Prediction of patients with follicular lymphoma outcome based on the FLIPI

*Factors adversely affecting survival in the FLIPI include age greater than 60 years; Ann Arbor stage III–IV; number of nodal sites greater than 4; serum LDH level greater than the upper limit of normal; and hemoglobin level less than 12 g/dL.Hematology 2007:216-

225

NK Lymphoma

Lymphoma ALCL

Pleomorphic large lymphocytes, Young adult

CD30+ (CD 30 also expressed in CTCL, lymphomatoid papulosis, regressing atypical histocytosis, HD and some

embryonal cell, pancreatic carcinoma) t(2;5)(p23;q35) NPM/ALK Nodal Extranodal: skin

Plasma cell labeling index (PCLI)

PLASMA CELL DYSCRASIA

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White Blood Cell Disease