The Scarlet Bedwetter

1
The Scarlet Bedwetter
L. Carolina I. Dumlao Internal Medicine Year
Level I
2
Objectives

• To distinguish cases of Paroxysmal Nocturnal
  Hemoglobinuria from other hemolytic anemias
• To identify clinical manifestations, especially
  life threatening sequelae of PNH

3
Case Identifying Data

• C.R.
• 39 year old
• Female
• Filipina
• Married
• Caloocan
• Call center

4
Case Chief complaint

• Anemia

5
Case History of present illness

• 30 years PTA low Hgb levels
• Iron supplementation
• 10 years PTA Hgb 4-5
• Transfused with 4u
• PRBC

6
Case History of present illness

• 6 months PTA Tea-colored urine
• at night or early
• morning
• 3 months PTA Shortness of breath
• () menorrhagia
• Hgb 7 2u PRBC

7
Case History of present illness

• Consulted hematologist
• Peripheral blood smear
• Microcytic/hypochromic
• Decreased WBC
• Normal platelet
• Iron deficiency anemia

8
Case History of present illness

• Iron studies normal
• Repeat PBS
• Normocytic/normochromic
• Decreased Hgb
• Decreased WBC

9
Case History of present illness

• Patient advised to undergo BMA
• ADMISSION

10
Case Review of Systems

• GENERAL No weight loss or weight gain. No focal
  weakness.
• SKIN No history of easy bruisability, no sores,
  no rashes nor pruritus
• HEENT No headaches, dizziness or vertigo. Does
  not wear corrective glasses, no history of eye
  pain, blurring of vision, excessive tearing,
  diplopia gross hearing is intact, no ear pain,
  discharge or infection no post- nasal drip or
  sinus pain no history of frequent sore throats
  
• PULMO No cough, no hemoptysis,
• CVS No history of orthopnea, chest pain, or
  bipedal edema. No palpitations noted.

11
Case Review of systems

• GIT No anorexia. No history of dysphagia,
  odynophagia, or jaundice. No hematemesis,
  hematochezia or melena. No note of change in
  character ot frequency of bowel movement.
• URINARY No history of UTIs. No intermittency,
  decreased caliber or incontinence.
• NEURO No history of syncope, seizures or tremors.
  No numbness or loss of sensation.
• HEMA No history of prolonged bleeding.
• ENDOCRINE No history of polyuria, polyphagia,
  polydipsia.
• No excessive sweating

12
Case Past health

• Non-hypertensive
• Non-diabetic
• No allergies
• No previous operations

13
Case Obstetric/Gynecologic history

• Menarche at 12 years of age
• Monthly, 2-3 pads per day, lasting 3 days
• Married for 10 years
• G0P0

14
Case Personal/Social history

• 2-3 pack smoker
• Stopped 1 year ago
• Occasional alcoholic beverage drinker

15
Case Family history

• Great grandfather - leukemia

16
Case Physical Exam

• VITAL SIGNS BP100/70mm Hg sitting HR92bpm
  
• RR 18breaths/min T36.8?C
• GENERAL Conscious, coherent, oriented to person,
  place and time.
• Weight 45kg Height 152.4cm BMI 19.4
• SKIN All extremities warm to touch. No
  discolorations, bruises or rashes.
• HEENT Normocephalic slightly icteric sclerae,
• pink conjunctivae no naso-aural discharge
• dry oral mucosa no tonsillopharyngeal
  congestion
• no neck mass noted no cervicolymphadenopathy
  noted.

17
Case Physical exam

• CHEST/LUNGS Equal chest expansion, no
  retractions clear breath sounds, no crackles
  or wheezing equal tactile and vocal fremitus
• CVS Adynamic precordium normal rate, regular
  rhythm, distinct S1/S2, no murmurs/gallops, PMI
  and apex beat at the 5th intercostal space left
  midclavicular line visible neck veins on
  supine position flattening with inspiration, full
  and equal pulses bilaterally (Gr. 3/5).
• ABDOMEN flabby abdomen normoactive bowel sounds,
  soft, no guarding or direct tenderness,
  rebound tenderness was absent no splenomegaly
• EXTREMITIES Normal muscle bulk and tone no
  cyanosis, edema or deformities no limitation
  of motion pale nail beds

18
Case Salient features

• 39/F
• Chronic anemia
• Tea-colored urine
• Iron studies normal
• Peripheral blood smear normocytic/normochromic

19
Anemia flowchart
Anemia
Volume depleted
Normovolemic
Reticulocytes increased
Reticulocytes decreased
Transfusion
Green, H. Decision Making in Medicine An
algorithmic approach
20
Anemia flowchart
Reticulocytes increased
Coombs test positive
Coombs test negative
Warm Ab
Cold Ab
-Autoimmune hemolytic anemia -Drug induced AHA
-1?/2?cold agglutinin -PCH
21
Anemia flowchart
Coombs test NEGATIVE
Normal spleen
Spleen palpable
Congenital
Acquired
-Hypersplenism -Thalassemia -Abnormal Hemoglobin
(eg. HbSC)
-Drug related -Enzyme defect -Membrane
defect -Abnormal HgB

• Microangiopathic hemolytic anemia
• Paroxysmal nocturnal hemoglobinuria

22
Case Initial impression

• Paroxysmal nocturnal hemoglobinuria
• Myelodysplastic syndrome
• Leukemia

23
Case Course in the wards

• On admission CBC
• Hgb 9.5
• Hct 31.8
• RBC 2.9
• WBC 3,550
• Eosinophils 1.00
• Segmenters/Neutrophils 61.00
• Lymphocytes 31.00
• Monocytes 7.00
• Plt Ct 173,000

24
Course in the wards

• 1st Hospital Day
• Underwent BMA
• Flow cytometry (sent to SLMC)
• PNH Panel report (By Flow Cytometry Peripheral
  Blood)
• Discharged
• Prednisone 20mg OD every other day
• Folic acid 500mg OD

25
PNH Panel Report
Marker Lineage of cells commonly labelled labelled Reference Range
CD 55 Broad (decay accelerating factor DAF for C3 and C5, GPI-linked deficiency leads to paroxysmal nocturnal hemoglobinuria PNH) 1.98 65.42-81.82
CD59 Protectin or Membrane Inhibitor of Reactive Lysis (MIRL) 6.23 73.24-87.00
26
Case Diagnosis

• Paroxysmal Nocturnal Hemoglobinuria

27
Paroxysmal Nocturnal Hemoglobinuria (PNH)

• acquired chronic non-immunologic hemolytic anemia
  
• arises from a somatic mutation in a hematopoietic
  stem cell (PIG-A)
• Deficient in surface protein normally attached to
  cell membrane by a GPI anchor
• renders the red cells highly susceptible to
  complement mediated lysis resulting in the
  characteristic hemolysis.

28
History

• Investigator Year
  Contribution
• Gull 1866 Described nocturnal and
  paroxysmal nature of intermittent
  haematinuria in a young tanner.
• Strubing 1882 Distinguished PNH from
  paroxysmal cold haemoglobinuria and march
  haemoglobinuria.
• Attributed the problem to the red cells.
• van den Burgh 1911 Red cells lysed in
  acidified serum. Suggested a
• role for complement.
• Enneking 1928 Coined the name
  paroxysmal nocturnal haemoglobinuria.
• Marchiafava 1928- Described perpetual
  hemosiderinemia in absence of
• and Micheli 1931 hemolysis. Their names became
  eponymous for PNH in Europe.

29
History

• Ham 1937-1939 Identified the role of
  complement in lysis of PNH red cells.
  Developed the acidified serum test, also
  called the Ham test, which is still used to
  diagnose PNH.
  Demonstrated that only a portion of PNH red
  cells are abnormally sensitive to complement.
• Davitz 1986 Suggests defect in membrane
  protein anchoring system responsible
• Hall Rosse 1996 Flow cytometry for the
  diagnosis of PNH

30
Epidemiology

• Rare disease
• frequency unknown
• thought to be on the same order as aplastic
  anemia (2-6 per million)
• Median age at diagnosis
• 35 yrs
• PNH reported at extremes of age

31
Epidemiology

• FemaleMale ratio 1.21.0
• No increased risk of PNH in patient relatives
• Median Survival after diagnosis 10-15 yrs

32
Pathogenesis The Defect

• Defect - Somatic mutation of PIG-A gene
  (phosphatidylinositol glycan complementation
  group A) located on the X chromosome in a clone
  of a hematopoietic stem cell
• gt100 mutations in PIG - A gene known in PNH
• The mutations (mostly deletions or insertions)
  generally result in stop codons - yielding
  truncated proteins which may be non or partially
  functional

33
PNH Defect
Hillmen and Richards, Br J Haematol, 2000
34
Dual Pathogenesis Hypothesis
Hillmen and Richards, Br J Haematol, 2000
35
Molecular lesion

• Frameshift mutations
• Type III cells
• Total absence
• Missense point mutations
• Small amounts of GPI anchor proteins
• Partial expression/residual activity
• Type II cells

Richards, et. al. Cytometry 42 223-233 (2000)
36
Pathogenesis Functional consequences of lack of
GPI linked proteins

• CD55 (decay accelerating factor) inhibits the
  formation or destabilizes complement C3
  convertase (C4bC2a)
• CD59 (membrane inhibitor of reactive lysis,
  protectin, homologous restriction factor)
  Protects the membrane from attack by the C5-C9
  complex
• Inherited absences of both proteins in humans
  have been described
• Most inherited deficiencies of CD55 - no distinct
  clinical hemolytic syndrome
• Inherited absence of CD59 - produces a clinical
  disease similar to PNH with hemolysis and
  recurrent thrombotic events

37
CD55 inhibits assembly, regulating complement
cascade at C3 step
38
CD59 limits polymerization of C9 in membrane
C5b-9 complex
39
Missing proteins of importance

• Complement regulating proteins
• CD59 (aka MAC inhibitor/protectin)
• Homologous restriction factor (HRF)
• CD55 (aka decay accelerating factor)
• Thrombosis regulating proteins
• CD87 (aka urokinase-type plasminogen activator
  receptor)

40
Clinical manifestations of PNH

• Highly variable and dependent upon the size of
  the abnormal clonal population in any individual
• Hemolysis
• mild to very brisk
• dependent upon
• size of abnormal clone (1-gt90)
• content of complement defense proteins
  (PNHII/III)
• presence of concomitant infection or other factor
  activating complement

41
Clinical Manifestations Hemolysis

• chronic hemolysis with acute exacerbations
  (hallmark)
• only 1/3 exhibit hemolysis at diagnosis

42
Clinical manifestations Hemolysis

• Recurrent attacks of intravascular hemolysis are
  usually associated with
• Hemoglobinuria
• Abdominal pain
• Dysphagia
• Impotence
• Dependent upon
• Size of abnormal clone (1-gt90)
• Content of complement defense proteins (PNH
  II/III)
• Presence of concomitant infection or other factor
  activating complement

43
Clinical manifestations Thrombosis

• Hepatic vein most common
• common cause of fatality
• Cerebral vein thrombosis
• sagittal sinus in particular
• Abdominal veins
• Dermal veins
• Pulmonary embolism - unusual

44
Clinical manifestations Cytopenia

• Relative/absolute bone marrow failure
• present to some degree in all patients
• relative granulocytopenia/thrombocytopenia
• decreased capacity to form myeloid colonies

45
Clinical Features Dilemma in diagnosis

• Variable expression of symptoms often causes
  considerable delay in the diagnosis

46
Presenting features in 80 patients with PNH
Signs and Symptoms of Px ()
Symptoms of anemia 28 (35)
Hemoglobinuria 21 (26)
Hemorrhagic signs and symptoms 14 (18)
Aplastic anemia 10 (13)
Gastrointestinal symptoms 8 (10)
Hemolytic anemia and jaundice 7 (9)
Iron-deficiency anemia 5 (6)
Thrombosis or embolism 5 (6)
Infections 4 (5)
Neurologic signs and symptoms 3 (4)
Dacie and Lewis, 1972
47
Laboratory Evaluation of PNH

• Acidified Serum Test (Ham Test 1939)
• Acidified serum activates alternative complement
  pathway resulting in lysis of patients rbcs
• May be positive in congenitial dyserythropoietic
  anemia
• Still in use today
• Sucrose Hemolysis Test (1970)
• 10 sucrose provides low ionic strength which
  promotes complement binding resulting in lysis of
  patients rbcs
• May be positive in megaloblastic anemia,
  autoimmune hemolytic anemia, others
• Less specific than Ham test

48
Diagnosis HAM test

• Lysis of PNH cells by activated complement
• Semi-quantitative
• Cannot differentiate Type III from Type II
• Cannot provide information on cell lineages other
  than red cells

49
Laboratory Evaluation of PNH

• PNH Diagnosis by Flow Cytometry (1986)
• Considered gold standard for diagnosis of PNH
  (1996)
• Detects actual PNH clones lacking GPI anchored
  proteins
• More sensitive and specific than Ham and sucrose
  hemolysis test

50
PNH Diagnosis by Flow Cytometry

• Flow Cytometry is method of choice
• More studies are needed to better define whether
  the type (I, II, or III), cell lineage, and size
  of the circulating clone can provide additional
  prognostic information.
• Theoretically - should be very valuable

51
Flow cytometry

• Extent of PNH clone
• 2 classifications of PNH
• Hemolytic PNH
• Overt episodes
• Venous thrombosis in 50
• Hypoplastic PNH
• No overt hemolysis
• 10 die due to aplastic anemia

Richards, et. al. Cytometry 42 223-233 (2000)
52
Flow cytometry Red cells

• CD55 and CD59
• Negative or partially deficient cells must be
  present for both in order to establish a
  diagnosis of PNH
• Provides clearest discrimination between types
• May predict clinical phenotype

53
Flow cytometry Granulocytes

• CD55/CD16 or CD59/CD16 plus an additional
  antibody - CD15 or CD33
• Percentage of PNH granulocytes reflects most
  accurately size of PNH clone
• Size of PNH clone is a determinant of the
  clinical phenotype of individual patients
• Serial studies allow prediction of prognosis

Richards, et al Cytometry 42 223-233 (2000)
54
Flow cytometry Serial monitoring

• Clinical relevance is unclear.
• Size of PNH clones if not constant with time

British Journal of Hematology. 2000. 108 470-479
55
From Purdue Cytometry CD-ROM vol3 97
56
Natural History of PNH

• Long term study of 80 patients with PNH seen at
  one institution between 1940 and 1970
• Results
• median age at diagnosis 42 (16-75)
• median survival 10 years
• 28 survived more than 25 years
• 39 had one or more episodes of venous thrombosis
• 12 experienced spontaneous clinical recovery
• leukemia did not develop in any of the patients

Hillmen et al, NEJM, 1995
57
Clinical features Sequelae

• Major cause of death
• venous thrombosis
• complications from progressive pancytopenia
• 25 of PNH patients survive gt25 years - one half
  of these go on to spontaneous remission
• Remission patients
• hematological values revert to normal
• no PHN rbcs or granulocytes detected
• PNH lymphocytes - still detected but no clinical
  consequence
• Higher incidence of acute leukemia (6)
• preleukemic condition most likely bone marrow
  failure not PNH

58
Clinical features Sequelae

• Significant proportion of patients survive for
  prolonged periods (?25 over 25 years)
• ?15 will experience recovery from PNH with no
  sequelae attributable to their disease

Hillmen, et al, 1995
59
Sites of thrombosis in PNH
Sites and Types of Thrombosis Number of patients
Intraabdominal
Hepatic vein 8
Inferior vena cava 3
Mesenteric vein 4
Splenic vein 1
Hillmen et al, NEJM, 1995
60
Sites of thrombosis in PNH
Sites and Types of Thrombosis Number of patients
Other venous sites
Cerebral 4
Pulmonary embolism 9
Deep vein 7
Superficial 3
Arterial
Myocardial infarction 6
Cerebrovascular accident 2
Hillmen et al, NEJM, 1995
61
Natural history of PNH
Causes of death in 60 patients with PNH of patients
Probably related to PNH
Venous thrombosis
Hepatic Vein 7
Inferior vena cava 1
Cerebral vein 1
Mesenteric vein 2
Pulmonary embolism 3
Hemorrhage
Gastrointestinal 6
Hillmen et al, NEJM, 1995
62
Natural history of PNH
Causes of death in 60 patients with PNH of patients
Subarachonoid 3
Intracerebral 2
Miscellaneous
Liver failure 2
Intraabdominal event 1
Probably unrelated to PNH
Arterial thrombosis
Myocardial infarction 2
Cerebrovascular accident 6
Hillmen et al, NEJM, 1995
63
Natural history of PNH
Causes of death in 60 patients with PNH of patients
Bronchopnuemonia 3
COPD plus cor pulmonale 2
Cardiac tamponade 1
Constrictive pericarditis 1
Renal failure 1
Amyloidosis 1
Lymphoma 1
Bronchial carcinoma 1
Unknown 12
Hillmen et al, NEJM, 1995
64
Natural History of PNH
Hillmen et al, NEJM, 1995
65
Relationship to aplastic anemia (AA)

• AA described as pancytopenia with nonfunctioning
  bone marrow. Cytopenia in one or all cell
  lineages also common to PNH
• High percentage of patients with AA develop
  clinical PNH or have lab evidence of PNH
  abnormality at some point (52)
• Have lower risk of thrombosis

Fujioka Asai, 1989 Dunn, et al, 1991
66
Venous thrombosis

• Anticoagulation
• Recommended for patients with large PNH
  populations
• With no contraindications to anticoagulation

British Journal of Haematology 108 470-479
67
Pancytopenia

• 10 will die from aplastic anemia
• Good response to immunosuppresion
• Antilymphocyte globulin
• Cyclosporin A

Ebenbichler et al, 1996 Stoppa, et al,
1996 Paquette et al, 1997 Schubert et al. 1997
68
PNH and Leukemia

• Incidence similar to the risk of AML in aplastic
  anemia (5)
• AA predisposes to clonal hematopoietic disorders
  (AML, PNH, MDS)
• PNH clone does not increase the risk of AML/MDS
• Not preleukemic

British Journal of Haematology 108_470-179
69
Therapy

• Bone Marrow Transplantation
• Only curative treatment
• Immunosuppressive therapy
• Antilymphocyte globulin /or cyclosporine A
• Does not alter proportion of PNH hemopoiesis
• Growth Factors
• Some improvement
• no evidence that normal clones respond better
  than PNH clones

70
Treatment

• Supportive, prevent complement activation
• Prednisone
• Therapy for/prevention of thrombosis
• thrombolytics acutely
• anticoagulation
• Stimulate hematopoiesis
• G-CSF
• Immunosuppression
• ATG
• Cyclosporine

71
Stem Cell Transplantation in PNH
Flotho et al, Br J Haematol, 2002
72
Stem Cell Transplantation in PNH

• Summary of single institution trials
• Approximately 12 reported
• Number of patients ranges from 1-16
• Survival rates typically higher (58-100)
• Likely high degree of reporting bias, based on
  outcomes seen using registry data

73
Results

• Sustained engraftment 77
• Graft failure 17
• Grade 2-4 acute GVHD 34
• Chronic GVHD 33
• Causes of death
• graft failure (7), int. pneumonitis (4), GVHD
  (3), infection (3), ARDS (2), hemorrhage (1)

IBMTR Data Saso et al, Br J Haematol, 1999
74
Stem Cell Transplantation in PNH

• EBMT Database (unpublished)
• 46 transplants using HLA-ID sibs reported from
  1979 to 1997
• Median age at diagnosis 29 (10-46)
• Median interval from diagnosis to BMT 794 days
  (30-8680)
• Actuarial 5 yr survival 52
• Main cause of death acute GVHD

75
Stem Cell Transplantation in PNH

• Conclusions from reported series
• BMT may cure 50-60 of selected patients with
  HLA-identical siblings
• Most patients transplanted have been lt 30 years
  of age
• Regimen related toxicity and GVHD remain
  significant hurdles
• Role of alternative donor transplants unclear,
  though initial reports are not encouraging except
  in pediatric population

76
Prognosis Based on Management
Luzzatto, Haematologica, 2001
77
PNH Management Guidelines
Luzzato, ASH, 2001
78
Updates

• Eculizimab (Soliris)
• 1st drug to be approved for the treatment of PNH
• Complement inhibitor
• Humanized monoclonal antibody against terminal
  complement protein C5

79
Update Eculizimab

• 600mg IV Q7 days x 4 doses, then
• 900mg IV, 7 days after 4th dose, then
• 900mg IV Q14 days
• Must give meningococcal vaccine 2 weeks before
  treatment

80
Thank you !