1. Neonatal Alloimmune Thrombocytopenia: Diagnosis, Management, Investigations
Donald M. Arnold, MD MSc
Medical Director, Platelet Immunology Laboratory
McMaster University
Transfusion Medicine Residency Teaching
June 11, 2008

2. Neonatal Alloimmune Thrombocytopnia
Definition:
Thrombocytopenia in a fetus or neonate caused by maternal antiplatelet alloantibodies, directed against a fetal platelet alloantigen, inherited from the father.

3. Neonatal Alloimmune Thrombocytopenia
Most common cause of severe TCP in infant
Most common cause of ICH in term newborns
First pregnancies, without warning
Otherwise healthy babies

4. NAT Fetus inherits platelet antigens from father
Transplacental passage of fetal platelet antigens
Mother forms IgG alloAbs that cross placenta
Maternal alloAb react with fetal platelets
AlloAb-sensitized platelets are cleared in RE system

6. NAT and HDN NAT HDN Affected cells Most common antigen
Affected pregnancy
Timing of sensitization
Affected Infant
Affected fetus
Main risk factor
Treatment
Prevention
Efficacy of prevention

7. NAT and HDN NAT HDN Affected cells Platelets Red blood cells
Most common antigen
Affected pregnancy
Timing of sensitization
Affected Infant
Affected fetus
Main risk factor
Treatment
Prevention
Efficacy of prevention

8. NAT and HDN NAT HDN Affected cells Platelets Red blood cells
Most common antigen
HPA-1a
Rh-D
Affected pregnancy
Timing of sensitization
Affected Infant
Affected fetus
Main risk factor
Treatment
Prevention
Efficacy of prevention

9. NAT and HDN NAT HDN Affected cells Platelets Red blood cells
Most common antigen
HPA-1a
Rh-D
Affected pregnancy
First
Second +
Timing of sensitization
Affected Infant
Affected fetus
Main risk factor
Treatment
Prevention
Efficacy of prevention

10. NAT and HDN NAT HDN Affected cells Platelets Red blood cells
Most common antigen
HPA-1a
Rh-D
Affected pregnancy
First
Second +
Timing of sensitization
16 weeks onwards
At birth, or during a procedure
Affected Infant
Affected fetus
Main risk factor
Treatment
Prevention
Efficacy of prevention

11. NAT and HDN NAT HDN Affected cells Platelets Red blood cells
Most common antigen
HPA-1a
Rh-D
Affected pregnancy
First
Second +
Timing of sensitization
16 weeks onwards
At birth, or during a procedure
Affected Infant
TCP, bleeding
Hemolysis, jaundice, kernicterus
Affected fetus
Main risk factor
Treatment
Prevention
Efficacy of prevention

12. NAT and HDN NAT HDN Affected cells Platelets Red blood cells
Most common antigen
HPA-1a
Rh-D
Affected pregnancy
First
Second +
Timing of sensitization
16 weeks onwards
At birth, or during a procedure
Affected Infant
TCP, bleeding
Hemolysis, jaundice, kernicterus
Affected fetus
ICH
Hydrops fetalis
Main risk factor
Treatment
Prevention
Efficacy of prevention

13. NAT and HDN NAT HDN Affected cells Platelets Red blood cells
Most common antigen
HPA-1a
Rh-D
Affected pregnancy
First
Second +
Timing of sensitization
16 weeks onwards
At birth, or during a procedure
Affected Infant
TCP, bleeding
Hemolysis, jaundice, kernicterus
Affected fetus
ICH
Hydrops fetalis
Main risk factor
Previously affected infant
Rh-negative mothers
Treatment
Prevention
Efficacy of prevention

14. NAT and HDN NAT HDN Affected cells Platelets Red blood cells
Most common antigen
HPA-1a
Rh-D
Affected pregnancy
First
Second +
Timing of sensitization
16 weeks onwards
At birth, or during a procedure
Affected Infant
TCP, bleeding
Hemolysis, jaundice, kernicterus
Affected fetus
ICH
Hydrops fetalis
Main risk factor
Previously affected infant
Rh-negative mothers
Treatment
Supportive
Prevention
Efficacy of prevention

15. NAT and HDN NAT HDN Affected cells Platelets Red blood cells
Most common antigen
HPA-1a
Rh-D
Affected pregnancy
First
Second +
Timing of sensitization
16 weeks onwards
At birth, or during a procedure
Affected Infant
TCP, bleeding
Hemolysis, jaundice, kernicterus
Affected fetus
ICH
Hydrops fetalis
Main risk factor
Previously affected infant
Rh-negative mothers
Treatment
Supportive
Prevention
IVIG
Anti-D
Efficacy of prevention

16. NAT and HDN NAT HDN Affected cells Platelets Red blood cells
Most common antigen
HPA-1a
Rh-D
Affected pregnancy
First
Second +
Timing of sensitization
16 weeks onwards
At birth, or during a procedure
Affected Infant
TCP, bleeding
Hemolysis, jaundice, kernicterus
Affected fetus
ICH
Hydrops fetalis
Main risk factor
Previously affected infant
Rh-negative mothers
Treatment
Supportive
Prevention
IVIG
Anti-D
Efficacy of prevention ?
99%

17. Incidence: 1 in 1,000 to 1 in 2,000 births
NAT
Incidence: 1 in 1,000 to 1 in 2,000 births N
Incidence
Burrows and Kelton, 1993
15,932
1 in 1,700
Uhrynowska, 2000
24,101
1 in 2,400
Turner, 2005
26,000
1 in 5,000
Kjeldsen-Kragh, 2007
100,448

18. Severity of thrombocytopenia
<150, in 100
< 50, in 400
<20, NAT
Burrows, Kelton 1993

19. Predictors of Severity of NAT
History of NAT in a sibling Murphy, 2006
Worse with subsequent pregnancies Bussel, 1997
Worse with increased gestational age Kaplan, 1988
ICH in a previous sibling – greater severity Bussel, 1997

20. Treatment of affected neonates
Without warning
Prompt recognition
IVIg (2g/kg); Effective in 75% of cases
Platelet transfusions:
Antigen-negative (maternal) platelets
Random-donor platelets

21. = unmatched PLTs; = matched PLT; ∆ = IVIg; □= steroids
Kiefel Blood, 2006

22. Antenatal treatment (prevention)
IVIg 1g/kg/wk (2g/kg/wk for refractory)
IVIg + corticosteroids
Intrauterine platelet transfusions
Fetal blood sampling (FBS)

23. Outcome: increase in fetal platelet count
Risk-based treatment
High Risk (n= 40) Standard Risk (n= 39)
(previous ICH or PLT<20) (neither)
IVIg vs. IVIg + pred IVIg vs. pred
(1g/kg/wk) (1mg/kg) (1g/kg/wk) (0.5mg/kg)
PUBS (20 wks, repeat 3-8 wks)
Outcome: increase in fetal platelet count
Berkowitz, Bussel, 2006

24. Risk-based treatment HIGH RISK Mothers Pre 2-8 wks Birth
(platelet count)
Pre 2-8 wks Birth
IVIg alone* 7, , ,000
IVIg + pred , , ,000
* One ICH
Berkowitz, Bussel, 2006

25. STANDARD RISK Mothers*
Risk-based treatment
STANDARD RISK Mothers*
(platelet count)
Pre wks
IVIg alone >20, ,000
Pred alone >20, ,000
* 2 fetal deaths, 2 ICH
Berkowitz, Bussel, 2006

26. Risk-based treatment 11 SERIOUS COMPLICATIONS OF 175 PUBS (6%)
- 1 Fetal Death
- 9 Emergency C-Sections (14%)

27. Testing for NAT

28. Antigens on Platelets Blood group antigens (ABO) Common antigens (HLA)
Platelet specific antigens

29. Platelet Specific Antigens
Nomenclature
Chosen name, or name related to individual with antigen (PlA1,Zav, Gov).
Current recommendation: All antigens designated as HUMAN PLATELET ANTIGENS (HPA).
Antigens numbered in order of discovery.
Higher frequency allele is “a”.
Example: PLA1 = HPA-1a; PLA2 = HPA-1b

30. Platelet Specific Antigens
To date, 22 platelet specific alloantigens identified, with 6 diallelic system (HPA-1, 2, 3, 4, 5, 15). Almost all are associated with a single nucleotide substitution.

31. Major clinically important platelet antigens in NAT
HPA-1a PLA1
HPA-5a Br-b, Zav-b
HPA-5b Br-a, Zav-a
HPA-15a Gov-b
HPA-15b Gov-a
HPA-4a (Asian population) Pen-a
Associated with up to 5% of all NAT
HPA-3a Bak-a
HPA-2a Ko-b
HPA-2b Ko-a
Implicated in NAT, but occur rarely in the population
HPA-6b Ca-a, Tu-a
HPA-8b Sr-a
HPA-9b Max-a
HPA-13b Sit-a
Rarely implicated in NAT
HPA-1b PLA2
HPA-3b Bak-b

33. Gene discrepancy is not NAT!
Key Message
Gene discrepancy is not NAT!
“Genetic NAT” is 10 times more common than
actual NAT”.

34. Testing

36. Platelet Phenotyping

37. Phenotyping and Antibody Identification
Radioimmunoprecipitation

39. Investigation of Neonatal Alloimmune Thrombocytopenia
Platelet Antigen Typing:
Genotyping – Maternal and Paternal blood samples
Genotyping – Direct and Cultured amniocytes
Phenotyping – Maternal and Paternal samples
Platelet Antibody Investigation:
Radioimmunoprecipitation
Antigen Capture ElA
Flow Cytometry

40. Phenotype, antibody detection (RIP)
Mom: HPA-1a neg
Dad: HPA-1a pos
Mom: Anti-1a

41. Genotype (PCR)

42. Genotype (SSP)