Donor lymphocyte Donor lymphocyte infusion as a treatment infusion as a treatment of a complete DiGeorge of a complete DiGeorge

syndromesyndromeK.ZdráhalováK.Zdráhalová22, E.Mejstříková, E.Mejstříková1,21,2,T.Kalina,T.Kalina1,21,2,P.Sedláček,P.Sedláček22, ,

A. Janda A. Janda11, H.Žižková, H.Žižková33, Z.Sieglová, Z.Sieglová33,A.Šedivá,A.Šedivá11, , J.BartůňkováJ.Bartůňková11, J.Starý, J.Starý22, P.Kobylka, P.Kobylka33 , P.Hubáček , P.Hubáček 2,42,4, ,

O.HrušákO.Hrušák1,21,2

22Department of Pediatric Hematology and Oncology, Department of Pediatric Hematology and Oncology, 11Department of Immunology, Department of Immunology,

33Institute of Hematology and Blood Transfusion,Institute of Hematology and Blood Transfusion,44Department of Pediatrics,Department of Pediatrics,Prague, Czech RepublicPrague, Czech Republic

Introduction - DiGeorge syndromeIntroduction - DiGeorge syndrome• the most common deletion syndrome in humans – the most common deletion syndrome in humans – • monoallelic microdeletion of monoallelic microdeletion of 22q11.222q11.2 (DiGeorge (DiGeorge and velo-cardio-facial sy, conotruncal anomaly)and velo-cardio-facial sy, conotruncal anomaly)

• variable phenotype even in pts with the same deletionvariable phenotype even in pts with the same deletion

• manifestation:manifestation: ("CATCH22")("CATCH22")• mainly heart defects; mainly heart defects; hypoparathyroidismhypoparathyroidism• thymic hypoplasia thymic hypoplasia immunodeficiency, facial immunodeficiency, facial dysmodysmorrphism, developmental and behavioral problemsphism, developmental and behavioral problems

• Deletion or interstiDeletion or interstittial deletion ial deletion of of 10p13 10p13 - - other rare cause of DiGeorge sy other rare cause of DiGeorge sy (type II)(type II)

Patient:Patient: male, born June 18 male, born June 18thth 2004, now 10 months 2004, now 10 months

Family history:Family history: healthy parents, 0 siblings healthy parents, 0 siblings

Personal history:Personal history: 1 1stst pregnancy, polyhydramnios pregnancy, polyhydramnios amniocentesis amniocentesis normal karyotype 46, XY normal karyotype 46, XY

term delivery, fetal hypoxia term delivery, fetal hypoxia Cesarian section, Cesarian section, resuscitation, intubation, artificial ventilationresuscitation, intubation, artificial ventilation

• esophageal atresia + tracheoesophageal fistula:esophageal atresia + tracheoesophageal fistula: D+2 operationD+2 operation• bilateral choanal atresia:bilateral choanal atresia: D+5 operation D+5 operation • congenital heart defects:congenital heart defects: D+13 operation D+13 operation

• Stigmatisation:Stigmatisation:* faceface* genitalsgenitals* eyeseyes* CNSCNS

• CHARGE associationCHARGE association ((CColoboma, oloboma, HHeart disease, eart disease, AAtresia choanae, tresia choanae, RRetarded growth + etarded growth + development and/or CNS anomalies, development and/or CNS anomalies, GGenital anomalies and/or enital anomalies and/or hypogonadism, hypogonadism, EEar anomalies and/or deafness)ar anomalies and/or deafness)

Problems:Problems:• recurrent infections, septicemiasrecurrent infections, septicemias• recurrent respiratory distress recurrent respiratory distress ventilation ventilation • apneas, irritability, states of altered consciousnessapneas, irritability, states of altered consciousness

Immunology:Immunology:• lymphocyte subsets in 2 months of age: lymphocyte subsets in 2 months of age: CD3+ 0%CD3+ 0% NK 40% NK 40% CD4+ 0% CD4+ 0% CD8+ 0% CD8+ 0% CD19+ 58%CD19+ 58%

absent T absent T cellscells

• response to mitogens: absentresponse to mitogens: absent

• MRI -MRI - absent thymusabsent thymus

0,0001

0,001

0,01

0,1

1

10

CD4 (10^9/L)CD8 (10^9/L)CD3 (10^9/L)CD19 (10^9/L)NK (10^9/L)

prior DLI 16.12.05prior DLI 16.12.05at 12.8.04at 12.8.04

Complete DiGeorge SyndromeComplete DiGeorge Syndrome

(diagnosis at 2 months)(diagnosis at 2 months)

• microdeletion 22q11 not foundmicrodeletion 22q11 not found

• prophylaxis started : prophylaxis started : cotrimoxazole + itraconazole + IVIGcotrimoxazole + itraconazole + IVIG

2 months of age2 months of age

1st donor lymphocyte infusion1st donor lymphocyte infusion

age 6 monthsage 6 months

unrelated donor from register, 8/10 unrelated donor from register, 8/10 (B, Cw)(B, Cw)

no conditioningno conditioning

no GVHD preventionno GVHD prevention

1x 101x 1066/kg CD3+; 0.2x 10/kg CD3+; 0.2x 1066/kg CD34+/kg CD34+

due to mistake non irradiated blood products due to mistake non irradiated blood products administered (administered (7 times prior 17 times prior 1stst DLI, 1 time after DLI, 1 time after 11stst DLI DLI))

1st donor1st donor lymphocytelymphocyte infusioninfusion (cont.)(cont.)

Complications:Complications: D+10:D+10:• isolated skin isolated skin aGVHD aGVHD ((stage 3, grade II)stage 3, grade II)• sepsis sepsis • cardiopulmonary cardiopulmonary instability instability • capillary leak sycapillary leak sy

Chimerism:Chimerism: D+10 donorD+10 donor detected detected

0,00010,0001

0,0010,001

0,010,01

0,10,1

11

1010

CD4 (10^9/L)CD4 (10^9/L)

CD8 (10^9/L)CD8 (10^9/L)

CD3 (10^9/L)CD3 (10^9/L)

Between 1st and 2nd DLI

after ATGafter ATG

00 1010 days post 1days post 1stst DLI DLI 88

after ATGafter ATG

aGVHDaGVHD, Capillary leak sy

Immunosupressive therapy:Immunosupressive therapy:• rATG FreseniusrATG Fresenius (25mg/kg 3x D+10, D+12, D+14)(25mg/kg 3x D+10, D+12, D+14)• CsACsA• corticosteroidscorticosteroids - MP (2mg/kg)- MP (2mg/kg)

• GVHD resolvedGVHD resolved

• corticosteroids corticosteroids - - 2 weeks 2 mg/kg, 1 week 1 mg/kg, 2 weeks 2 mg/kg, 1 week 1 mg/kg, 1 week 0.5 mg/kg, then tapered (D+35)1 week 0.5 mg/kg, then tapered (D+35)

• CsA continuedCsA continued

**********• D+33 last extubation!D+33 last extubation! - aged 7 months- aged 7 months

2nd donor lymphocyte infusion2nd donor lymphocyte infusion

age 7 months, D+36 after 1age 7 months, D+36 after 1stst DLI DLI

the same donorthe same donor

no conditioningno conditioning

prevention of GVHD: CsA (continued)prevention of GVHD: CsA (continued)

0.89x 100.89x 1066/kg CD3+/kg CD3+

2nd donor lymphocyte infusion2nd donor lymphocyte infusion (cont.) (cont.)

Complications:• D+27 EBV infection: (B cell proliferation, oligoclonality, IgM; no clinical manifestation)

• withdrawal of CsA

• Rituximab (375 mg/m2)

• proliferation of CD8+ activated T cells started

CD19CD1900

0,0010,001

0,010,01

0,10,1

11

1010

CD4CD4

CD8CD8

EBVEBVrituximabrituximab

00 days post days post 22ndnd DLI DLI

2727

10^

9/L

10^

9/L CD3CD3

1st DLI1st DLIacute GVHDacute GVHD

d 0

d 8

d 10

d 19

d 34

2nd DLI2nd DLI

d 1

0d

10

d 2

5d

25

EBV infection and prior rituximab EBV infection and prior rituximab

d 4

1d

41

d 5

5d

55

proliferation of activated T cells, proliferation of activated T cells, severe liver GVHDsevere liver GVHD

last non last non irradiated irradiated trftrf

NNo proof of engraftment of non irradiated blood transfusionso proof of engraftment of non irradiated blood transfusions

chimerism in non separated chimerism in non separated blood: recipient mainly, donor blood: recipient mainly, donor detecteddetected

chimerism in FACSorted chimerism in FACSorted T lymphocytes CD3+ T lymphocytes CD3+ (D+41 and D+55 CD4+ (D+41 and D+55 CD4+ and CD8+): mainly donorand CD8+): mainly donor

Chimerism after 1st and 2nd DLIChimerism after 1st and 2nd DLI

D+34:D+34: jaundice jaundice - bilirubin 4 mg/dL- bilirubin 4 mg/dL

dif.dg.:dif.dg.: • hepatic GVHDhepatic GVHD • EBV lymphoproliferationEBV lymphoproliferation (EBV in blood 0; in organs?)(EBV in blood 0; in organs?)

• hepatic infectionhepatic infection - not found- not found

******************

D+35D+35 2nd Rituximab2nd Rituximab (375 mg/m(375 mg/m22))

D+41 D+41 corticosteroidscorticosteroids (MP 1 mg/kg) hepatic aGVHD(MP 1 mg/kg) hepatic aGVHD

neutropenia - granulo 215!neutropenia - granulo 215!

D+55 D+55 preventive ATB, antimycoticspreventive ATB, antimycotics

D+45 - D+49: D+45 - D+49: agranulocytosisagranulocytosis (0 granulo) (0 granulo)

• corticosteroidscorticosteroids (MP 2 mg/kg)(MP 2 mg/kg)• CsACsA• G-CSFG-CSF 5x5x

D+52: granulo 3000D+52: granulo 3000 bilirubin 13.7 mg/dLbilirubin 13.7 mg/dL

isolated liver GVHD stage 3, grade IIIisolated liver GVHD stage 3, grade III

D+56: D+56: rATG FreseniusrATG Fresenius 1 1 dose, 25mg/kgdose, 25mg/kg

D+57: D+57: bilirubin 23.8 mg/dL bilirubin 23.8 mg/dL

isolated liver GVHD stage 4, grade IVisolated liver GVHD stage 4, grade IV

******************• corticosteroids corticosteroids (D+63 1.5 mg/kg, D+83 1 mg/kg)(D+63 1.5 mg/kg, D+83 1 mg/kg)• CsA continues CsA continues

Current status:Current status: • D+108 after 2nd DLI, age 10 months• bilirubin 7.6 mg/dL• CD8 activated T cells absolutely decreased• slight gradual psychomotor development

Patient aged 10 months

Thank you.