Our experience in PGD for HLA matching

The procedure was applied to 197 PGD cycles (150 for β-thalassaemia, 19 for ALL, 16 for DBA, 3 for Histiocytosis, 3 for Adrenoleukodystrophy , 2 for Duncan Sindrome, 1 for Chronic granulomatous disease, 1 for FANCG, 1 for Wiscott-Aldrich syndrome and 1 for Mannosidosis-Alpha) performed for 106 couples. A total of 1488 embryos were tested for HLA typing in combination with a genetic disease, and 329embryos for HLA typing only (Table 2). A total of 2492 blastomeres was analysed, in 2292 (92.0%) of which a successful amplification was obtained (Table 1). Two hundred blastomeres resulted in the amplification failure for all the markers/loci tested. The amplification efficiency of the individual STR markers on blastomeres ranged from 92.2 to 100%, with an overall amplification rate of 98.2%. The ADO rates varied between the different loci investigated, ranging from 0.0 to 10.8%

A reliable HLA haplotype was obtained in 2292/2292 (100%) of the blastomeres with positive PCR results.

Totally, 1704 (93.8%) embryos were diagnosed; however testing for chromosome 6 copy number revealed 187 (11.0%) embryos with aneuploidies, including a total of 38 (2.2%) trisomies, 143 (8.4%) monosomies and 6 (0.4%) uniparental dysomies, which affected the HLA matching diagnosis for these embryos (Table 4), leading to a conclusive diagnosis in only 1517 (89.0%) embryos.

In total, 311 (18.3%) embryos resulted to be HLA matched with the affected siblings. In the 159cycles of HLA matching combined a with single gene disorder, although the overall number of HLA-identical embryos was 259 (18.5%), only 196 (14.0%) resulted also in an unaffected genotype. In the couples undergoing preimplantation HLA typing without testing for a causative gene, 52 (17.2%) embryos appeared to be HLA compatible with the affected children. Recombination was found in 38 (2.2%) embryos.

Overall 211 embryos were transferred to the patients in 132 of the 197 cycles performed (1.6 + 0.7 on average), and 50 pregnancies were achieved. Eight pregnancies resulted only biochemical, 11 spontaneously miscarried and one resulted ectopic, and was then terminated.

Thirdy-one ongoing pregnancies were obtained. The ongoing pregnancy rate was 15.7% per cycle and 23.5% per transfer.

Fourteen pregnancies are still ongoing, while 3 twin and 14 singleton pregnancies, produced the birth of 20 healthy children, which were confirmed to be HLA identical to their affected sibling by blood samples HLA testing. Stem cells collected after delivery from the umbilical cords blood were transplanted to the affected siblings of 7 couples, resulting in a successful hematopoietic reconstruction for all patients.

Table 1: Multiplex PCR and HLA haplotyping results

 

Multiplex PCR results

HLA+PGD

%

HLA

%

Total

%

No. of cells analyzed

2077

 

415

 

2492

 

No. of cells with amplification failure of all markers (%)

163

7,8

37

8,9

200

8,0

No. of cells with a positive amplification of HLA markers(%)

1914

92,2

378

91,1

2292

92,0

HLA haplotyping results

 

 

 

 

 

 

No. of embryos analyzed

1488

 

329

 

1817

 

No. of embryos diagnosed (%) *

1402

94,2

302

91,8

1704

93,8

No. of embryos without HLA diagnosis (%) ?

230

15,5

70

21,3

300

16,5

No. of embryos with conclusive HLA diagnosis (%) $

1258

89,7

259

85,8

1517

89,0

No of HLA identical embryos (%)

259

18,5

52

17,2

311

18,3

No of HLA identical healthy embryos (%)

196

14,0

52

17,2

248

14,6

No. of HLA non-identical embryos (%)

1143

81,5

250

82,8

1393

81,7

No. of embryos with recombination (%)

32

2,3

6

2,0

38

2,2

Embryos without amplification (%)

86

5,8

27

8,2

113

6,2

 

 

 

 

 

 

 

No. of abnormal embryos (%)

144

10,3

43

14,2

187

11,0

Monosomy chromosome 6  (%)

109

7,8

34

11,3

143

8,4

Trisomy chromosome 6 (%)

30

2,1

8

2,6

38

2,2

UPD (%)

5

0,4

1

1,9

6

0,4


*(No. of embryos analyzed) ? (No. of embryos without diagnosis because of a total PCR failure on the single cell or both cells  biopsed) $(No. of embryos analyzed) ? (No. of abnormal embryos) ? (No. of embryos without diagnosis because of a total PCR failure on the unique cell biopsed) ? (No. of abnormal embryos)+(No. of embryos w/o diagnosis because of PCR failure on both biopsied cells or on the single cell biopsied)



Table 2: Results and outcome from 197 preimplantation HLA matching cycles


 
Clinical results

HLA+PGD

%

HLA

%

Total

%

No. of couples treated

88

 

18

 

106

 

Maternal age + SD

31.8 ? 5.2

 

37.5 ? 4.3

 

32.8 ? 5.4

 

No. of cycles performed

159

 

38

 

197

 

No. of oocytes retrieved

2394

 

533

 

2927

 

 

15,1

 

14,0

 

14,9

 

No. of oocytes injected (%)

1830

76,4%

413

77,5%

2243

76,6%

No. of oocytes fertilized (%)

1634

89,3%

358

86,7%

1992

88,8%

No. of embryos analyzed (%)

1488

91,1%

329

91,9%

1817

91,2%

Per cycle

9,4

 

8,7

 

9,2

 

No. of transfers

105

 

27

 

132

 

Per cycle

66,0%

 

71,1%

 

67,0%

 

No. of embryos transferred

165

 

46

 

211

 

Mean no. of embryos transferred

1.5 + 0.7

 

1.8 + 0.9

 

1.6 + 0.7

 

Outcome

 

 

 

 

 

 

No. of positive HCG pregnancies

41

 

9

 

50

 

UNEMBRYONIC

2

 

0

 

2

 

Biochemical

7

 

1

 

8

 

Clinical

34

 

8

 

42

 

Miscarried

8

 

3

 

11

 

Ectopic

1

 

0

 

1

 

Ongoing

26

 

5

 

31

 

Ongoing per cycle

16,4%

 

13,2%

 

15,7%

 

Ongoing per transfer

24,8%

 

18,5%

 

23,5%

 

No. of embryos implanted (fetuses)

38

 

8

 

46

 

Implantation rate

23,0

 

17,4

 

21,8

 

 

 

 

 

 

 

 

No. of pregnancies went to term

15

 

2

 

17

 

No. of babies born

18

 

2

 

20

 

No. of pregnancies still ongoing

11

 

3

 

14

 

No. of transplantations done

6

 

1

 

7

 

 

Table 3:
Number of families treated with distinct indications for PGD and/or HLA. ongoing pregnancies. life births and cord blood transplantations (CBTs).

Diseasese

Adrenoleukodystrophy

3

2

1

50,0

0

1

50,0

0

1

0

0

β-Thalassemia

150

81

38

38,8

7

31

31,6

8

10

13

15

5

Chronic granulomatous disease

1

1

1

100,0

0

1

100,0

0

0

1

2

0

Diamond Blackfan Anemia

16

3

4

36,4

1

3

27,3

2

1

0

0

0

Duncan Syndrome

2

1

0

0,0

0

0

0,0

0

0

0

0

FANCG

1

1

0

0

0

0

0

0

0

0

0

0

Histiocytosis

3

1

1

33,3

0

1

33,3

0

0

1

1

0

Leukemia

19

14

4

30,8

0

4

30,8

1

2

1

1

Mannosidosis-Alpha

1

1

0

0,0

0

0

0,0

0

0

0

0

0

Wiskott-Aldrich Syndrome

1

1

1

100,0

0

1

100,0

0

0

1

1

1

Total

197

106

50

37,9

8

42

31,8

11

14

17

20

7

 



Next :
Accuracy of PGD for HLA matching

Back to the top Print Page
 
  You are in the section
PGD Conditions

  Home Page
  info@preimplantationgeneticdiagnosis.it

Genoma Group Srl ? 2014 - All rights reserved