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178-P
160
178-P
Abstracts / Human Immunology 73 (2012) 49–167
SBT AND RSSO FAIL TO DETECT HETEROZYGOUS HLA LOCI IN TWO AML PATIENTS. Cheryl L. Thompson 1, Donna Phelan 1, Gerald Morris 2, T. Mohanakumar 3. 1 HLA Laboratory, Barnes-Jewish Hospital, St. Louis, MO, USA; 2 Department of Pathology and Immunology, Washington University, St. Louis, MO, USA; 3 Department of Surgery, Washington University, St. Louis, MO, USA. Aim: We present the cases of two AML patients HLA typed by rSSO and SBT as homozygous, and subsequently typed as heterozygous by SSP. Methods: Potential bone marrow transplant recipients were typed for HLA-A, -B, -C, -DR, -DQ by low resolution rSSO (LabType, One Lambda) and high resolution SBT (SeCore, Life Technologies), using DNA extracted (Qiagen EZ, 20 ng/ll) from EDTA anticoagulated peripheral blood. SSP (Olerup) was used as a secondary method for clarification. Results: Patient #1, a 71y.o. female caucasian in AML blast crisis (86% blasts in peripheral blood), was typed by rSSO and SBT. Results at the A locus were discrepant, DR-DQ association was uncommon, and a possible homozygous B⁄44:27 warranted confirmation. Further SSP testing to investigate the discrepancy revealed a full heterozygous HLA typing. Buccal cell DNA typed by SBT confirmed heterozygous DQB1⁄05:02 and ⁄06:04 [Table 1]. Patient #2, a 68 y.o. female diagnosed with AML, was typed by rSSO and SBT. Discrepancies were detected at A and B loci, and the C locus appeared homozygous. SSP method revealed a heterozygous HLA-A, -B, -C typing [Table 2]. Conclusions: These two cases reinforce the importance of close scrutiny when reviewing HLA molecular typing results. HLA heterozygous patients may present as homozygous from peripheral blood with large proportions of leukemic cells. We suggest using multiple methods and/or DNA from sources other than peripheral blood to confirm homozygous HLA loci in this patient population.
Table 1 Patient #1 Peripheral Blood DNA Typing Results Method
A
B
C
DRB1
DQB1
rSSO SBT SSP
⁄
⁄
⁄
⁄
⁄
02,⁄03 ⁄ 03:01/01N ⁄ 02:01,⁄03:01
44 ⁄ 44:27 ⁄ 40:01,⁄44:27
13,⁄16 ⁄ 13:02, ⁄16:01 ⁄ 13:02,⁄16
07 ⁄ 07:04/11 ⁄ 03:04, ⁄07:04
⁄ ⁄
05 05:02 05:02,⁄06:04
Table 2 Patient #2 Peripheral Blood DNA Typing Results Method
A
B
C
DRB1
DQB1
rSSO SBT SSP
⁄
⁄
⁄
⁄
⁄
02, ⁄32 ⁄ 02:01 ⁄ 02, ⁄32:01
40, ⁄44 ⁄ 44:02/19N ⁄ 40:01, ⁄44
05 ⁄ 05:01 ⁄ 03, ⁄05
04, ⁄13 ⁄ 04:02, ⁄13:01/117 NT
03, ⁄06 03:02, ⁄06:03 NT ⁄
178-P
Abstracts / Human Immunology 73 (2012) 49–167
SBT AND RSSO FAIL TO DETECT HETEROZYGOUS HLA LOCI IN TWO AML PATIENTS. Cheryl L. Thompson 1, Donna Phelan 1, Gerald Morris 2, T. Mohanakumar 3. 1 HLA Laboratory, Barnes-Jewish Hospital, St. Louis, MO, USA; 2 Department of Pathology and Immunology, Washington University, St. Louis, MO, USA; 3 Department of Surgery, Washington University, St. Louis, MO, USA. Aim: We present the cases of two AML patients HLA typed by rSSO and SBT as homozygous, and subsequently typed as heterozygous by SSP. Methods: Potential bone marrow transplant recipients were typed for HLA-A, -B, -C, -DR, -DQ by low resolution rSSO (LabType, One Lambda) and high resolution SBT (SeCore, Life Technologies), using DNA extracted (Qiagen EZ, 20 ng/ll) from EDTA anticoagulated peripheral blood. SSP (Olerup) was used as a secondary method for clarification. Results: Patient #1, a 71y.o. female caucasian in AML blast crisis (86% blasts in peripheral blood), was typed by rSSO and SBT. Results at the A locus were discrepant, DR-DQ association was uncommon, and a possible homozygous B⁄44:27 warranted confirmation. Further SSP testing to investigate the discrepancy revealed a full heterozygous HLA typing. Buccal cell DNA typed by SBT confirmed heterozygous DQB1⁄05:02 and ⁄06:04 [Table 1]. Patient #2, a 68 y.o. female diagnosed with AML, was typed by rSSO and SBT. Discrepancies were detected at A and B loci, and the C locus appeared homozygous. SSP method revealed a heterozygous HLA-A, -B, -C typing [Table 2]. Conclusions: These two cases reinforce the importance of close scrutiny when reviewing HLA molecular typing results. HLA heterozygous patients may present as homozygous from peripheral blood with large proportions of leukemic cells. We suggest using multiple methods and/or DNA from sources other than peripheral blood to confirm homozygous HLA loci in this patient population.
Table 1 Patient #1 Peripheral Blood DNA Typing Results Method
A
B
C
DRB1
DQB1
rSSO SBT SSP
⁄
⁄
⁄
⁄
⁄
02,⁄03 ⁄ 03:01/01N ⁄ 02:01,⁄03:01
44 ⁄ 44:27 ⁄ 40:01,⁄44:27
13,⁄16 ⁄ 13:02, ⁄16:01 ⁄ 13:02,⁄16
07 ⁄ 07:04/11 ⁄ 03:04, ⁄07:04
⁄ ⁄
05 05:02 05:02,⁄06:04
Table 2 Patient #2 Peripheral Blood DNA Typing Results Method
A
B
C
DRB1
DQB1
rSSO SBT SSP
⁄
⁄
⁄
⁄
⁄
02, ⁄32 ⁄ 02:01 ⁄ 02, ⁄32:01
40, ⁄44 ⁄ 44:02/19N ⁄ 40:01, ⁄44
05 ⁄ 05:01 ⁄ 03, ⁄05
04, ⁄13 ⁄ 04:02, ⁄13:01/117 NT
03, ⁄06 03:02, ⁄06:03 NT ⁄