Application of a digital PCR method for WT1 to myeloid neoplasms in CR and deep ELN WT1 molecular response (< 10 copies).

Bone marrow WT1 mRNA levels assessed by the ELN method are useful to establish prognostic correlations in myeloid malignancies treated with chemotherapy or hematopoietic stem cell transplantation (HCT). Those patients with WT1 levels below ten copies have a good outcome. However, some of these patients relapse. To further characterize this group of cases, we applied a new and sensitive digital (ddPCR) WT1 method. A consecutive series of 49 patients with treated myeloid malignancies and with an ELN WT1 quantitation of < 10 copies were included in the study. All cases (47 AML and 2 MDS) have received intensive chemotherapy or HCT. One to four micrograms of total RNA were retrotranscribed to obtain ≥ 10,000 ABL1 copies using the ELN protocol. Only those cases with a good quality cDNA were used in the ddPCR WT1 test. The ddPCR Gene Expression WT1 Assay of Bio-Rad© was used to perform the PCR amplification, and the microdroplets were quantified in the Bio-Rad's QX200 droplet reader. Eighteen patients showed a negative WT1 ddPCR assay (0 copies/µl), whereas 31 cases were positive (results ranged from 1 to 15.2 copies/µl). Survival analysis showed statistically significant differences in terms of OS between both groups, 83 ± 8% vs. 46 ± 9% (p = 0.024). A statistically significant correlation was also found between ddPCRWT1 results and CD123+ cell number detected by flow cytometry (p = 0.024). Larger series of patients tested with the current ddPCRWT1 method will solve whether it could be used to stratify patients with myeloid malignancies achieving deep WT1 molecular response (< 10 copies).

Cunninghamella bertholletiae infection (mucormycosis) in a patient with acute T-cell lymphoblastic leukemia.

Cunninghamella spp. are unusual opportunistic pathogens that have been identified with increased frequency in immunocompromised patients. Clinical infection by this fungus is almost always devastating and usually fatal. Infections with this group of organisms have been seen most frequently in patients with hematological malignancy. Here we report the case of a patient with acute leukemia who developed multiorganic failure as a consequence of hematological dissemination by Cunninghamella bertholletiae. The case highlights the mortality associated with this fungal infection in immunocompromised patients, confirms the risk factors associated with non-candida fungal infections and shows a clinical presentation mimicking myocardial infarct and cerebrovascular stroke.

The direct antiglobulin test in a hospital setting.

To evaluate the current use of the DAT in our hospital,we reviewed the charts of all patients who had a DAT performed in our laboratory. The collected data included DAT results and a previously completed laboratory evaluation of suspected hemolytic anemia. Four hundred sixty-three DATs were performed in our laboratory from April 1999 to October 2001. The DAT was negative in 434 (93.7%) cases and positive in 29 (6.3%) cases. A complete laboratory evaluation of suspected hemolytic anemia was seen in 179 (38.7%) cases. The incidence of a positive DAT was higher in the group of patients with > 2 signs of hemolysis (4/34 cases; 11.8%) than in the group of patients with