5,10-methylenetetrahydrofolate reductase (MTHFR) is a central regulatory enzyme in folate metabolism. A common functional polymorphism of the MTHFR gene occurs at C677T. The 677TT genotype produces an enzyme with only 30% of the activity of the wild-type (677CC) enzyme. The MTHFR polymorphism has been shown to affect the sensitivity of patients to folate-based drugs such as methotrexate (MTX) that is used for GVHD prophylaxis following allogeneic hematopoietic stem cell transplantation (HSCT).

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... o assess the significance of C677T genotypes in HSCT using MTX as a GVHD prophylaxis, we analyzed DNA from 159 patients with a hematological disease and their HLA-identical sibling donors using PCR-RFLP method. The frequencies of CC, CT and TT genotypes in the patients were 35%, 52% and 13%, respectively, and in the donors were 30%, 62% and 8%, respectively. There was no significant difference in the distribution patterns of the C677T genotypes between patients and donors (P 5 0.19). Multivariate analysis revealed a significantly lower incidence of grade I-IV acute GVHD in patients with the 677TT genotype (relative risk, 0.35; 95% confidence interval, 0.13-0.95; P 5 0.040) and a non-malignant disease (0.22; 0.05-0.89; 0.034). There was a significant association between a lower incidence of grade II-IV acute GVHD and the use of bone marrow for transplantation (0.32; 0.11-0.91; 0.032). There was no association between the incidence of acute GVHD and the donor C677T genotypes. We analyzed the incidence of acute GVHD in relation to the MTHFR genotype using the Kaplan-Meier method. The incidence of grade I-IV acute GVHD in the patients with 677TT genotype was significantly lower than in those with 677CC/CT genotype (19% versus 45%, P 5 0.035). There was a trend for a lower incidence of grade II-IV acute GVHD in patients with 677TT genotype compared with 677CC/CT genotype (5% versus 24%, P 5 0.077). The C677T genotypes in the patient or donor were not associated with the treatment-related mortality, relapse rate, relapse-free survival and overall survival. These results suggest that greater immunosuppression by MTX due to low MTHFR enzyme activity decreases the risk of acute GVHD in recipients of allogeneic HSCT. Further studies are needed to confirm that the MTHFR C677T polymorphism can predict the outcome of HSCT using prophylactic MTX to prevent GVHD. BAFF, a non-redundant cytokine produced by myeloid cells, plays a critical role in the normal homeostatic maintenance, activation and function of B cells. Elevated circulating levels of BAFF, however, have been observed in systemic autoimmune disorders and, in murine models, have been linked to a failure to delete auto-reactive B cells. We similarly observed elevated plasma BAFF levels in 77 patients in an ongoing NCI CGVHD natural history protocol, with a median of 2845 pg/ml (range 92 to 17058), as compared to 556 pg/ml (range 75 to 1834) in 18 normal donors. Furthermore, in a subset of 22 patients in which severity of CGVHD could be assessed by the presence of marked erythema or sclerosis on 10 to 90% of their body surface areas (BSA), the BAFF level correlated with the percentage of affected BSA (Spearman r 5 1.63). We then explored the factors that might contribute to elevated BAFF levels. In recipients recovering from either autologous or allogeneic transplant (without GVHD) we observed the highest BAFF levels at day 0 (median of 10534 and 12240 pg/ml respectively), when B cells were severely depleted. As B cell populations recovered to normal levels post transplant, plasma BAFF concentrations declined (Spearman r 5 -.80 and r 5 -.60, respectively), consistent with homeostatic cytokine-consumption dynamics. Despite comparably high levels of BAFF (median of 11342 pg/ ml) at transplant day 0 in 16 patients who later developed CGHVD, BAFF levels in the cross-sectional, natural history patient population were only moderately correlated with the degree of post transplant B cell recovery (r 5 -.48). Since inflammatory triggers can induce elevated BAFF production by dendritic cells, we assessed plasma levels of cytokines indicative of an inflammatory process. In 34 patients, the plasma levels of IP-10 and sTNFRII correlated positively with BAFF levels (r 5 1.627 and r 5 1.642, respectively), consistent with active inflammatory processes in those CGVHD patients with elevated BAFF levels. In a multi-step regression model, the levels of circulating B cells, plasma IP-10 and sTNFRII combined to strongly predict BAFF levels (R 5 .834). These findings suggest that both homeostatic recovery of B cell populations consuming BAFF and inflammatory cytokine cascades initiated by donor-anti-host reactivity combine to regulate BAFF levels post transplant. a major complication of allogeneic stem cell transplantation (al-loSCT). Risk factors include patient age, sex matching, CMV status and degree of match. Regulatory T cells are critical for immune tolerance processes such as aGVHD, and express the transcription factor FOXP3, a member of the forkhead/winged-helix family, identified as a key regulatory gene required for the development and activity of these cells. It has been suggested that genetic expression of FOXP3 is inversely correlated with the severity of the GVHD. We studied donors DNA looking for 5 polymorphisms on the promoter region of the FOXP3 gene, and we tried to correlate them with presence and degree of aGVHD. Patients and Methods: We studied donors of stem cells for allogeneic stem cell transplants. We looked for the presence of the following polymorphisms by PCR: POL01-5906 T/A rs2869211; POL03-3279 A/C rs3761548; POL04-2383 C/T rs3761549; POL05-1383 C/T rs2232364; POL06-924 A/G rs2232365. Results: Our sample consisted of 31 donors, all siblings. In them we found only 2 of the 5 FOXP3 polymorphisms, either as homozygous or heterozygous. These polymorphisms were found in 15/31 donors, with 12 being homozygous (38.7%), and 3 heterozygous (9.7%). These genes polymorphisms were POL03 y POL06. The most observed polymorphism was POL-06 with 9 cases, while POL-03 was found in 6 donors. Only sex difference and CMV status had an elevated hazard ratio for developing GVHD (HR 5 1.18, CI95%: 0.18 to 7.64; p 5 0.85) and (HR 5 3.0, CI95%: 0.07 to 126; p 5 0.46) respectively. We found no statistically significant difference in the incidence of GVHD between patients who had received cells from donor with or without a FOXP3 polymorphism (p 5 0.87). When we analyzed the risk of presenting GVHD, results suggest that having one of the 2 positive polymorphisms of the FOXP3 gene could have a protective effect for the patient. For POL03 HR 5 0.87, CI95%:0.18 to 4.14; p 5 0.86, and for POL06, HR 5 1, CI95%: 0.37 to 2.64, p 5 0.67. Conclusions: Even though our sample is still small to make conclusive remarks, we believe that our results point toward some level of protection from acute GVHD in patients receiving cells from donors expressing POL03 and POL06. It is also worthwhile mentioning the relatively high frequency of these polymorphisms, as well as the absence of the other 3.