DataSheet_2_Association between gene polymorphisms in the cyclophosphamide metabolism pathway with complications after haploidentical hematopoietic stem cell transplantation.pdf

Allogeneic hematopoietic stem cell transplantation (allo-HSCT) is a curative treatment for patients with hematologic malignances. Haploidentical HSCT (Haplo-HSCT) is an alternative option for patients who do not have an HLA-matched donor. The use of post-transplantation high dose cyclophosphamide (PT-Cy) is commonly employed for graft-versus-host disease (GVHD) prophylaxis in haplo-HSCT. Cyclophosphamide (Cy) is an alkylating agent with antineoplastic and immunosuppressive activity, whose bioactivation requires the activity of polymorphic enzymes in the liver to produce phosphoramide mustard, which is a DNA alkylating agent. To identify polymorphisms in the genes of Cy metabolism and correlate them with post-HSCT complications [GVHD, sinusoidal obstruction syndrome (SOS), hemorrhagic cystitis (HC) and transplant-related mortality (TRM)], we designed a custom next-generation sequencing panel with Cy metabolism enzymes. We analyzed 182 patients treated with haplo-HSCT with PT-Cy from 2007 to 2019, detecting 40 variants in 11 Cy metabolism genes. Polymorphisms in CYP2B6, a major enzyme involved in Cy activation, were associated with decreased activity of this enzyme and a higher risk of Graf-versus-host disease (GVHD). Variants in other activation enzymes (CYP2A6, CYP2C8, CYP2C9, CYP2C19) lead to decreased enzyme activity and were associated with GVHD. Polymorphisms in detoxification genes such as glutathione S-transferases decreased the ability to detoxify cyclophosphamide metabolites due to lower enzyme activity, which leads to increased amounts of toxic metabolites and the development of III-IV acute GVHD. GSMT1*0 a single nucleotide polymorphism previously recognized as a risk factor for SOS was associated with a higher risk of SOS. We conclude that polymorphisms of genes involved in the metabolism of cyclophosphamide in our series are associated with severe grades of GVHD and toxicities (SOS and TRM) after haplo-HSCT and could be used to improve the clinical management of transplanted patients.</p

Table_1_Association between gene polymorphisms in the cyclophosphamide metabolism pathway with complications after haploidentical hematopoietic stem cell transplantation.xlsx

Allogeneic hematopoietic stem cell transplantation (allo-HSCT) is a curative treatment for patients with hematologic malignances. Haploidentical HSCT (Haplo-HSCT) is an alternative option for patients who do not have an HLA-matched donor. The use of post-transplantation high dose cyclophosphamide (PT-Cy) is commonly employed for graft-versus-host disease (GVHD) prophylaxis in haplo-HSCT. Cyclophosphamide (Cy) is an alkylating agent with antineoplastic and immunosuppressive activity, whose bioactivation requires the activity of polymorphic enzymes in the liver to produce phosphoramide mustard, which is a DNA alkylating agent. To identify polymorphisms in the genes of Cy metabolism and correlate them with post-HSCT complications [GVHD, sinusoidal obstruction syndrome (SOS), hemorrhagic cystitis (HC) and transplant-related mortality (TRM)], we designed a custom next-generation sequencing panel with Cy metabolism enzymes. We analyzed 182 patients treated with haplo-HSCT with PT-Cy from 2007 to 2019, detecting 40 variants in 11 Cy metabolism genes. Polymorphisms in CYP2B6, a major enzyme involved in Cy activation, were associated with decreased activity of this enzyme and a higher risk of Graf-versus-host disease (GVHD). Variants in other activation enzymes (CYP2A6, CYP2C8, CYP2C9, CYP2C19) lead to decreased enzyme activity and were associated with GVHD. Polymorphisms in detoxification genes such as glutathione S-transferases decreased the ability to detoxify cyclophosphamide metabolites due to lower enzyme activity, which leads to increased amounts of toxic metabolites and the development of III-IV acute GVHD. GSMT1*0 a single nucleotide polymorphism previously recognized as a risk factor for SOS was associated with a higher risk of SOS. We conclude that polymorphisms of genes involved in the metabolism of cyclophosphamide in our series are associated with severe grades of GVHD and toxicities (SOS and TRM) after haplo-HSCT and could be used to improve the clinical management of transplanted patients.</p

DataSheet_4_Association between gene polymorphisms in the cyclophosphamide metabolism pathway with complications after haploidentical hematopoietic stem cell transplantation.pdf

Allogeneic hematopoietic stem cell transplantation (allo-HSCT) is a curative treatment for patients with hematologic malignances. Haploidentical HSCT (Haplo-HSCT) is an alternative option for patients who do not have an HLA-matched donor. The use of post-transplantation high dose cyclophosphamide (PT-Cy) is commonly employed for graft-versus-host disease (GVHD) prophylaxis in haplo-HSCT. Cyclophosphamide (Cy) is an alkylating agent with antineoplastic and immunosuppressive activity, whose bioactivation requires the activity of polymorphic enzymes in the liver to produce phosphoramide mustard, which is a DNA alkylating agent. To identify polymorphisms in the genes of Cy metabolism and correlate them with post-HSCT complications [GVHD, sinusoidal obstruction syndrome (SOS), hemorrhagic cystitis (HC) and transplant-related mortality (TRM)], we designed a custom next-generation sequencing panel with Cy metabolism enzymes. We analyzed 182 patients treated with haplo-HSCT with PT-Cy from 2007 to 2019, detecting 40 variants in 11 Cy metabolism genes. Polymorphisms in CYP2B6, a major enzyme involved in Cy activation, were associated with decreased activity of this enzyme and a higher risk of Graf-versus-host disease (GVHD). Variants in other activation enzymes (CYP2A6, CYP2C8, CYP2C9, CYP2C19) lead to decreased enzyme activity and were associated with GVHD. Polymorphisms in detoxification genes such as glutathione S-transferases decreased the ability to detoxify cyclophosphamide metabolites due to lower enzyme activity, which leads to increased amounts of toxic metabolites and the development of III-IV acute GVHD. GSMT1*0 a single nucleotide polymorphism previously recognized as a risk factor for SOS was associated with a higher risk of SOS. We conclude that polymorphisms of genes involved in the metabolism of cyclophosphamide in our series are associated with severe grades of GVHD and toxicities (SOS and TRM) after haplo-HSCT and could be used to improve the clinical management of transplanted patients.</p

DataSheet_1_Association between gene polymorphisms in the cyclophosphamide metabolism pathway with complications after haploidentical hematopoietic stem cell transplantation.pdf

Allogeneic hematopoietic stem cell transplantation (allo-HSCT) is a curative treatment for patients with hematologic malignances. Haploidentical HSCT (Haplo-HSCT) is an alternative option for patients who do not have an HLA-matched donor. The use of post-transplantation high dose cyclophosphamide (PT-Cy) is commonly employed for graft-versus-host disease (GVHD) prophylaxis in haplo-HSCT. Cyclophosphamide (Cy) is an alkylating agent with antineoplastic and immunosuppressive activity, whose bioactivation requires the activity of polymorphic enzymes in the liver to produce phosphoramide mustard, which is a DNA alkylating agent. To identify polymorphisms in the genes of Cy metabolism and correlate them with post-HSCT complications [GVHD, sinusoidal obstruction syndrome (SOS), hemorrhagic cystitis (HC) and transplant-related mortality (TRM)], we designed a custom next-generation sequencing panel with Cy metabolism enzymes. We analyzed 182 patients treated with haplo-HSCT with PT-Cy from 2007 to 2019, detecting 40 variants in 11 Cy metabolism genes. Polymorphisms in CYP2B6, a major enzyme involved in Cy activation, were associated with decreased activity of this enzyme and a higher risk of Graf-versus-host disease (GVHD). Variants in other activation enzymes (CYP2A6, CYP2C8, CYP2C9, CYP2C19) lead to decreased enzyme activity and were associated with GVHD. Polymorphisms in detoxification genes such as glutathione S-transferases decreased the ability to detoxify cyclophosphamide metabolites due to lower enzyme activity, which leads to increased amounts of toxic metabolites and the development of III-IV acute GVHD. GSMT1*0 a single nucleotide polymorphism previously recognized as a risk factor for SOS was associated with a higher risk of SOS. We conclude that polymorphisms of genes involved in the metabolism of cyclophosphamide in our series are associated with severe grades of GVHD and toxicities (SOS and TRM) after haplo-HSCT and could be used to improve the clinical management of transplanted patients.</p

DataSheet_3_Association between gene polymorphisms in the cyclophosphamide metabolism pathway with complications after haploidentical hematopoietic stem cell transplantation.pdf

Allogeneic hematopoietic stem cell transplantation (allo-HSCT) is a curative treatment for patients with hematologic malignances. Haploidentical HSCT (Haplo-HSCT) is an alternative option for patients who do not have an HLA-matched donor. The use of post-transplantation high dose cyclophosphamide (PT-Cy) is commonly employed for graft-versus-host disease (GVHD) prophylaxis in haplo-HSCT. Cyclophosphamide (Cy) is an alkylating agent with antineoplastic and immunosuppressive activity, whose bioactivation requires the activity of polymorphic enzymes in the liver to produce phosphoramide mustard, which is a DNA alkylating agent. To identify polymorphisms in the genes of Cy metabolism and correlate them with post-HSCT complications [GVHD, sinusoidal obstruction syndrome (SOS), hemorrhagic cystitis (HC) and transplant-related mortality (TRM)], we designed a custom next-generation sequencing panel with Cy metabolism enzymes. We analyzed 182 patients treated with haplo-HSCT with PT-Cy from 2007 to 2019, detecting 40 variants in 11 Cy metabolism genes. Polymorphisms in CYP2B6, a major enzyme involved in Cy activation, were associated with decreased activity of this enzyme and a higher risk of Graf-versus-host disease (GVHD). Variants in other activation enzymes (CYP2A6, CYP2C8, CYP2C9, CYP2C19) lead to decreased enzyme activity and were associated with GVHD. Polymorphisms in detoxification genes such as glutathione S-transferases decreased the ability to detoxify cyclophosphamide metabolites due to lower enzyme activity, which leads to increased amounts of toxic metabolites and the development of III-IV acute GVHD. GSMT1*0 a single nucleotide polymorphism previously recognized as a risk factor for SOS was associated with a higher risk of SOS. We conclude that polymorphisms of genes involved in the metabolism of cyclophosphamide in our series are associated with severe grades of GVHD and toxicities (SOS and TRM) after haplo-HSCT and could be used to improve the clinical management of transplanted patients.</p

Cumulative incidence analysis of three different outcomes among high and low <i>PTCH1</i> expression groups established using previously published thresholds.

<p>HR, hazard ratio; exp, expression; IFFS, imatinib failure-free survival; MMR, major molecular response; PFS, progression-free survival; CML-OS, CML-related overall survival; N, number. The solid line indicates high <i>PTCH1</i> expression and the dashed line low <i>PTCH1</i> expression. Non-CML-related death was considered a competing risk in every variable analyzed by the cumulative incidence procedure. In addition, discontinuation of imatinib due to side effects was considered a competing risk for the analysis of IFFS. The HR is not shown if no event was recorded in any of the groups due to non-precise estimations. (A) IFFS analyzed by the cumulative incidence procedure and the Fine-Gray test. (B) PFS analyzed by the cumulative incidence procedure and the Fine-Gray test. (C) CML-OS analyzed by the cumulative incidence procedure and the Fine-Gray test.</p

Cumulative incidence or Kaplan-Meier analysis of four different outcomes in the high and low <i>PTCH1</i> expression groups.

<p>HR, hazard ratio; exp, expression; IFFS, imatinib failure-free survival; MMR, major molecular response; CCyR, complete cytogenetic response; PFS, progression-free survival; CML-OS, CML-related overall survival; N, number. The solid line indicates high <i>PTCH1</i> expression (>0.0259) and the dashed line low <i>PTCH1</i> expression. Non-CML-related death was considered a competing risk in every variable analyzed by the cumulative incidence procedure. In addition, discontinuation of imatinib due to side effects was considered a competing risk for the analysis of IFFS. The HR is not shown if no event was recorded in any of the groups due to non-precise estimations. (A) IFFS analyzed by the cumulative incidence procedure and the Fine-Gray test. (B) MMR analyzed by Kaplan-Meier plot, log-rank test, and Cox regression (for HR estimation). (C) CCyR analyzed by Kaplan-Meier plot and log-rank test. (D) PFS analyzed by the cumulative incidence procedure and the Fine-Gray test. (E) CML-OS analyzed by the cumulative incidence procedure and the Fine-Gray test.</p

Patient characteristics.

<p>Patient characteristics.</p

<i>PTCH1</i> is a reliable marker for predicting imatinib response in chronic myeloid leukemia patients in chronic phase

<div><p>Patched homolog 1 gene (<i>PTCH1</i>) expression and the ratio of <i>PTCH1</i> to <i>Smoothened</i> (<i>SMO</i>) expression have been proposed as prognostic markers of the response of chronic myeloid leukemia (CML) patients to imatinib. We compared these measurements in a realistic cohort of 101 patients with CML in chronic phase (CP) using a simplified qPCR method, and confirmed the prognostic power of each in a competing risk analysis. Gene expression levels were measured in peripheral blood samples at diagnosis. The <i>PTCH1</i>/<i>SMO</i> ratio did not improve <i>PTCH1</i> prognostic power (area under the receiver operating characteristic curve 0.71 vs. 0.72). In order to reduce the number of genes to be analyzed, <i>PTCH1</i> was the selected measurement. High and low <i>PTCH1</i> expression groups had significantly different cumulative incidences of imatinib failure (IF), which was defined as discontinuation of imatinib due to lack of efficacy (5% vs. 25% at 4 years, <i>P</i> = 0.013), probabilities of achieving a major molecular response (81% vs. 53% at first year, <i>P</i> = 0.02), and proportions of early molecular failure (14% vs. 43%, <i>P</i> = 0.015). Every progression to an advanced phase (n = 3) and CML-related death (n = 2) occurred in the low <i>PTCH1</i> group (<i>P</i><0.001 for both comparisons). <i>PTCH1</i> was an independent prognostic factor for the prediction of IF. We also validated previously published thresholds for <i>PTCH1</i> expression. Therefore, we confirmed that <i>PTCH1</i> expression can predict the imatinib response in CML patients in CP by applying a more rigorous statistical analysis. Thus, <i>PTCH1</i> expression is a promising molecular marker for predicting the imatinib response in CML patients in CP.</p></div

Flow diagram of sample selection and study design.

<p>RELMC, Spanish CML registry; IF, imatinib failure; IFFS, imatinib failure-free survival; CML-OS, overall survival related to CML. *Clinical data for 40 patients from two hospitals were collected <i>ad hoc</i>. †Groups established with previous thresholds were only compared in a univariate analysis.</p