Ahmet H. Elmaagacli

A multicenter prospective phase 2 randomized study of extracorporeal photopheresis for treatment of chronic graft-versus-host disease

Chronic graft-versus-host disease (cGVHD) is a major limitation of successful hematopoietic cell transplantation. The safety and efficacy of extracorporeal photopheresis (ECP) for 12 to 24 weeks together with standard therapy was compared with standard therapy alone in patients with cutaneous manifestations of cGVHD that could not be adequately controlled by corticosteroid treatment. The primary efficacy end point was a blinded quantitative comparison of percent change from baseline in Total Skin Score (TSS) of 10 body regions at week 12. Ninety-five patients were randomized to either ECP and standard therapy (n = 48) or standard therapy alone (n = 47). The median percentage improvement in TSS at week 12 was 14.5% for the ECP arm and 8.5% for the control arm (P = .48). The proportion of patients who had at least a 50% reduction in steroid dose and at least a 25% decrease from baseline in TSS was 8.3% in the ECP arm at week 12 and 0% in the control arm (P = .04). The nonblinded investigator assessment of skin complete or partial responses revealed a significant improvement in favor of ECP (P < .001). ECP was generally well tolerated. These results suggest that ECP may have a steroid-sparing effect in the treatment of cGVHD. Clinical trials registered at www.ClinicalTrials.gov as NCT00054613.

Early human cytomegalovirus replication after transplantation is associated with a decreased relapse risk: evidence for a putative virus-versus-leukemia effect in acute myeloid leukemia patients

The impact of early human cytomegalovirus (HCMV) replication on leukemic recurrence was evaluated in 266 consecutive adult (median age, 47 years; range, 18-73 years) acute myeloid leukemia patients, who underwent allogeneic stem cell transplantation (alloSCT) from 10 of 10 high-resolution human leukocyte Ag-identical unrelated (n = 148) or sibling (n = 118) donors. A total of 63% of patients (n = 167) were at risk for HCMV reactivation by patient and donor pretransplantation HCMV serostatus. In 77 patients, first HCMV replication as detected by pp65-antigenemia assay developed at a median of 46 days (range, 25-108 days) after alloSCT. Taking all relevant competing risk factors into account, the cumulative incidence of hematologic relapse at 10 years after alloSCT was 42% (95% confidence interval [CI], 35%-51%) in patients without opposed to 9% (95% CI, 4%-19%) in patients with early pp65-antigenemia (P < .0001). A substantial and independent reduction of the relapse risk associated with early HCMV replication was confirmed by multivariate analysis using time-dependent covariate functions for grades II to IV acute and chronic graft-versus-host disease, and pp65-antigenemia (hazard ratio = 0.2; 95% CI, 0.1-0.4, P < .0001). This is the first report that demonstrates an independent and substantial reduction of the leukemic relapse risk after early replicative HCMV infection in a homogeneous population of adult acute myeloid leukemia patients.

Mutations in innate immune system NOD2/CARD 15 and TLR-4 (Thr399Ile) genes influence the risk for severe acute graft-versus-host disease in patients who underwent an allogeneic transplantation.

Background. NOD2 and TLR-4 genes belong to the innate immune system that detects invading pathogens through several pattern-recognition receptors. Here we analyzed 403 patients for NOD2 gene mutations and 307 patients for TLR-4 gene mutations (Thr399Ile) with their respective donors and correlated the results with the incidence of acute graft-versus-host disease (aGVHD), severe acute GVHD (saGVHD), the risk for transplant-related mortality (TRM), overall survival (OS) and incidence of infectious complications. Methods. We performed a retrospective single-center study. Genotyping of TLR-4 and NOD2 were evaluated by real-time polymerase chain reaction. Results. Surprisingly, we found a significant reduced incidence of aGVHD, saGVHD, and intestinal GVHD for patients with NOD2 gene mutations on the donor side with 50%, 0% and 2% compared to patients with the wild-type NOD2 gene with 65%, 17%, and 26%, respectively (P<0.02). However, the incidence of saGVHD increased in patients with NOD2 mutations on the patient and donor (P/D) side with 44% versus 17% compared to patients with the wild-type gene (P<0.03). TLR-4 gene mutations at P/D side had an increased risk for saGVHD with 42% versus 15% of patients with wild-type gene (P<0.04). OS, TRM, and incidence of infectious complications were not influenced by the mutated genes. Multivariate analysis confirmed that NOD2 gene mutations on the donor side had a reduced risk for saGVHD (P<0.001), whereas mutations of the NOD2 gene on P/D side had an increased risk for saGVHD (P<0.01) in our analysis. Conclusions. These results suggest that NOD2 mutations have influence on the occurrence of acute GVHD after transplantation.

Improvement of inflammatory bowel disease after allogeneic stem-cell transplantation

Because causes of inflammatory bowel disease (IBD) remain obscure and a curative therapy is still lacking, the influence of stem-cell transplantation (SCT) on IBD is of major interest. We retrospectively analyzed the course of seven patients with Crohn’s disease and four patients with idiopathic ulcerative colitis who underwent allogeneic SCT between July 1994 and August 2002 for acute and chronic myeloid leukemia and myelodysplastic syndrome. After a median follow-up of 34 months posttransplantation, 10 patients are alive. None of the patients showed IBD activity after SCT, except one patient with mild persistent symptoms of Crohn’s disease early after transplant. Colonoscopy after complete discontinuation of prophylactic posttransplant immunosuppression revealed no pathologic findings. These observations imply that host immune dysregulation plays a central role in the perpetuation of IBD. It may be influenced by the implementation of a new allogeneic immune system resulting from the transplantation of hematopoietic stem cells.

The amount of BCR-ABL fusion transcripts detected by the real-time quantitative polymerase chain reaction method in patients with Philadelphia chromosome positive chronic myeloid leukemia correlates with the disease stage

Abstract The use of the real-time reverse-transcription polymerase-chain reaction (RT-PCR) method to quantify BCR-ABL transcripts before and after allogeneic transplant was prospectively studied in 65 patients with chronic myeloid leukemia (CML). The expression of the BCR-ABL transcript was determined and normalized using the glyceraldehyde-3-phosphate dehydrogenase (GAPDH) housekeeping gene product as an endogenous reference. In the single step real-time PCR assay, tenfold serial dilutions of cDNA of the K5652 cell line remained positive down to 100 pg cDNA only. However, molecular relapses of CML after transplant were only safely detectable when a nested real-time PCR assay was performed, which was able to detect 1–10 pg cDNA from a tenfold serial dilution. The median normalized BCR-ABL transcript level was measured as 0.004% in 17 patients with a molecular relapse, 0.4% in 7 patients with a cytogenetic relapse, 2.6% in 36 patients with a stable phase of CML, and 36% in 5 patients with a relapse in a blast crisis. The analyzed median normalized amount of BCR-ABL transcript differed significantly (P<0.001) between the various disease stages. In ten CML patients with relapse, the real-time PCR method was used to monitor the response of various immunotherapies as donor leukocyte infusions, withdrawal of immunosuppression, or interferon-α application. The results of the quantitative evaluation of BCR-ABL transcripts reflected very well the clinical effect of the different applied immunotherapies. The new real-time PCR method seems to be a suitable technique for the early detection of relapse after allogeneic transplant in patients with the BCR-ABL transcript. Its ability to distinguish between molecular and cytogenetic relapse (P<0.001) allows early therapeutic decisions.

Progressive Improvement in Cutaneous and Extracutaneous Chronic Graft-versus-Host Disease after a 24-Week Course of Extracorporeal Photopheresis—Results of a Crossover Randomized Study

In a prior multicenter randomized controlled trial, we found that a 12-week course of extracorporeal photopheresis (ECP) plus standard immunosuppressive therapy resulted in several beneficial outcomes in patients with corticosteroid-refractory/intolerant/dependent chronic graft-versus-host disease (GVHD). Here, we report the results of an open-label crossover ECP study in 29 eligible participants randomized initially to the standard of care non-ECP (control) arm. Eligible for the crossover ECP study were control arm patients who either (1) had progression of cutaneous chronic GVHD (cGVHD), defined as >25% worsening from baseline as measured by the percent change in the total skin score (TSS) at any time, or (2) had less than 15% improvement in the TSS, or had a ≤25% reduction in corticosteroid dose at week 12 of the initial study. ECP was administered 3 times during week 1, then twice weekly until week 12, followed by 2 treatments monthly until week 24. The median age of the study cohort was 43 (20-67) years and 90% had extensive cGVHD. The median months from onset of cGVHD to start of ECP were 26 (range: 4-79). Twenty-five of 29 patients (86%) completed the 24-week course of ECP. Complete or partial skin response at week 24 was noted in 9 patients (31%). The median percent of decrease in TSS from baseline to weeks 12 and 24 was -7.9 and -25.8, respectively. In 4 (17%) and 8 (33%) patients, a ≥50% reduction in corticosteroid dose at weeks 12 and 24 was observed. Extracutaneous cGVHD response was highest in oral mucosa with 70% complete and partial resolution after week 24. In conclusion, progressive improvement in cutaneous and extracutaneous cGVHD was observed after a 24-week course of ECP in patients who previously had no clinical improvement or exhibited worsening of cGVHD while receiving standard immunosuppressive therapy alone in a randomized study. These results confirm previous findings and support the notion that prolonged ECP appears to be necessary for optimal therapeutic effects in corticosteroid-refractory cGVHD patients.

Therapeutic application of small interfering RNA directed against bcr-abl transcripts to a patient with imatinib-resistant chronic myeloid leukaemia

RNA interference is referred to as the recently discovered process of sequence-specific, post-transcriptional gene silencing that is initiated by double-stranded RNA molecules known as small interfering RNAs (siRNA). We herein present a first report on the in vivo application of targeted non-virally delivered synthetic bcr-abl siRNA in a female patient with recurrent Philadelphia chromosome-positive chronic myeloid leukaemia (CML) resistant to imatinib (Y253F mutation) and chemotherapy after allogeneic haematopoietic stem cell transplantation. We found a remarkable inhibition of the overexpressed bcr-abl oncogene resulting in increased apoptosis of CML cells. In vivo siRNA application was well tolerated without any clinically adverse events. Our findings imply that the clinical application of synthetic siRNA is feasible, safe and has real potential for genetic-based therapies using synthetic non-viral carriers.

Reduced risk of persisting cytomegalovirus pp65 antigenemia and cytomegalovirus interstitial pneumonia following allogeneic PBSCT.

In order to evaluate the risk of cytomegalovirus (CMV) associated disease after allogeneic stem cell transplantation (SCT), 158 consecutive patients at risk for infection were analyzed. BMT was performed in 101 patients and peripheral blood stem cell transplantation (PBSCT) in 57 patients. CMV antigenemia was found in 57 cases (56%) after BMT and 27 cases (47%) after PBSCT, respectively. CMV antigenemia resistant to a 14-day course of GCV was found in 26 patients (26%) after BMT but in only four patients (7%) after PBSCT (P < 0.01). Eighteen patients (11%) developed cmv disease, 14 post BMT and four post PBSCT. Lethal CMV-related interstitial pneumonia (CMV-IP) occurred in 13 cases of whom 12 patients were bone marrow recipients (P = 0.04). The subgroup of seronegative patients with a CMV seropositive donor had a significantly lower risk of developing CMV antigenemia, GCV-resistant CMV antigenemia (P < 0.01) and CMV-related disease (P = 0.01). In conclusion, the incidence of persistent CMV antigenemia and CMV-IP was significantly reduced when allogeneic transplantation was performed with peripheral blood stem cells instead of bone marrow. These findings suggest that our previous in vitro data on improved immune reconstitution after allogeneic PBSCT as compared to allogeneic BMT have clinical relevance. Bone Marrow Transplantation (2000) 25, 665–672.

Relation of an interleukin-23 receptor gene polymorphism to graft-versus-host disease after hematopoietic-cell transplantation

Polymorphisms in cytokine genes can influence immune responses and inflammation and thereby affecting the outcome of hematopoietic stem-cell transplantation. We analyzed a single-nucleotide polymorphism in the gene for the interleukin-23 receptor (IL-23R) (1142G>A) in a cohort of 221 transplant recipients and their human leukocyte antigen (HLA)-identical sibling donors and in a second cohort of 186 transplant recipients and their HLA-identical unrelated donors. Genotypes were tested for an association with graft-versus-host disease (GVHD) by multivariate analysis. The donors IL-23R genotype was significantly associated with a reduced risk of acute GVHD in both cohorts for patients after transplant. Analysis of all 407 transplant recipients showed that IL-23R (1142G>A, Arg381Gln) genotype of the donor was associated with a decreased risk of grades 2–4 acute GVHD (31.6 compared to 51.0%, P=0.02) and grades 3–4 severe acute GVHD (3.9 compared to 23.4%, P=0.003). Death in remission was significantly lower in patients transplanted from donors with variant IL23-R (11.7 versus 27.7%, P=0.028), whereas overall survival or relapse rates were not influenced significantly by the IL-23R genotype. Among recipients of hematopoietic cells from HLA-identical donors, the IL-23R (Arg381Gln) gene variant on the donor side has a protective effect on the occurrence of acute GVHD in recipients after transplantation.

Outcome of allogeneic stem cell transplantation in patients with myelofibrosis

Summary:Myelofibrosis, either de novo or following pre-existing hematologic diseases, can be cured by allogeneic hematopoietic stem cell transplantation (SCT), but SCT is associated with significant morbidity and mortality, making the choice and timing of transplantation difficult. In all, 20 patients (seven female and 13 male), with a median age of 45 years (range 22–57 years), with idiopathic myelofibrosis (n=12), post-thrombocythemic (n=3) or post-polycythemic (n=2) myeloid metaplasia or leukemic transformation (n=3), underwent allogeneic SCT at our center between 1994 and 2003. With regard to the pre-transplant presence of risk factors such as hemoglobin levels ⩽10 mg/dl, grade III marrow fibrosis or peripheral blast counts >1%, patients were divided into high- and low-risk groups. The estimated 3-year survival post transplant was 38.5% for all patients. The 3-year probability of survival within the high-risk group (n=11) characterized by the presence of at least two risk factors was 16%. Low-risk patients (n=9) with at most one risk factor had an estimated 3-year survival of 67%. Thus, previously defined risk determinants for the outcome of allogeneic transplantation for myelofibrosis may provide useful information facilitating treatment strategies. Our data suggest that transplantation should be taken into consideration before poor prognostic variables develop.