Thiotepa, Busulfan and Cyclophosphamide As a Preparative Regimen for Allogeneic Transplantation for Advanced Chronic Myelogenous Leukemia

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Bone Marrow Transplantation, (1999) 23, 977–981  1999 Stockton Press All rights reserved 0268–3369/99 $12.00 http://www.stockton-press.co.uk/bmt Thiotepa, busulfan and cyclophosphamide as a preparative regimen for allogeneic transplantation for advanced chronic myelogenous leukemia

D Przepiorka1, I Khouri1, P Thall2, R Mehra1, M-S Lee3, C Ippoliti1, S Giralt1, J Gajewski1, K van Besien1, B Andersson1,MKo¨rbling1, AB Deisseroth1 and R Champlin1

Departments of 1Blood and Marrow Transplantation, 2Biomathematics and 3Laboratory Medicine, The University of Texas MD Anderson Cancer Center, Houston, Texas, USA

Summary: blast crisis and 15–40% for transplantation in accelerated or second chronic phase.1–7 Busulfan-based preparative Thirty-six adults with chronic myelogenous leukemia regimens have been favored by some for CML as a result (CML) in second or greater chronic phase, accelerated of the potent anti-myeloid activity and ease of adminis- phase, or blast crisis underwent marrow or blood stem tration of busulfan, and results with the busulfan–cyclopho- cell transplantation from an HLA-matched sibling using sphamide regimen are similar to those achieved with cyclo- high-dose thiotepa, busulfan and cyclophosphamide phosphamide and total body irradiation for patients with (TBC) as the preparative regimen. All evaluable advanced CML.8 N,NЈ,NЈ-triethylenethiophosphoramide patients engrafted and had complete donor chimerism. (thiotepa) is a highly myelosuppressive alkylating agent One patient failed to clear meningeal leukemia, and one that lacks overlapping toxicities with busulfan or cyclopho- patient had one of 30 metaphases positive for the Phila- sphamide.9 We evaluated the combination of thiotepa with delphia chromosome at 2 months post transplant. The busulfan and cyclophosphamide as a preparative regimen remainder of the patients studied had eradication of to determine if addition of a third alkylating agent would CML documented by cytogenetics and/or Southern blot improve outcome without increased toxicity. The maximal for BCR gene rearrangement, and 13 of 15 patients tolerated dose of this combination was previously identified studied became negative for the BCR gene rearrange- in a phase I study.10 Here we report the results using this ment by polymerase chain reaction. Three-year relapse novel regimen for transplantation of patients with advanced rate is 42% (95% CI, 19–64%). The relapse rate was CML from HLA-identical donors. significantly lower for patients transplanted without blast crisis (9% vs 100%, P Ͻ 0.001). Eight (22%, 95% CI, 10–39%) patients had severe or fatal veno-occlusive Materials and methods disease (VOD). Elevated liver enzymes within 1 month prior to transplantation and transplantation using mar- Eligibility criteria row were significantly associated with the occurrence of VOD. Three-year survival is 28% (95% CI, 13–43%). From May 1991 to January 1996, patients with CML past Survival was significantly higher for patients trans- first chronic phase having a t(9;22) documented by cyto- planted without blast crisis (45% vs 0%, P = 0.01). TBC genetics or Southern blot for rearrangement of the BCR is an effective preparative regimen for CML in acceler- gene were eligible for treatment on consecutive phase I– ated phase but not refractory blast crisis, and it should II studies of thiotepa, busulfan and cyclophosphamide as be used with caution in patients with prior hepatopathy preparative regimen. Other requirements included age 15– who have an increased risk of severe VOD. 55 years, Zubrod performance status 0–2, adequate organ Keywords: allogeneic bone marrow transplantation; function, and an HLA-matched related donor.10,11 Protocols chronic myelogenous leukemia; thiotepa; busulfan were approved by the institutional review board of the MD Anderson Cancer Center, and written informed consent was obtained from all patients and donors.

Advanced phase of disease is a strong predictor of poor outcome after HLA-identical marrow transplantation for Treatment 1 chronic myelogenous leukemia (CML). Long-term sur- The preparative regimen consisted of thiotepa 150– vival is reported to be 0–25% for patients transplanted in 250 mg/m2/day i.v. on days Ϫ9, Ϫ8 and Ϫ7, busulfan 1 mg/kg orally every 6 h on days Ϫ6, Ϫ5 and Ϫ4 (for a total of 12 doses), and cyclophosphamide 60 mg/kg/day i.v. on Correspondence: Dr D Przepiorka, Baylor College of Medicine, Center Ϫ Ϫ for Cell and Gene Therapy, 1102 Bates St, Suite 1100, Houston, TX days 3 and 2 (TBC) with precautions and dose adjust- 10,11 77030, USA ments as described previously. Marrow or blood stem Received 15 July 1998; accepted 19 December 1998 cells were collected as reported,10,11 and T cell depletion TBC for advanced CML D Przepiorka et al 978 was not used. Graft-versus-host disease (GVHD) prophy- intervals for proportions were based on the binomial laxis consisted of tacrolimus (FK506) or cyclosporine distribution. (CsA) with methylprednisolone, methotrexate, or immuno- toxin.10,11 Standard post-transplant care was provided. Thirty-four patients received ﬁlgrastim post transplant to Results accelerate neutrophil recovery. Patient characteristics Monitoring post transplant Thirty-six adults were transplanted (Table 1). Twenty had clinical signs of accelerated disease, two were in у2nd Patients were reviewed by one of the study collaborators chronic phase, ﬁve were in lymphoid blast crisis, and nine at least weekly to day 100 and less frequently thereafter. were in myeloid blast crisis. Thirteen of 15 patients treated Regimen-related toxicity was scored using the system of with induction chemotherapy were refractory, and two had Bearman et al12 except that death occurring after day 28 relapsed after allogeneic transplantation using a TBI-based due to regimen-related toxicity was still considered grade preparative regimen. 4. Monitoring for idiopathic interstitial pneumonitis, hemorrhagic cystitis and VOD continued to day 100. GVHD was graded according to the consensus criteria.13 Engraftment and GVHD Marrow biopsies and aspirates were examined at 1, 3, 6, All patients who survived at least 28 days engrafted. The 12, 18 and 24 months after transplantation and annually neutrophil count exceeded 0.5 × 109/l and 1.0 × 109/l at a thereafter. Hematopoietic chimerism was evaluated by median of 10 (range, 8–24) days and 11 (range, 9–27) days restriction fragment length polymorphisms (RFLP). post transplant, respectively. The platelet count exceeded Residual disease was evaluated by conventional cytogen- 20 × 109/l and 50 × 109/l at a median of 19 (9–100+) days etics, Southern blot hybridization for a BCR gene and 24 (10–100+) days post transplant, respectively. Com- rearrangement, and, in some cases, by polymerase chain plete chimerism was found by RFLP analysis in 17 of 20 reaction (PCR) for the bcr-abl transcript.14 patients tested. One patient had mixed chimerism transi-

Definitions Table 1 Patient and donor characteristics The criteria for blast crisis were у30% blasts in the marrow or blood, or any extramedullary collections of blasts, and Number of patients 36 the criteria for accelerated phase included у10% blasts in Median age 44 years (range) (20–55 years) marrow or blood; Ն20% basophils and eosinophils in mar- Patient sex row or blood; cytopenia unrelated to therapy; and leuko- Male/female 21/15 cytosis, thrombocytosis, adenopathy, splenomegaly or bone Disease status pain unresponsive to therapy. Patients with cytogenetic CP у 2, AP/BC 22/14 clonal evolution and no clinical evidence of accelerated Cytogenetics phase were not eligible for these protocols. Neutrophil High risk/Standard risk 16/20 recovery was defined as the first of 3 consecutive days that Prior treatment the absolute neutrophil count exceeded the target value, and Busulfan 6 platelet recovery was defined as the day that the platelet Hydroxyurea 28 count exceeded the target value with no platelet trans- Interferon 13 Interferon + other 11 fusions the following week. After transplantation, obser- Homoharringtonine 6 vation of the Philadelphia chromosome in two or more con- Induction chemotherapy 15 secutive studies without abnormalities in the blood counts Allogeneic transplantation 2 was considered a cytogenetic relapse. A white blood cell Resistant to therapy 12 9 count greater than 10 × 10 /l with cytogenetic or molecular Thiotepa dose level evidence of the Philadelphia chromosome but less than 5% 150 mg/m2 day 3 blasts and no extramedullary disease was considered a hem- 200 mg/m2 day 4 2 atologic relapse in chronic phase. High-risk cytogenetics 250 mg/m day 29 include variant Ph, double Ph, +8, and 3p abnormalities.15 Transplant Marrow/Blood stem cells 22/14 GVHD prophylaxis Statistical analyses Tacrolimus-based 8 Cyclosporine-based 28 At time of analysis, median time from transplantation for Median donor age 44 years all patients was 4.8 years (range, 2.4–7.1 years). Minimum (range) (14–65 years) follow-up for survivors was 3 years. Categorical data were Donor sex compared using the Fisher exact test. The actuarial risks Male/Female 20/16 of GVHD, relapse, survival and treatment-related mortality Patient–donor pair sex-mismatched 11 were estimated according to Kaplan and Meier, and the Patient–donor pair CMV-seronegative 0 logrank test was used to make comparisons. Confidence TBC for advanced CML D Przepiorka et al 979 ently at 3 months post transplant and remains in remission 42% (95% CI, 19–64%), and there were no cytogenetic 40 months post transplant, and two patients had mixed relapses without clinical evidence of recurrence. The risk chimerism within 3 months prior to hematologic relapse. of relapse correlated with phase of disease but not with type Grades 2–4 GVHD occurred in 49% (95%CI, 31–67%), of transplant, age or cytogenetic risk group (Table 3). and grades 3–4 GVHD in 21% (95% CI, 7–35%). Seventy- five percent of the patients survived at least 100 days, and Survival the incidence of chronic GVHD in these patients was 83% (95% CI, 63–100%). Three-year survival is 28% (95% CI, 13–43%). Survival correlated with phase of disease at transplantation Regimen-related toxicity (Figure 1) but not with type of transplant, age or cytogenetic risk group (Table 3). Median survival was 7 months The most common regimen-related complications were for patients transplanted in blast crisis. Nine patients died mucositis and elevated liver enzymes (Table 2). Four in relapse, eight with GVHD, seven with infection, and two patients (11%) had a grade 3 or 4 regimen-related toxicity, with regimen-related toxicity. Two-year treatment-related and two patients (6%) had a cumulative toxicity score у7. mortality is 48% (95% CI, 31–65%) and did not differ sig- Within the first 100 days post transplant, no patient nificantly by phase of disease. developed idiopathic interstitial pneumonitis, and 28% had hemorrhagic cystitis. Twenty-two percent (95% CI, 10– 39%) of the patients had severe VOD (total bilirubin Ն6 Discussion mg/dl), and two patients had fatal VOD. Severe VOD correlated with type of transplant (8/22 marrow recipients vs Although the combination of busulfan and cyclophospham- 0/14 blood stem cell recipients, P = 0.01), transaminases ide has gained favor as a preparative regimen for allogeneic more than twice the upper level of normal within 1 month transplantation, it has limited efficacy for patients with pre-transplant (7/12 with transaminitis vs 1/24 with normal advanced CML.5,6,8,16 In this study, we reduced the dose of liver enzymes, P = 0.006), and use of MTX for GVHD busulfan to minimize toxicity and added a third alkylating prophylaxis (5/10 using MTX vs 3/26 using MP, P = 0.02), agent, thiotepa, with the goal of improving efficacy. All but the risk of severe VOD was not related to phase of patients achieved a medullary remission with TBC, and disease, positive hepatitis serology pre-transplant, and prior only one patient, treated at the lowest thiotepa dose level, busulfan use or prior transplantation. failed to clear Ph-positive metaphases within the first 100 days post transplant. Furthermore, 79% of the patients studied were PCR-negative within 1 year post transplant, which Residual disease and relapse appears superior to the 32% rate reported by Miyamura et All evaluable patients had Ͼ5% blasts in the marrow when al17 in a meta-analysis of all published series of patients assessed 1 month post transplant, but one patient had per- with CML transplanted past first chronic phase. These sistent leptomeningeal leukemia. Within the first 3 months results demonstrate that even with the reduced dose of bus- after transplantation, the BCR gene was germline in mar- ulfan, TBC is effective for early control of disease. row samples from all 22 patients evaluated, and cytogen- For patients with CML in blast crisis, a second chronic etics showed eradication of the Ph chromosome in 28 of phase can be achieved in 16–49% using combination 29 (97%) evaluated. The two patients with mixed chimer- chemotherapy, with a higher response rate reported for ism by RFLP analysis 3 months prior to hematologic patients with lymphoid blast morphology.18,19 Median sur- relapse had a germline BCR gene and normal cytogenetics vival is approximately 6 months for all blast crisis patients at that timepoint. Thirteen (87%) of 15 patients studied ach- and less than 3 months for those refractory to standard-dose ieved eradication of residual disease by PCR for bcr-abl induction therapy. Median survival post transplant for our transcripts; 11 of 14 (79%) were negative within 1 year patients in blast crisis, 7 months, was somewhat longer than post transplant, and one of five (20%) was positive more with standard-dose induction therapy, despite the fact that than 1 year post transplant. The actuarial relapse rate is almost all were refractory to induction chemotherapy, but the relapse rate was high for this group, and there are no long-term survivors. It is clear that TBC is not an active Table 2 Regimen-related toxicity regimen for patients with CML in refractory blast crisis. Grade In this regard, it has no apparent advantage over standard preparative regimens which are associated with a 0–25% 1,2,4,7 01 2 3 4 survival. However, the efficacy of TBC for untreated blast crisis is unknown. Mucosal 2 19 14 1 0 By contrast, only one of our patients with CML in accel- Hepatic 8 18 7 1 2 erated or second chronic phase has relapsed, and the long- Cardiac 30 3 3 0 0 term survival, 45%, is somewhat better than that reported Renal 35 0 1 0 0 historically for this subgroup of patients.1–3,7 The improved Gastrointestinal 13 21 2 0 0 Bladder 34 1 1 0 0 disease control may reflect the addition of thiotepa to the Pulmonary 36 0 0 0 0 preparative regimen; however, potential differences in Nervous system 35 0 1 0 0 patient characteristics preclude a truly meaningful comparison of outcomes in different reports. TBC for advanced CML D Przepiorka et al 980 Table 3 Univariate analysis of clinical outcomes

Outcome Variables 3-year actuarial rates % P (logrank)

Survival Transplant: BMT vs SCT 23 vs 36 0.24 Phase: BC vs CP у 2, AP 0 vs 45 0.01 Age (years): Ͼ 50 vs Ͻ 50 10 vs 35 0.16 Cytogenetics: high vs standard risk 25 vs 30 0.69

Relapse Transplant: BMT vs SCT 55 vs 24 0.10 Phase: BC vs CP у 2, AP 100 vs 9 Ͻ0.001 Age (years): Ͼ50 vs Ͻ 50 40 vs 44 0.40 Cytogenetics: High vs standard risk 49 vs 36 0.29

1 Acknowledgements

The study was supported in part by the American Cyanamid Cor- 0.8 poration, the Tony Anderson Fund, and grants CA-16672 and CA- 49639-05 from the National Institutes of Health.

0.6 References

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