Heike Pfeifer

Targeted Therapy With the T-Cell–Engaging Antibody Blinatumomab of Chemotherapy-Refractory Minimal Residual Disease in B-Lineage Acute Lymphoblastic Leukemia Patients Results in High Response Rate and Prolonged Leukemia-Free Survival

PURPOSE Blinatumomab, a bispecific single-chain antibody targeting the CD19 antigen, is a member of a novel class of antibodies that redirect T cells for selective lysis of tumor cells. In acute lymphoblastic leukemia (ALL), persistence or relapse of minimal residual disease (MRD) after chemotherapy indicates resistance to chemotherapy and results in hematologic relapse. A phase II clinical study was conducted to determine the efficacy of blinatumomab in MRD-positive B-lineage ALL. PATIENTS AND METHODS Patients with MRD persistence or relapse after induction and consolidation therapy were included. MRD was assessed by quantitative reverse transcriptase polymerase chain reaction for either rearrangements of immunoglobulin or T-cell receptor genes, or specific genetic aberrations. Blinatumomab was administered as a 4-week continuous intravenous infusion at a dose of 15 μg/m2/24 hours. RESULTS Twenty-one patients were treated, of whom 16 patients became MRD negative. One patient was not evaluable due to a grade 3 adverse event leading to treatment discontinuation. Among the 16 responders, 12 patients had been molecularly refractory to previous chemotherapy. Probability for relapse-free survival is 78% at a median follow-up of 405 days. The most frequent grade 3 and 4 adverse event was lymphopenia, which was completely reversible like most other adverse events. CONCLUSION Blinatumomab is an efficacious and well-tolerated treatment in patients with MRD-positive B-lineage ALL after intensive chemotherapy. T cells engaged by blinatumomab seem capable of eradicating chemotherapy-resistant tumor cells that otherwise cause clinical relapse.

Standardized MRD quantification in European ALL trials : Proceedings of the Second International Symposium on MRD assessment in Kiel, Germany, 18-20 September 2008

Assessment of minimal residual disease (MRD) has acquired a prominent position in European treatment protocols for patients with acute lymphoblastic leukemia (ALL), on the basis of its high prognostic value for predicting outcome and the possibilities for implementation of MRD diagnostics in treatment stratification. Therefore, there is an increasing need for standardization of methodologies and harmonization of terminology. For this purpose, a panel of representatives of all major European study groups on childhood and adult ALL and of international experts on PCR- and flow cytometry-based MRD assessment was built in the context of the Second International Symposium on MRD assessment in Kiel, Germany, 18–20 September 2008. The panel summarized the current state of MRD diagnostics in ALL and developed recommendations on the minimal technical requirements that should be fulfilled before implementation of MRD diagnostics into clinical trials. Finally, a common terminology for a standard description of MRD response and monitoring was established defining the terms ‘complete MRD response’, ‘MRD persistence’ and ‘MRD reappearance’. The proposed MRD terminology may allow a refined and standardized assessment of response to treatment in adult and childhood ALL, and provides a sound basis for the comparison of MRD results between different treatment protocols.

Imatinib compared with chemotherapy as front-line treatment of elderly patients with Philadelphia chromosome-positive acute lymphoblastic leukemia (Ph+ALL).

Elderly patients with Philadelphia chromosome‐positive acute lymphoblastic leukemia (Ph+ALL) have a poor prognosis, with a low complete remission (CR) rate, high induction mortality, and short remission duration. Imatinib (IM) has a favorable toxicity profile but limited antileukemic activity in advanced Ph+ALL. Imatinib in combination with intensive chemotherapy has yielded promising results as front‐line therapy, but its value as monotherapy in newly diagnosed Ph+ALL is not known.

Valproic Acid Stimulates Proliferation and Self-renewal of Hematopoietic Stem Cells

Histone deacetylase inhibitors have attracted considerable attention because of their ability to overcome the differentiation block in leukemic blasts, an effect achieved either alone or in combination with differentiating agents, such as all-trans retinoic acid. We have previously reported favorable effects of the potent histone deacetylase inhibitor valproic acid in combination with all-trans retinoic acid in patients with advanced acute myeloid leukemia leading to blast cell reduction and improvement of hemoglobin. These effects were accompanied by hypergranulocytosis most likely due to an enhancement of nonleukemic myelopoiesis and the suppression of malignant hematopoiesis rather than enforced differentiation of the leukemic cells. These data prompted us to investigate the effect of valproic acid on normal hematopoietic stem cells (HSC). Here we show that valproic acid increases both proliferation and self-renewal of HSC. It accelerates cell cycle progression of HSC accompanied by a down-regulation of p21(cip-1/waf-1). Furthermore, valproic acid inhibits GSK3beta by phosphorylation on Ser9 accompanied by an activation of the Wnt signaling pathway as well as by an up-regulation of HoxB4, a target gene of Wnt signaling. Both are known to directly stimulate the proliferation of HSC and to expand the HSC pool. In summary, we here show that valproic acid, known to induce differentiation or apoptosis in leukemic blasts, stimulates the proliferation of normal HSC, an effect with a potential effect on its future role in the treatment of acute myeloid leukemia.

Therapy with imatinib mesylate (Glivec) preceding allogeneic stem cell transplantation (SCT) in relapsed or refractory Philadelphia-positive acute lymphoblastic leukemia (Ph+ALL).

Imatinib has pronounced antileukemic activity in Ph+ALL, although responses are usually short. To determine whether imatinib may facilitate allogeneic SCT in relapsed or refractory Ph+ALL, we evaluated 46 consecutive, not previously transplanted patients who were enrolled in phase II studies of imatinib. Of 30 patients eligible for SCT, 22 (73%) were actually transplanted. Ten patients were in complete hematologic remission (CHR) (n = 5) or had a complete marrow response (CMR) (n = 5) at the time of SCT, 12 patients had again relapsed or were refractory. After SCT, 18 patients were in complete remission, one patient was refractory, three patients died prior to response assessment. Seven patients (32%) are in ongoing complete remission with a median follow-up of 9.4 (range 1.7–23.8) months. Seven patients (32%) relapsed a median of 5.2 months after SCT. Transplant-related mortality (TRM) was 36%. Probability of disease-free survival (DFS) is 25.5 ± 9.8% overall and 51.4 ± 17.7% when SCT was performed in CHR or CMR, compared with 8.3 ± 8% for SCT during overt leukemia (P = 0.06). In conclusion, imatinib is a well-tolerated salvage therapy prior to allogeneic SCT in patients with Ph+ALL, but requires that SCT be performed within a few weeks of starting treatment to avoid resistance. Disease status at time of transplantation is an important determinant of DFS and TRM.

Management of Philadelphia chromosome–positive acute lymphoblastic leukemia (Ph+ ALL)

The tyrosine kinase inhibitor (TKI) imatinib has become an integral part of front-line therapy for Ph(+) ALL, with remission rates exceeding 90% irrespective of whether imatinib is given alone or combined with chemotherapy. Treatment outcome with imatinib-based regimens has improved compared with historic controls, but most patients who do not undergo allogeneic stem cell transplantation (SCT) eventually relapse. Acquired resistance on TKI treatment is associated with mutations in the bcr-abl tyrosine kinase domain in the majority of patients, and may be detected at low frequency prior to TKI treatment in a subset of patients. Second generation TKIs, eg, dasatinib and nilotinib, show activity against most of the bcr-abl tyrosine kinase domain (TKD) mutations involved in acquired imatinib resistance, but clinical benefit is generally short-lived. Accordingly, SCT in first complete remission (CR) is considered to be the best curative option. Molecular monitoring of minimal residual disease levels appears to have prognostic relevance and should be used to guide treatment. International standardization and quality control efforts are ongoing to ensure comparability of results. Mutation analysis during treatment relies increasingly on highly sensitive PCR techniques or denaturing HPLC and may assist in treatment decisions, eg, in case of molecular relapse. Results from current studies of second-generation TKI as front-line treatment for Ph(+) ALL are promising and show high molecular response rates, but follow-up is still too short to determine their impact on remission duration and long-term survival. Strategies to improve outcome after SCT include the pre-emptive use of imatinib, which appears to reduce the relapse rate. In patients ineligible for transplantation, novel concepts for maintenance therapy are needed. These could involve novel immunotherapeutic interventions and combinations of TKI.

TCRB gene rearrangements in childhood and adult precursor-B-ALL: frequency, applicability as MRD-PCR target, and stability between diagnosis and relapse.

Using the multiplex PCR tubes of the BIOMED-2 Concerted Action, TCRB gene rearrangements were detected in 35% of childhood (n=161) and adult (n=172) precursor-B-ALL patients (Vβ–(Dβ)–Jβ in 25%; Dβ–Jβ in 15%). The presence of TCRB rearrangements showed a significant relation with age (highest frequency of 46% between 5 and 10 years of age) and the presence of TEL-AML1 transcripts, and was associated with relatively high frequencies of IGK-Kde, TCRG, and Vδ2-Jα rearrangements. In 62 out of 65 patients with Southern blot-detected Vβ–(Dβ)–Jβ and/or Dβ–Jβ rearrangements, at least one TCRB gene rearrangement was detected by PCR. Based on combined Southern blot and PCR analysis, oligoclonal TCRB gene rearrangements were observed in only 12% of patients. Analysis of paired diagnosis and relapse samples (n=26) showed that 20 out of 24 (83%) Vβ–(Dβ)–Jβ rearrangements and eight out of 14 (57%) Dβ–Jβ rearrangements remained stable. Using real-time quantitative PCR, a quantitative range ⩽10−4 was obtained in 64% of TCRB gene rearrangements and in 86% of cases a sensitivity ⩽10−4 was obtained. In conclusion, TCRB gene rearrangements occur in 35% of precursor-B-ALL patients and are relatively stable and sensitive PCR targets for detection of minimal residual disease, particularly if this concerns complete Vβ–(Dβ)–Jβ rearrangements.

Dasatinib and low-intensity chemotherapy in elderly patients with Philadelphia chromosome-positive ALL

Prognosis of Philadelphia-positive (Ph(+)) acute lymphoblastic leukemia (ALL) in the elderly has improved during the imatinib era. We investigated dasatinib, another potent tyrosine kinase inhibitor, in combination with low-intensity chemotherapy. Patients older than age 55 years were included in the European Working Group on Adult ALL (EWALL) study number 01 for Ph(+) ALL (EWALL-PH-01 international study) and were treated with dasatinib 140 mg/day (100 mg/day over 70 years) with intrathecal chemotherapy, vincristine, and dexamethasone during induction. Patients in complete remission continued consolidation with dasatinib, sequentially with cytarabine, asparaginase, and methotrexate for 6 months. Maintenance therapy was dasatinib and vincristine/dexamethasone reinductions for 18 months followed by dasatinib until relapse or death. Seventy-one patients with a median age of 69 years were enrolled; 77% had a high comorbidity score. Complete remission rate was 96% and 65% of patients achieved a 3-log reduction in BCR-ABL1 transcript levels during consolidation. Only 7 patients underwent allogeneic hematopoietic stem cell transplantation. At 5 years, overall survival was 36% and up to 45% taking into account deaths unrelated to disease or treatment as competitors. Thirty-six patients relapsed, 24 were tested for mutation by Sanger sequencing, and 75% were T315I-positive. BCR-ABL1(T315I) was tested by allele-specific oligonucleotide reverse transcription-quantitative polymerase chain reaction in 43 patients and detection was associated with short-term relapses. Ten patients (23%) were positive before any therapy and 8 relapsed, all with this mutation. In conclusion, dasatinib combined with low-intensity chemotherapy was well-tolerated and gave long-term survival in 36% of elderly patients with Ph(+) ALL. Monitoring of BCR-ABL1(T315I) from diagnosis identified patients with at high risk of early relapse and may help to personalize therapy.

First-line treatment of Philadelphia chromosome-positive acute lymphoblastic leukaemia in adults.

The tyrosine kinase inhibitor (TKI) imatinib has become an integral part of front-line therapy for Philadelphia chromosome-positive acute lymphoblastic leukaemia, with remission rates exceeding 90% irrespective of whether imatinib is given alone or combined with chemotherapy. Treatment outcome with imatinib-based regimens has improved compared with historic controls, but most patients who do not undergo allogeneic stem cell transplantation (SCT) eventually relapse. Second-generation TKI, e.g. dasatinib and nilotinib, show activity against most of the bcr-abl tyrosine kinase domain mutations involved in acquired imatinib resistance, but clinical benefit is generally short lived. Accordingly, SCT in first complete response is considered to be the best curative option. Strategies to improve outcome in patients ineligible for transplantation as well as after SCT include front-line treatment with more effective TKI to increase molecular response rates. Following SCT, the pre-emptive use of imatinib appears to reduce the relapse rate. Novel immunotherapeutic interventions and combinations of TKI are also being explored.

Increased levels of 2-hydroxyglutarate in AML patients with IDH1-R132H and IDH2-R140Q mutations

To the Editor: Acute myeloid leukemia (AML) is a clonal malignancy of myeloid blood progenitor cells. Karyotyping provides the most powerful prognostic information. Especially for AML cases with normal karyotype, recurring mutations with prognostic significance in genes such as FLT3, NPM1, CEBPA, WT1 and MLL have been identified (1). Recently, isocitrate dehydrogenase (IDH) 1 mutations, which are well characterized in gliomas, were also reported for patients with AML (2). In astrocytomas, oligodendrogliomas and mixed oligoastrocytomas, it was found that IDH1-R132H mutations are the most frequent genetic alterations (3). Notably, IDH1 mutations are emerging as a major diagnostic and prognostic marker in gliomas. Other malignancies only rarely exhibit IDH mutations. However, recently, it has been shown that up to 10% of patients with AML carry mutations in IDH1, with approximately half of the patients harboring R132H and R132C substitutions, respectively (2, 4–7). Additionally, mutations in IDH2 can be found in up to 20% of patients with AML, with the R140Q mutation being detected more often compared to R172K (7). Of interest, the cytogenetic profile of patients with IDH1 mutations is strongly associated with a normal karyotype, intermediate cytogenetic risk and the prevalence of NPM1 mutations (2, 8). The impact of IDH mutations on prognosis