Mats Merup

Up-Front Autologous Stem-Cell Transplantation in Peripheral T-Cell Lymphoma: NLG-T-01

PURPOSE Systemic peripheral T-cell lymphomas (PTCLs) respond poorly to conventional therapy. To evaluate the efficacy of a dose-dense approach consolidated by up-front high-dose chemotherapy (HDT) and autologous stem-cell transplantation (ASCT) in PTCL, the Nordic Lymphoma Group (NLG) conducted a large prospective phase II study in untreated systemic PTCL. This is the final report, with a 5-year median follow-up, of the NLG-T-01 study. PATIENTS AND METHODS Treatment-naive patients with PTCL age 18 to 67 years (median, 57 years) were included. Anaplastic lymphoma kinase (ALK) -positive anaplastic large-cell lymphoma (ALCL) was excluded. An induction regimen of six cycles of biweekly CHOEP (cyclophosphamide, doxorubicin, vincristine, etoposide, and prednisone) was administered (in patients age > 60 years, etoposide was omitted). If in complete or partial remission, patients proceeded to consolidation with HDT/ASCT. RESULTS Of 166 enrolled patients, 160 had histopathologically confirmed PTCL. The majority presented with advanced-stage disease, B symptoms, and elevated serum lactate dehydrogenase. A total of 115 underwent HDT/ASCT, with 90 in complete remission at 3 months post-transplantation. Early failures occurred in 26%. Treatment-related mortality was 4%. At 60.5 months of median follow-up, 83 patients were alive. Consolidated 5-year overall and progression-free survival (PFS) were 51% (95% CI, 43% to 59%) and 44% (95% CI, 36% to 52%), respectively. Best results were obtained in ALK-negative ALCL. CONCLUSION Dose-dense induction followed by HDT/ASCT was well tolerated and led to long-term PFS in 44% of treatment-naive patients with PTCL. This represents an encouraging outcome, particularly considering the high median age and adverse risk profile of the study population. Therefore, dose-dense induction and HDT/ASCT are a rational up-front strategy in transplantation-eligible patients with PTCL.

Lenalidomide inhibits the malignant clone and up-regulates the SPARC gene mapping to the commonly deleted region in 5q− syndrome patients

Myelodysplastic syndromes (MDSs) are a group of hematopoietic stem cell disorders characterized by ineffective hematopoiesis and peripheral blood cytopenias. Lenalidomide has dramatic therapeutic effects in patients with low-risk MDS and a chromosome 5q31 deletion, resulting in complete cytogenetic remission in >60% of patients. The molecular basis of this remarkable drug response is unknown. To gain insight into the molecular targets of lenalidomide we investigated its in vitro effects on growth, maturation, and global gene expression in isolated erythroblast cultures from MDS patients with del(5)(q31). Lenalidomide inhibited growth of differentiating del(5q) erythroblasts but did not affect cytogenetically normal cells. Moreover, lenalidomide significantly influenced the pattern of gene expression in del(5q) intermediate erythroblasts, with the VSIG4, PPIC, TPBG, activin A, and SPARC genes up-regulated by >2-fold in all samples and many genes involved in erythropoiesis, including HBA2, GYPA, and KLF1, down-regulated in most samples. Activin A, one of the most significant differentially expressed genes between lenalidomide-treated cells from MDS patients and healthy controls, has pleiotropic functions, including apoptosis of hematopoietic cells. Up-regulation and increased protein expression of the tumor suppressor gene SPARC is of particular interest because it is antiproliferative, antiadhesive, and antiangiogenic and is located at 5q31-q32, within the commonly deleted region in MDS 5q− syndrome. We conclude that lenalidomide inhibits growth of del(5q) erythroid progenitors and that the up-regulation of SPARC and activin A may underlie the potent effects of lenalidomide in MDS with del(5)(q31). SPARC may play a role in the pathogenesis of the 5q− syndrome.

Cloning of two candidate tumor suppressor genes within a 10 kb region on chromosome 13q14, frequently deleted in chronic lymphocytic leukemia

Previous studies have indicated the presence of a putative tumor suppressor gene on chromosome 13q14, commonly deleted in patients with B-cell chronic lymphocytic leukemia (B-CLL). We have previously defined a minimally deleted region of 130 kb centromeric to the marker D13S272, and constructed a PAC and cosmid contig encompassing this area. In the present study we have made a detailed restriction and transcriptional map of the region of interest. Using these tools we have screened a panel of 206 primary CLL clones and three cell lines. In five CLL cases we found limited deletions defining the region of interest to an area of no more than 10 kb. Two adjacent genes, termed Leu1 and Leu2 (leukemia-associated gene 1 and 2), were mapped to the minimally deleted region, with several patients showing deletion borders within these genes. The Leu1 and Leu2 genes show little homology to previously published genes at the nucleotide and expected translated amino acid sequence level. Mutational analysis of the Leu1 and 2 genes in 170 CLL samples revealed no small intragenic mutations or point mutations. However, in all cases of 13q14 loss examined, the first exon of both genes, which are only 300 bp apart, were deleted. We conclude that the Leu1 and Leu2 genes are strong candidates as tumor suppressor gene(s) involved in B-CLL leukemogenesis.

A phase II trial of pegylated interferon alpha-2b therapy for polycythemia vera and essential thrombocythemia: feasibility, clinical and biologic effects, and impact on quality of life.

Conventional interferon‐α (IFN) is an effective treatment for patients with myeloproliferative disorders. However, many patients discontinue therapy because of side effects.

Different outcome of allogeneic transplantation in myelofibrosis using conventional or reduced-intensity conditioning regimens

Allogeneic haematopoietic stem cell transplantation remains the only curative treatment of myelofibrosis with myeloid metaplasia (MMM). Previous reports have indicated significant treatment‐related mortality (TRM) for patients transplanted after myeloablative conditioning but superior survival has been reported after reduced‐intensity conditioning (RIC). We report the results of a survey of all allogeneic transplantations for MMM performed in Sweden at six transplant units between 1982 and 2004. Twenty‐seven patients were transplanted; 17 with a myeloablative conditioning regimen and 10 with RIC. The median age was 50 years (5–63 years) at transplantation. After a median follow up of 55 months, 20 patients are alive. TRM was 10% in the RIC group and 30% in the myeloablative group. There was no difference in survival for high or low‐risk patients according to Cervantes score or between sibling and unrelated donor transplantations.

Effects of PRIMA-1 on chronic lymphocytic leukaemia cells with and without hemizygous p53 deletion

The tumour suppressor gene p53 is the most commonly mutated gene in solid tumours. Although less common in haematological malignancies, 10–15% of B‐cell chronic lymphocytic leukaemia (B‐CLL) cases carry a p53 mutation. Recently, the compound P53‐dependent reactivation and induction of massive apoptosis (PRIMA‐1) has been shown to induce cytotoxic effects and apoptosis in human tumour cells by restoration of the transcriptional activity of mutated p53. This is believed to be mediated by a change in the conformation of mutated p53 protein, restoring DNA binding and activation of p53 target genes. We studied the effects of PRIMA‐1 and commonly used anti‐leukaemic drugs on B‐CLL cells from 14 patients with and without hemizygous p53 deletion. Cells obtained from peripheral blood or bone marrow were exposed to PRIMA‐1 and fludarabine alone or in combination. PRIMA‐1 showed cytotoxic effects on B‐CLL cells from samples with and without hemizygous p53 deletion. Furthermore, conventional B‐CLL drugs were less effective in cell samples with hemizygous p53 deletion and the response depended on the size of the p53 deleted clone. Finally, we found evidence for synergistic and additive effects of PRIMA‐1 in combination with fludarabine.

Amplification of multiple regions of chromosome 12, including 12q13–15, in chronic lymphocytic leukaemia

Abstract: Trisomy 12 is a frequent abnormality in chronic lymphocytic leukaemia (CLL). The biological importance of trisomy 12 is still poorly understood but it has been suggested that one or several genes are duplicated leading to malignant transformation. We present a case with amplification of 12q13–22 found in a clinically aggressive relapse of CLL. A smaller region, 12q13–15, was amplified most frequently and a YAC containing the MDM2 gene gave the highest number of signals. Additionally, in a subclone an amplicon containing at least 5 copies of a cosmid from 12q23–24 was detected. The case shows that small duplications of chromosome 12, not revealed by cytogenetic analysis, may occur in CLL. Also, it shows that cytogenetic clonal evolution can occur in CLL without morphological evidence of blast transformation. Our results indicate that the 12q13–15 region carries an important gene for CLL progression.

Mutated and non-mutated TP53 as targets in the treatment of leukaemia

TP53 is mutated in 10–20% of cases of chronic lymphocytic leukaemia (CLL) and 3–8% of cases of acute myeloid leukaemia (AML). Recently, two classes of compounds that restore the function of p53 in tumours have been described. PRIMA‐1 (p53‐dependent reactivation and induction of massive apoptosis) restores the wild‐type conformation of mutant TP53, whereas RITA (reactivation of p53 and induction of tumour cell apoptosis) increases intracellular levels of p53. We evaluated the effects of RITA alone and in combination with PRIMA‐1 or conventional cytostatics on leukaemic cells isolated from AML and CLL patients. AML samples with −17, which are more resistant to daunorubicin and cytarabine compared with samples without −17, were effectively killed by PRIMA‐1. RITA, which stabilizes the function of wild‐type p53, induced apoptosis in AML cells. In contrast to that seen with PRIMA‐1, AML patient samples without −17 were significantly more sensitive to RITA. Similarly, RITA exerted dose‐dependent apoptosis and cytotoxicity in CLL cells, which was significantly more pronounced in samples without hemizygous TP53 deletion. Notably, a synergistic effect was observed in all CLL samples with RITA and fludarabine in combination. In both AML and CLL cells exposure to RITA resulted in induction of intracellular p53. We conclude that small molecules targeting p53 might be of clinical importance in the future for treating drug‐resistant leukaemia.

PRIMA-1 induces apoptosis in acute myeloid leukaemia cells with p53 gene deletion

The p53 tumour suppressor gene located on chromosome 17p13 is the most frequently mutated gene in human tumours. About 5–8% of cases with acute myeloid leukaemia (AML) carry the p53 mutation. Recently, the compound p53‐dependent reactivation and induction of massive apoptosis (PRIMA‐1) has been shown to induce cytotoxic effects and apoptosis in human tumour cells by restoration of the transcriptional activity of mutated p53. This is believed to be mediated by a change in the conformation of mutated p53 protein, restoring DNA binding and activation of p53 target genes. We studied the effects of PRIMA‐1 and commonly used antileukaemic drugs on AML cells from 62 patients. Cells were obtained from peripheral blood or bone marrow and were exposed to PRIMA‐1, cytarabine, daunorubicin, chlorodeoxyadenosine and fludarabine. This study showed that PRIMA‐1 had cytotoxic effects on AML cells. Conventional AML drugs were less effective in cells with hemizygous p53 deletion. Interestingly, our data indicated that PRIMA‐1 was more effective in this subgroup of patients compared with patients with normal chromosome 17. Our data suggest that the concept of restoration of p53 protein by PRIMA‐1 or a PRIMA‐1‐based new drug may increase the efficacy of AML treatment in patients with p53 mutations.

Negligible clinical effects of thalidomide in patients with myelofibrosis with myeloid metaplasia.

We conducted a nonrandomized prospective phase II study of thalidomide in anemic patients with myelofibrosis with myeloid metaplasia (MMM), with or without preceding polycythemia vera or essential thrombocythemia, with a primary aim to improve anemia. Thalidomide was given in escalating doses with a target dose of 800 mg daily, but the median dose of thalidomide that was actually tolerated was 400 mg daily. Fifteen patients were entered into the study and 14 were evaluable for response. Five of 14 (36%) patients discontinued thalidomide before 3 mo because of side effects, and none of these five patients had a response at the time when thalidomide was stopped. When evaluated after 3 mo of therapy, none of the remaining nine patients exhibited a discernible clinical response. Three patients showed progressive disease defined as >50% increase in the need for red cell transfusions. Treatment was poorly tolerated, with all patients reporting side effects of thalidomide, the most prominent being fatigue documented in 80% of patients. Two patients died while on study, one from acute myelogenous leukemia and one from pneumonia. We conclude that thalidomide given in doses employed in the treatment of multiple myeloma gives no clinically relevant hematological effects in advanced MMM and is hampered by a very high incidence of side effects.