Partial Cytogenetic Remission Research Articles

Benefit of TKI-Treatment in CML after Failing Molecular or Cytogenetic Milestones

No information is available on long-term survival of tyrosine kinase inhibitor (TKI)- treated patients failing the response milestones as defined by the European LeukemiaNet (ELN). Such information would help with decision making in failing patients when risks associated with comorbidities, drug toxicities or transplantation preclude switching to another TKI or other treatments. To systematically analyze survival after reaching, or not reaching, ELN-response milestones, 1342 imatinib-treated patients from CML-study IV with newly diagnosed CML in chronic phase and regular molecular and cytogenetic tests were studied. Survival was analyzed by landmark analyses at 3, 6, 12 and 24 months and followed up to 14 years. In patients who failed the failure milestones (>10% BCR::ABL1 IS at 6 months, >1% BCR::ABL1 IS at 12 months), 10- to 12-year survival ranged around 70%, 10-20% less than in responding patients. An example is shown in Fig. 1a which depicts survival of patients reaching, or not reaching 1% BCR::ABL1 IS at 12 months. Similar survival curves were observed for the other landmarks that indicate failure. Inbetween response rates of >1%-10% BCR::ABL1 IS at 12 months indicated survival probabilities similar to those of patients failing 1% BCR::ABL1 IS, whereas patients reaching >1%-10% BCR::ABL1 IS at 24 months showed a survival similar to that of patients failing 10% BCR::ABL1 IS. Switching to alternative therapies, mostly to dasatinib and nilotinib, was observed in 372 patients (26.9%) and did not change the main results. Although cytogenetic tests were increasingly replaced by molecular tests in the course of the study, sufficient cytogenetic analyses were available for comparison with molecular analyses. Complete and partial cytogenetic remissions have been shown to be roughly equivalent to 1% and 10% BCR::ABL1 IS, respectively. Wheras reaching, or not reaching cytogenetic and molecular reponse markers at 6 and 12 months was equivalent, not reaching a complete or partial cytogenetic remission at 24 months identified a small subgroup of patients (5.9%) with inferior survival (Fig. 1b). Age (more or less than 60 years) had no major impact on survival differences, but on hazard ratios for early death. Hazard ratios at 10% BCR::ABL1 IS by 6 months were 4.0 (95%CI:2.0-8.1) for the younger patients, but only 0.9 (95% CI: 0.4-1.8) for the elderly. The reasons for early death in the younger patients were mostly CML-related, whereas the elderly primarily died of other reasons. The data show that TKI-treated patients not reaching failure milestones still may derive benefit from continuing TKI-treatment and provide a basis for individualized decisions, if failing patients are confronted with risks of alternative treatments.

Cytogenetic and Molecular Remission in Chronic Myeloid Leukemia in Togo

The availability of tyrosine kinase inhibitors (TKIs) in Togo through the GIPAP program has revolutionized the management of chronic myeloid leukemia and increased patient life expectancy. However, diagnosis and molecular monitoring of therapeutic efficacy remain a real challenge due to the inaccessibility of cytogenetic and molecular biology tools. We’ve conducted, a cross-sectional, descriptive study that ran from December 21, 2022, to January 20, 2023, with the aim to evaluate the cytogenetic and molecular remission in CML patients followed at the CHU Campus de Lomé in Togo. Patients diagnosed with chronic-phase CML who had been treated for at least three months with imatinib or dasatinib and had achieved complete hematological remission were included. Dosage of BCR-ABL transcript levels was performed by RQ-PCR in Seattle, USA, using Dried Blood Spot. The transcript detection limit was 0.003% i.e., MR4. A total of 38 patients were included, 68.4% of them were treated with imatinib and 31.6% with dasatinib. In terms of remission, 28.9% had not achieve a partial cytogenetic remission while 15.8% of patients had achieved a MR4 remission and 7.9% a major molecular remission. Major molecular remission was achieved at a mean follow-up time of 95 months for patients taking imatinib and 22 months for those taking dasatinib. Patients in MR4 were on imatinib after a mean of 68 months. All in one, our study showed a high rate of treatment failure. Better access to cytogenetic and molecular biology tools is needed to improve management.

Monitoring By Chromosome Banding Analysis (CBA) and FISH Of Circulating CD34+ Cells In Low-Risk MDS Patients Treated In The Le-Mon-5 Study With Lenalidomide Monotherapy Reveals 82% Cytogenetic Responders With Different Response – , Evolutionary -, and Remission Patterns and No Increased Karyotype Evolution (KE)

IntroductionBesides a more reliable and frequent measurement of cytogenetic response, it was our aim to find out whether lenalidomide treatment in patients with IPSS low- or intermediate I-risk MDS can foster karyotype evolution (KE) and thus increase the risk of leukemic transformation. Thus, it is an important goal of the Le-Mon-5 study to examine the cytogenetic course under treatment with lenalidomide. In this study, only lower risk MDS patients with an isolated del(5q) are included. MethodsWe performed a rigid initial pretreatment screening of bone marrow (bm) aspirates by chromosome banding (CBA) as well as FISH-analysis to ensure an isolated del(5q). For initial screening as well as frequent cytogenetic follow-up every two to three months (FISH analysis of immunomagnetically enriched CD34+ peripheral blood cells (PBC) = CD34+ pb FISH), we used panels of 8 to 13 FISH probes covering the most common aberrations in MDS (Braulke et al. Leuk Res, 2010, 2013). Using this method we intended a reliable surveillance of cytogenetic changes occurring during therapy. Complete cytogenetic remission (CCyR) was defined as: no metaphases with del(5q) and abnormal FISH result below the laboratory threshold (5% EGR1-loss in CD34+ PBC) and partial cytogenetic remission (PCyR) was defined as: >50% reduction of clone size, also including cases with a CCyR at only one time point and those with either only CBA-CCyR or FISH-CCyR. ResultsFrom the initially screened 145 MDS patients, 84 could be included in the study according to the study inclusion criteria. Currently, follow-up data (at least 2 different time points) are available for 67 patients. An ongoing (>1 time point) CCyR was observed in 22 (33%) patients after a median follow-up of 18 (6-33) months. Thirty-three patients (49%) had a PCyR. Considering only CD34+ pb FISH results 31 (46%) patients showed a complete molecular-cytogenetic remission.A cytogenetic response was observed after a median of 6 (2-12) months after initiation of therapy for a median duration of 10 (2-25) months. In twelve patients (18%) the size of the del(5q) clone did not change. The median del(5q)-clone size at start of therapy was 60% in responders. After 10 cycles maximum clone size reduction was achieved (n=26) (median: 2.6%; range: 0% - 33.6%), after 24 months of observation clone size in responders (n=22) still was reduced to a median of 10.8% (range: 0.5% -76%) (Figure 1). [Display omitted] In 10 of 67 patients (15%) a KE occurred in the del(5q) clone after a median time of 18 months (6-42). This is not increased compared to the rate of spontaneous KE (13%) in a representative MDS cohort (n=729) studied recently by our group by banding analysis (Cevik et al. DGHO 2013, accepted for oral presentation). Remarkably, in one patient a KE with loss of a p53 allele and a 20q- deletion resolved after the 8th cycle of lenalidomide and after the 10th cycle a complete cytogenetic remission was achieved (Figure 2). In two additional cases, the del(5q) clone was completely eliminated, however a new cell clone with independent abnormalities (t(3;3)(q21;q26), second case: inv(11)(p15q23)del(11)(q13q22)) emerged under lenalidomide. There is a growing body of evidence that these clones were preexistent initially but below the cytogenetic detection level and were possibly suppressed by the del(5q) clone until its cytogenetic remission. [Display omitted] ConclusionsOur interim results show that:•A cytogenetic response to lenalidomide can be achieved in >80% of pts. with isolated del(5q) and lower-risk MDS•CBA can be complemented reasonable by CD34+ pB FISH, which allows a reliable monitoring of response and KE•Cytogenetic response to lenalidomide is durable (median 10 months as yet)•KE is not increased in this patient group under treatment with lenalidomide•In one case with KE and acquisition of additional p53 allelic loss and 20q-deletion continuation of lenalidomide finally lead to a CCyR Disclosures:Platzbecker:Celgene: Honoraria. Nolte:Celgene: Honoraria, Research Funding. Giagounidis:Celgene: Honoraria. Götze:Celgene Corp: Honoraria. Schlenk:Celgene: Honoraria, Research Funding; Pfizer: Honoraria, Research Funding; Chugai: Research Funding; Amgen: Research Funding; Novartis: Research Funding; Ambit: Honoraria. Bug:Celgene: Honoraria, Research Funding. Germing:Celgene: Honoraria. Haase:Celgene: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding.

A short report on chronic myeloid leukemia from Post Graduate Institute of Medical Education and Research, Chandigarh

Post Graduate Institute (PGI) Chandigarh is a premier institute of North India. There are approximately 70,000 admissions per year. The adult clinical hematology department sees more than 2000 new patients per year. A preliminary analysis of 299 chronic myeloid leukemia patients registered from January 2001 until December 2007 was done. Out of these, 256 (86%) patients were in chronic phase (CP). The median age at presentation was 40 years. At 6 months of follow-up 95% of patients who were started on Imatinib mesylate based therapy remained in CP. Partial cytogenetic remission was seen in 69% of patients while complete cytogenetic response was seen in only 20% of patients at 6 months on Imatinib mesylate.

Utility of Peripheral Blood for Cytogenetic and Mutation Analysis in Myelodysplastic Syndrome

Abstract 1707With the advent of high throughput and high resolution techniques, >80% of myelodysplastic syndrome (MDS) patients harbour somatic mutations and/or genomic aberrations, which provide diagnostic and prognostic utility; however, frequent bone marrow (BM) aspirates are required. In a significant minority, the BM is hypocellular and fibrotic with suboptimal in vitro growth and, additionally, the procedure causes discomfort particularly in the elderly. This led us to investigate the use of peripheral blood (PB) and serum to identify and monitor BM derived genetic markers using high resolution single nucleotide polymorphism array (SNP-A) karyotyping and 454 parallel sequencing (454-PS) of a 22 gene myeloid panel comprising of all the exons of DNMT3a, RUNX1, CEBPα, TP53, EZH2, TET2 and ZRSR2 and mutations ‘hotposts' for NPM1, FLT3, ASXL1, IDH1, IDH2, MPL, JAK2, BRAF, cCBL, NRAS, KRAS, C-KIT, SF3B1, SRSF2, and U2AF1.We selected 23 MDS patients with concurrent BM and PB samples and detected 45 mutations in TET2, SF3B1, ASXL1, TP53, DNMT3a, FLT3, U2AF1, NRAS, cCBL, JAK2 and IDH2 in BM and subsequently analysed their PB using 454-PS with independent validation performed by Sanger sequencing (SS). All the mutations identified in BM were detected in PB with the exception of a single NRAS (BM-11%). Nine patients had a single mutation in SF3B1, ASXL1, TP53, TET2, DNMT3a and U2AF1 with the remaining patients having multiple coexisting mutations. Overall there was no significant difference in the mutation burden between the PB (median 25%(1.5%-50%)) and the BM (median 33%(5%-68%)). Concurrent analysis of unamplified and whole genome amplified PB DNA from 3 patients with mutations in TET2, U2AF1, ASXL1 and NRAS showed no difference in their mutation profile. As expected, in three patients post therapy the mutation burden in the PB was lower than in the pre-treatment BM sample.The lower mutation burden and sequence context in the PB contributed significantly to the quality of SS analysis. Prior knowledge of the mutation site resulted in 98% concordance (smallest clone size - 1.5%) between BM and PB, however, a blind approach decreased this to 84% (smallest clone size - 15%). In addition, serum was available from 14 patients (22 known mutations in BM) and SS of serum DNA identified 12 mutations correctly (U2AF1, FLT3, SF3B1, TET2, TP53, ASXL1, DNMT3a and IDH2, 4 as wildtype (TET2, cCBL, ASXL1 and SF3B1) and 6 failed to amplify (ASXL1, TP53 and TET2) without any preference for specific genes and the failure attributed to the quantity of serum DNA.Karyotype aberrations in PB were assessed using Affymetrix SNP 6.0 arrays on 31 MDS patients; normal karyotype (n=11), del5q (n=9), del7q/-7 (n=5), trisomy 8 (n=2), complex (n=2), isodiXq13 (n=1) and t(2:4)(q33;q27). An overall karyotype concordance of 94% was observed in PB with the 2 discordant cases showing normal karyotype in PB and BM by SNP-A but having monosomy 7 (partial cytogenetic remission after 5-azacitidine) and t(2:4)(q33;q27) respectively in their BM by MC. The mean copy number (CN) in the PB was lower than BM (PB vs BM); deletions (CN of 1.8 vs 1.6) and gains(CN of 2.2 vs 2.4), implying a smaller abnormal clone in the PB. Concurrent SNP-A karyotype from BM from 9 patients was concordant with SNP-A karyotype from PB, although a patient with complex karyotype determined by SNP-A in the BM and having 30 aberrations had only 15 aberrations detected in the PB.To determine if the 5q deletion was lineage restricted, we enriched PB for CD3+, CD19+ and CD3-CD19- populations from 4 patients. FISH and SNP-A karyotyping showed the presence of the 5q deletion using both techniques in all three fractions, however at a lower level in PB lymphocytes indicating the presence of a smaller clone.In conclusion, our study showed an excellent concordance between BM and PB, both for karyotype and mutational analyses using high resolution SNP-A karyotyping and 454-PS suggesting its clinical utility as a surrogate for BM, thereby, avoiding the discomfort of repeated BM aspirates and help in monitoring response to therapy more frequently. Disclosures:No relevant conflicts of interest to declare.

Loss of the Y Chromosome Predicts Lower Complete Cytogenetic Remission Rate in Patients (pts) with Chronic Myeloid Leukemia (CML)

AbstractAbstract 4804 Background:Chronic myeloid leukemia (CML) is one of the classic myeloproliferative neoplasms characterized by a reciprocal translocation of BCR and ABL t(9;22)(q34;q11). Tyrosine kinase inhibitors (TKI) have revolutionized the management of CML in inducing rapid and prolonged responses. However; clonal evolution (CE) is considered a poor prognostic factor and a criterion for accelerated phase (AP) CML by the World Health Organization (WHO). Deletion of chromosome Y (−Y) is frequently considered an age-related abnormality and the exact prognostic value has not yet been determined. Aim:To determine if –Y carries an impact on the clinical outcome of male pts with CML. Methods:All male patients diagnosed with chronic phase CML in our institution between 1993 and 2011 were screened for -Y. Data were collected in a retrospective manner and compared (using t-test) to male patients with sole BCR-ABL translocation after excluding patients with advanced stages (accelerated phase, blast phase). Demographics, laboratory tests, cytogenetic analysis, molecular testing and survival data were abstracted. Kaplan-Meier estimates of overall survival were used via JMP software v9.0. Institutional Review Board (IRB) approval was obtained for this study in accordance with the Helsinki Declaration. Results:20 of 162 (12%) males with CML were found to have –Y abnormality (group 1). CML male patients with sole Philadelphia chromosome abnormality were the control cohort (group 2). In group1; the median age was 57 years, BMI 27.7, hemoglobin 12.2 g/dL, white blood cell count (WBC) 32.8 x109/L, platelet 270 x109/L, and peripheral blood blasts 1%. Sokal risk was low in 30%, intermediate in 65% and high in 5% of pts. Nine pts (45%) were treated with interferon (IFN) prior to TKI. In group 2; the median age was 54 years, hemoglobin 12 g/dL, WBC 57 x109/L, and platelet 282 x109/L. Sokal risk was low in 37%, intermediate in 47%, and high in 16%. 46 of 142 patients (32%) had received previous interferon therapy. All patients in both groups had chronic phase CML at the time of diagnosis, with a median bone marrow cellularity of 95%. In group 1, 14 of 20 pts (70%) received imatinib, all of whom achieved a complete hematological response (CHR), 7 of 14 pts (50%) had partial cytogenetic response, 2 of 14 pts (14%) achieved a complete cytogenic response (CCyR);1 (7%) pt achieved CCyR within 12 months and an additional 1 (7%) by 18 months). Two pts of 12 (16%) achieved at least a major molecular response (MMR); one of whom (8%) achieved a complete molecular response (CMR). In comparison, 107of 142 pts (75%) in group 2 received imatinib, all of whom achieved CHR. Twenty-one pts (20%) achieved partial cytogenetic remission. CCyR was more frequently achieved than group 1 (48/107 pts (45%), p 0.026); 24 pts (22%) achieved CCyR within 12 months of therapy and an additional 10 pts by 18-months. MMR and/or CMR was higher in group 2 compared to group 1 (34 /101 (34%), p 0.18); 16 (16%) of which were CMR. In group 1; 6 (30%) pts had disease progression; 4 of 20 pts (20%) progressed to blast phase (BP) and 2 pts (10%) progressed to AP, compared to 32 (22%) pts in group 2 (p 0.17); 22 (15%) of whom progressed to BP and 10 (7%) patients progressed to AP. Median overall survival was 110 months in group 1 compared to 155 months in group 2 (log rank p=0.48). On multivariate analysis, CCyR was an independent factor for a better OS (p 0.03), but not –Y (p 0.7). Conclusion:Loss of the Y chromosome in chronic myelogenous leukemia is an infrequent phenomenon (12%). Although patients with –Y had a statistically significant less chance to achieve CCyR, loss of the Y chromosome did not affect the progression rate or overall survival. Larger scale studies are needed to confirm our observations [Display omitted] Disclosures:No relevant conflicts of interest to declare.

Interest of the Association Azacitidine-Lenalidomide As Front Line Therapy in High-Risk Myelodysplasia (MDS) or Acute Myeloid Leukemia (AML) with Complex Karyotype

Abstract 5035 Background.Results of intensive chemotherapy (ICT) in myelodysplasia (MDS) or secondary acute myeloid leukaemia (sAML) are poor, particularly in the presence of complex cytogenetic. Allogeneic haematopoietic stem cell transplantation (HSCT) remains the only curative treatment but only a minority of patients is eligible for HSCT notably because of difficulties to assess disease control underlining the need for novel strategies. Although, the use of hypomethylating agents has improved the overall survival (OS) of MDS with excess of blasts and might be interesting in AML (Fenaux, Lancet Oncol 2009), the prognosis of high risk MDS and AML remains poor (Itzykson, Blood 2011). Lenalidomide has demonstrated its efficiency in MDS with del5q and, has also been tested in patients with high-risk MDS or AML (Ades, Blood 2009). Aim.We report our experience of the association of azacitidine and lenalidomide in patients presenting with high risk MDS or sAML and who were unfit for ICT. Patients and treatment.Eight consecutive patients were referred to our haematological department from August 2008 to March 2010 with a diagnosis of high-risk MDS or sAML because of the presence of a complex karyotype including in all cases abnormalities of the 5q chromosome. At diagnosis, median age of patients was 66.5 years (range, 23–76); median WBC was 2×10.9/L (range, 0.11–7.1) and median marrow blast was 23% (range, 11%-37%). Four patients had secondary therapy-related AML and four patients had type-2 refractory anemia with excess of blasts (RAEB). None of them was eligible for an intensive chemotherapy because of age and/or poor performans status and/or comorbidities. They received cycles of 28 days of azacitidine (75 mg/m2 Day 1 to 5 or 7 SC) and lenalidomide (10 mg per day, day 1 to 14 or 21, oral) as first line treatment for a median of 4.5 cycles (range, 1–13 cycles). Results.Main side effects were cytopenias: 7 patients on the 8 developed grade 4 thrombocytopenia. Five patients, 4 with baseline ANC more than 1000/mm3 developed grade 4 neutropenia. Severe infections occurred in neutropenic patients: 6 sepsis (4 fevers of unknown origin, 1 facial cellulitis, 1 fungal infection and 1 CMV reactivation.) including 2 septic shock. However, no death was related to the toxicity of the treatment.Of the eight patients, 6 achieved a haematological response (3 complete response (CR), 3 partial response (PR)) and 2 progressed under treatment. Among the 6 responding patients, initial responses occurred at a median of 10 weeks (range, 6–28 weeks) from the beginning of therapy. Two patients were allografted, one with a reduced intensity conditioning in haematological and cytogenetic CR at transplant, and the other one with a sequential type reduced intensity conditioning in complete haematological but partial cytogenetic remission at HSCT. With a median follow-up of 60 weeks for the responding patients, (range, 0–72), the 4 responding patients who were not allografted because of age did experience relapse at a median of 12 weeks (range, 8–36 weeks) from initial response while the two patients who were allografted are still alive in CR at 15 and 14 months after HSCT, respectively. Considering the whole population, with a median follow up of 18 months, the median OS was 15 months (range, 1–18 months), and median progression-free survival (PFS) was 9.5 months (range, 0–18 months). Conclusion.The association of azacitidine and lenalidomide in the context of high risk MDS and sAML might offer a significant probability of remission with tolerable toxicity. For patients considered for an allogeneic HSCT, the use in first line therapy of such novel combination strategy rather than intensive chemotherapy represents an interesting option particularly in the case of complex cytogenetic. Disclosures:Off Label Use: Interest of the association Azacitidine-Lenalidomide as front line therapy in high-risk myelodysplasia (MDS) or acute myeloid leukemia (AML) with complex karyotype.

A Phase I Study of a Combination of 5-Azacyitidine Followed by Lenalidomide In High-Risk MDS or AML Patients with Chromosome 5 Abnormalities – Interim Results of the “AZALE” Trial

AbstractAbstract 4000Lenalidomide has shown single agent activity in patients with MDS (Myelodysplastic Syndromes) and a del(5q) cytogenetic abnormality. Further, studies with the DNA methyltransferase inhibitor 5-azacytidine (5-aza) have been conducted in high-risk MDS (IPSS INT-2 or HIGH) and patients with acute myeloid leukemia (AML) resulting in considerable responses with a low rate of extramedullary toxicity compared to conventional induction chemotherapy (IC). Given the poor outcome of high-risk MDS and AML patients with chromosome 5 abnormalities, there is a significant clinical need to perform studies with new regimens in this patient population.We report first results of an ongoing phase I clinical trial evaluating the maximum tolerated dose (MTD) of lenalidomide in combination with 5-aza in patients with either high-risk MDS, refractory/relapsed AML or de novo AML not eligible for conventional IC with chromosome 5 abnormalities including monosomy 5 or del(5q). Given the mechanism of action of both drugs and also in contrast to a recent study in non-del(5q) MDS patients, a sequential approach was chosen. In fact, induction therapy consisted of 5-aza (75mg/m2 days 1–5) followed by increasing doses of lenalidomide (starting with 10mg p.o., days 6–19). In patients achieving a complete remission this was followed by a combined maintenance therapy every 8 weeks until disease progression.To determine the MTD, a standard “3+3” design was used. The dose limiting toxicity (DLT) is determined during the first cycle only and is defined as either inability to deliver the full dosing schedule of lenalidomide due to any ≥ Grade 3 non-hematologic toxicity or absence of hematological recovery after completing the 1st cycle despite complete marrow blast clearance or treatment delay of ≥ 4 weeks as a result of unresolved grade 4 non-hematological toxicity.Of 8 patients currently enrolled, median age was 67 years (range, 45 to 74 years), interval from primary MDS or AML diagnosis was 9 months (range, 1 to 100 months). IPSS categories were INT-2 (n = 1) and HIGH (n = 3) whereas 4 patients were included with advanced AML. It is of note, that all but two patients had a complex karyotype including a del(5q) abnormality. Prior treatment included IC (n=1), IC plus allogeneic HSCT (n=3) and/or single agent 5-aza (n=3) while 4 patients had received supportive care only prior to study entry.A median of 2 induction cycles were administered. During the first cycle of cohort I (10mg lenalidomide) and cohort II (15mg lenalidomide) grades 3 to 4 non-hematologic toxicities included febrile neutropenia (n = 3), enterocolitis (n = 1) and pneumonia (n=3) whereas therapy-induced grade 3–4 neutropenia or thrombocytopenia occurred in four and five patients, respectively. The MTD has not been reached yet. One patient (12.5%) with AML showed rapid progression while receiving the 1st cycle. Out of the remaining seven patients, one (12.5%) achieved a marrow CR together with a partial cytogenetic remission, and six patients (75%) had stable disease. Interestingly, two out of these achieved a partial cytogenetic remission.These preliminary data of an ongoing phase I trial demonstrate the safety and the potential of a combination of 5-aza and lenalidomide in patients with advanced MDS or AML and a del(5q). Disclosures:Platzbecker:Celgene: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding. Haase:Celgene: Honoraria, Membership on an entity's Board of Directors or advisory committees. Götze:Celgene: Honoraria, Membership on an entity's Board of Directors or advisory committees. Kuendgen:Celgene: Honoraria, Membership on an entity's Board of Directors or advisory committees. Giagounidis:Celgene: Honoraria, Membership on an entity's Board of Directors or advisory committees. Hofmann:Celgene: Honoraria, Membership on an entity's Board of Directors or advisory committees.

Significance of Bone Abnormalities In CML Patients on Imatinib Mesylate Therapy

AbstractAbstract 4759Both Dasatinib and Nilotinib have proven themselves as more effective but costly first line therapy in chronic myeloid leukemia (CML) than imatinib mesylate (IM). Hence, IM would continue to be used as first line therapy in developing countries because of economic reasons. Thus simple diagnostic tools are required which can triage CML patients into different risk categories. Presence of bone abnormalities (osteoblastic, osteolytic and mixed) in CML was generally associated with poor outcome in pre-imatinib era. Nuclear bone scintigraphy is a simple test that has universal availability. The significance of these abnormalities in imatinib-era has not been investigated. We proposed to evaluate the significance of bone abnormalities as seen on nuclear scintigraphy in CML at diagnosis and at follow-up. The bone scintigraphy was performed with 99m Tc methylene di phosphonate 3-hours after an intravenous injection of 740 MBq of the radiotracer. Regions of interest (ROI) were marked using standard software. A second ROI was marked on the diaphyseal region of the involved long bone. The ratio of average counts per pixel between the involved and the normal areas was calculated for each long bone. The average ratios of the right and left half of body were calculated separately. This average scintigraphic quantification ratio were compared before and after IM therapy. The standard investigations including bone marrow examination and cytogenetics studies were done at diagnosis and at follow-up. The bone scintigraphic abnormalities were correlated with cytogenetic remission after 6 months of IM therapy. The starting dose of IM in chronic phase was 400 mg/day and in advanced phase 600 mg/day. The study was approved by institute review board and was carried out according the declaration of Helsinki. Forty-three newly diagnosed CML patients gave consent for the study. The mean age of patients was 40.8+14.6 and male female ratio was 25:18. Thirty-two patients were in chronic phase and 11 were in advanced phase (accelerated or blast crisis). The prevalence of bone scintigraphy abnormalities were found in 76.7% patients (75% in chronic phase and 81% in advanced phase, p =0.06). No bone abnormalities were detected in 10 patients. At a median follow-up of 7 months (range 5–8 months of IM therapy), 16% patients achieved completed cytogenetic remission (CCR) and 80% achieved partial cytogenetic remission (pCR). The average bone quantification decreased from 3.34+1.05 to 2.07+0.44 among patients in CCR and 2.47+0.57 among patients in pCR. One CML patient showed increase in bone quantification from 3.61 at baseline to 4.14 at 6 months and on bone marrow examination, he was diagnosed in blast crisis. No relationship of bone abnormalities was found with Sokal score at baseline. Bone abnormalities as seen on nuclear scintigraphy are thus common among patients with CML. They tend to resolve on IM therapy. However, a longer follow-up of patients is required for further evaluation of this technique in CML patients and whether it can replace sophisticated prognostic molecular techniques which are inaccessible in resource-constraints settings. Disclosures:No relevant conflicts of interest to declare.

Coexistence of chronic myeloid leukemia and pulmonary plasmacytoma mimicking primary lung cancer

A 61-year-old man was diagnosed with the simultaneous occurrence of chronic myeloid leukemia (CML) and infiltrative intrathoracic plasmacytoma, radiologically mimicking bronchogenic carcinoma. Following the administration of imatinib mesylate (IM; 400mg/day), both hematologic and partial cytogenetic remission of CML were achieved. However, the pulmonary plasmacytoma was persistently aggravated. High-dose dexamethasone was added to the IM therapy because the patient refused radiotherapy to control the aggravated pulmonary plasmacytoma. Finally, he died due to pneumonia and multi-organ failure during concurrent administration of IM and high-dose dexamethasone.

Discontinuation of Lenalidomide in Patients with Transfusion-Dependent Low- and Intermediate-1 Risk Myelodysplastic Syndromes with Del(5q): Sustained Remissions, but Not Cure.

Abstract 3807Poster Board III-743 BackgroundLenalidomide has high erythroid remitting activity in patients with del(5q) myelodysplastic syndrome (MDS). The actual recommendation is to continue drug administration until relapse of red blood cell transfusion dependence. We address the question whether it is safe and advisable to discontinue lenalidomide after achieving a hematologic and/or cytogenetic response Patients13 patients (8 female, 5 male) were treated with lenalidomide between December 2003 and January 2007. Median age was 69 years. All had low- or intermediate-1 risk MDS with del(5q) chromosomal abnormality. All patients were red blood cell (RBC) transfusion dependent. All patients started treatment with 5 or 10 mg lenalidomide for 28 out of 28 days or for 21 out of 28 days. Discontinuation of lenalidomide was usually due to patient's choice, and occurred after achievement of RBC transfusion independence. Lenalidomide administration duration ranged from 6 weeks to 24 months ResultsFour patients had a total lenalidomide exposure of > 12 months (12, 15, 18 and 24 months), seven patients between 4 and 12 months and two patients had lenalidomide exposures of <4 months (3,5 and 1 month). Five patients achieved partial cytogenetic remission, and eight patients complete cytogenetic remission (CCR), i.e. disappearance of all del(5q) metaphases in conventional cytogenetics. Within the five patients with partial cytogenetic remission, both patients with very short lenalidomide treatment (1 and 3.5 months) achieved transfusion independence of 21 months duration. At relapse, one was progressing to RAEB-1. One patient who received lenalidomide for four months remained RBC transfusion independent for 12 months. At relapse, she resumed lenalidomide for 6 months and became transfusion independent again (16+ months). One patient received lenalidomide for eight months and remains transfusion independent after 26+ months with normalized and stable hemoglobin value. One patient received lenalidomide for 12 months and remains transfusion independent for 30+ months, but hemoglobin levels have fallen recently to 11 g/dL, indicating imminent relapse. Eight patients achieved complete cytogenetic remission: Two administered lenalidomide for 24 months, one for 18 months, three for 15 months, one for 12 months and one for 9 months. Duration of RBC transfusion independence in these patients was dependent on the length of lenalidomide administration after achievement of CCR: One patient stopped lenalidomide at the time of CCR. She remains transfusion independent after 18+ months, but del(5q) karyotype has reoccurred after 12 months. Three patients stopped lenalidomide two, four, and seven months after CCR. Transfusion-independence is ongoing at 34+, 36+ and 38+ months. In all patients del(5q) has reoccurred. Three patients discontinued after >12 months of CCR achievement. They are transfusion independent for 27+ months, 41+ months, and 66+ months. The latter patient had reoccurrence of one metaphase out of 20 with del(5q) after 60 months. Finally, one patient took lenalidomide only six months beyond CCR and remains in transfusion independence for 70+ months. However, this patient was trated with a fludarabine-containing regimen for follicular lymphoma 48 months ago DiscussionDiscontinuation of lenalidomide seems to be feasible in patients with del(5q) MDS, preferably in those who achieve CCR. Best results regarding long-term transfusion independence and long-term complete cytogenetic remissions are seen in patients who continue lenalidomide for 12 months after CCR achievement. Disclosures:Giagounidis:Celgene: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau. Göhring:Celgene Corp.:. Schlegelberger:Celgene Corp.:. Kuendgen:Celgene: Honoraria. Platzbecker:Celgene: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding.

A Retrospective Study of Introferon-Alpha Treatment on Chinese Patients with Chronic Myeloid Leukemia

Introduction: Chronic Myeloid Leukemia (CML) is one of the myeloproliferative malignancies characterized by the reciprocal t(9;22)(q34;q11) chromosome, which causes the generation of Bcr-Abl fusion protein. Interferon-alpha (IFN-alpha) was introduced into clinical application from 1980s' and showed its treatment benefit mainly in chronic phase CML (CML-CP) patients including achieving hematological remission, prolonging survival. Our retrospective study aimed to evaluate its clinical efficacy as well as toxicities in Chinese CML-CP patients.Methods: This is a retrospective, multicentric clinical trial. Chinese CML patients in chronic phase received IFN-alpha treatment from 1992 to 2006 were enrolled. Other agents such as hydroxyurea or cytarabine treatment were permitted during treatment excepted for tyrosine protein kinase inhibitors.Results: 1037 CML-CP patients were into analysis, the median age was 42 yrs (range 9–80) and male: female ratio of 1.6:1. The median time for interferon treatment was 12 months (range from 1 month to 136 months). 86.32% of patients received 300Mu of interferon, three times a week, 10.88% in 300Mu of daily injection, while another 2.8% in 500Mu of interferon treatment, three times a week. Main of additional treatment included hydroxyurea, which was used in 8.9% of patients, and cytarabine treatment in 3.3% of patients. We evaluate the best response reaction during IFN-alpha treatment, 496 out of 1037 patients obtained hematological remission (47.8%). 33 in complete cytogenetic remission (CCR, 3.2%) but unfortunately 60% of them were lost CCR after median of 6–11 months; 89 in partial cytogenetic remission (8.6%); 40 in major cytogenetic remission (3.9%); 301 in minor cytogenetic remission (29%); 78(7.5%) of CML-CP patients worsen into accelerate phase or blast crisis. Non-Hematological toxicities during IFN-alpha treatment mainly included flu-like symptom (24.9%) and fatigue (23.5%) which were transient and most of patients got recovery after second injection whereas 20% of them could not endurable. While hematological toxicities were neutropenia (22.1%, 33% of them were in grade III or IV), and thrombocytopemia (16.9%, 33% of them in grade III or IV).Conclusion: The IFNalpha is one candidate agent for CML-CP patients in China with moderate toxicities during treatment, but it has little influence on achieving or maintaining complete cytogenetic remission.

Long-Term Cytogenetic Follow-up of MDS Patients with 5q- Treated within the MDS-003 (CC-5013-MDS-003) Study: Evolution to Complex Clones and Progression to AML.

Lenalidomide is the first drug to consistently induce transfusion independence and complete cytogenetic remissions in MDS patients with deletion of 5q. However, recently concerns were raised whether lenalidomide treatment may increase the risk of leukemic transformation. The selective activity of lenalidomide against the deletion 5q clone may allow pre-existing aberrant clones to expand if the dominant deletion 5q clone is eradicated in a circumstance in which few normal stem cells remain. This conditional selection of pre-existing clones (M. Cazzola, Haematologica 2008; 93:967–72) could represent a potential mechanism for leukemic transformation. We therefore analyzed 22 European patients with transfusion-dependent anemia due to low- or intermediate-1-risk deletion 5q MDS treated with lenalidomide who were enrolled in the MDS-003 (CC-5013-MDS-003) study between 12/2003 and 2/2004. For these patients, sufficient cytogenetic, morphologic and clinical follow-up data were available. Ten of these patients achieved a complete (n = 6, 3/6 transiently) or partial (n = 4, 3/4 transiently) cytogenetic remission and 11 patients (5/11 transiently) achieved an erythroid response resulting in transfusion independence. Seven of the 22 patients (32%) developed complex clones containing typical aberrations like der(7q), +8, del(12p), del(17p) and +21 within 5 to 42 months (median 19 months) after study entry, and within 15 and 72 months (median 44 months) after diagnosis. Among these 7 patients, the findings at baseline included that 2 patients had the deletion 5q plus additional chromosomal aberrations, i.e. del(12p) and trisomy 21, respectively, 2 patients had an isolated deletion 5q karyotype and low-risk MDS, and 3 patients had an elevated blast count ≥ 5%. Six of the these 7 patients progressed to AML, and one patient lost the complex clone with continued lenalidomide treatment, showing the isolated deletion 5q again, and achieved a partial remission from RAEB-1 to RCMD. Both patients with low risk MDS and isolated deletion 5q developed complex clones 12 and 9 months after study entry and 19 and 24 months after diagnosis, respectively, and progressed into AML. Moreover, two patients without cytogenetic response progressed to AML 5 and 6 months after study entry, and 15 and 72 months after diagnosis, respectively. In total, 8 patients (36%), 2 of them with 5q- syndrome, 4 with RCMD and 2 with RAEB-1 according to the WHO classification, progressed into AML. FISH analyses were performed retrospectively to clarify whether complex karyotypes were already present at the time of diagnosis. In no case were such aberrations detected. The FISH data also showed that 13% to 84% (median 50%) of the cells contained a del (5q) abnormality, demonstrating that in all patients who underwent karyoptypic evolution, a substantial number of normal cells was present in the bone marrow. Overall, these data show that karyotypic evolution of 5q- clones into complex clones plays a significant role in the progression of low and int-1 MDS with 5q- into AML. No evidence of conditional selection by FISH analyses was observed. However, the presence of small complex clones at baseline may have been missed because FISH has a detection limit of up to 10%. It remains unclear whether the development of complex clones occurs during the natural course of the disease, whether lenalidomide treatment increases chromosomal instability or whether pre-existing clones are selected during treatment. Clearly, these findings warrant careful follow-up of MDS patients treated with lenalidomide and show that a multi-center, double-blind, placebo-controlled study with a long observation time is needed to define long-term safety and efficacy of lenalidomide.

Safety and Efficacy of a Combination of Imatinib and Pegylated Interferon α2a (Pegasys®) in Newly Diagnosed Patients with Chronic Myelogenous Leukemia - A Phase II Cohort Study.

Imatinib (IM) and interferon α (IFN) are active in the treatment of CML patients (pts); synergistic effects of both drugs were demonstrated in vitro. To define efficacy and tolerability of a combination therapy with IM and pegylated IFNα2a (PEG-IFN), a phase II cohort study was performed in 40 newly diagnosed Ph+ chronic phase CML pts (25 m, 15 f; median age 52 yrs, range 22–74). Treatment was commenced at a median of 0.7 mo (0.1–2.5) after diagnosis. 18 pts (45%) were of low, 21 pts (53%) of intermediate, and 1 patient (2%) of high risk according to Hasford. Objectives were to determine efficacy and safety of PEG-IFN (sc) given in different combination schedules with IM (po) and to explore myeloblastin (proteinase 3, PR3) expression and its correlation with PR3-specific T-cells (PR3-CTL) as a putative surrogate marker for IFN response. After cytoreduction with hydroxyurea, pts in cohort I (n=20) received PEG-IFN-monotherapy (6 wks), then IM monotherapy (6 wks), followed by the combination. Cohort II pts (n=20) started with IM monotherapy (6 wks), followed by the combination. Target dose for IM was 400 mg/d, for PEG-IFN 180 μg/week during mono- and 135 μg/week during combination therapy. Dose and schedule were adjusted according to hematological and non-hematological side effects. After 48 and 96 wks, 28 (70%) and 35 pts (88%) achieved a complete (CCR), 6 and 1 pts a partial cytogenetic remission, respectively. In 5 pts, treatment was stopped due to side effects (n=4) or lack of efficacy (n=1). Thus, 33/40 pts (83%) actually had CCR at 96 wks. Major molecular response (MMR; BCR-ABL ratio of ≤0.1% according to the international scale) was achieved in 21 (53%) and 26 pts (65%) after 48 and 96 wks, respectively. Non-hematologic grade 3 toxicity included exanthema (n=4), increased transaminases (n=3), flu-like symptoms (n=1), and depression (n=1). Grade 3 leukocytopenia occurred in 10 pts, grade 3/4 thrombocytopenia in 5 and 1 pts, respectively. Overall, pts received 42% of the scheduled PEG-IFN and 87% of IM doses. Expression levels of PR3 mRNA were determined in 264 samples from 19 pts by a quantitative PCR assay. Median ratios PR3/glucose-6-phosphate dehydrogenase (in %, range) were 16 (0.068–1700) at baseline, 6.1 (0.26–840) after 6 wks PEG-IFN mono, 0.093 (0.0088–1.4) after 6 wks IM mono, 0.054 (0.02–150) and 0.091 (0.007–250) after 48 and 96 wks of the combination therapy, respectively. PR3 expression was significantly reduced by IM, but not by upfront PEG-IFN (p=0.001). PR3-CTL were prospectively monitored by a PR3 specific tetramer assay in 113 peripheral blood samples from 17 HLA-A2+ pts. Autoreactive PR3-CTL were detected at baseline in 2/14 pts (0.18 and 0.23%), after PEG-IFN monotherapy in 3/6 pts (0.04, 0.11, and 0.35%), and during combination therapy in 7/16 pts (peak levels median 0; range 0–1.5%). PR3-CTL increased after initial PEG-IFN, decreased after commencement of IM and were associated with a higher PR3 antigen load. In conclusion, the combination treatment of IM and PEG-IFN is well tolerated and results in a high rate of CCR and MMR. However, IM may modify PEG-IFN immunotherapy, for example by inhibition of autoantigen expression. Consecutive vs simultaneous treatment schedules may affect efficacy of IM/PEG-IFN combination therapies.

FISH-Analyses of Cirulating CD34+ Cells in MDS-Patients - A Suitable Method To Measure and Predict Response to 5-Azacytidine.

In this study we present a novel method to measure response to treatment (here 5-Azacytidine (5-Aza)) that is less invasive and can be more frequently performed than bone marrow biopsy (bmb). Usually chromosomal aberrations are proven in bone marrow cells by classical cytogenetics. Most of them are suitable for a detection by FISH-techniques. In our study we have followed chromosomal anomalies in circulating CD34+ cells from peripheral blood in patients (pts) with MDS treated with 5-Aza. We could observe that the size of an abnormal clone in general is larger in circulating CD34+ cells as compared to non-enriched bone marrow cells. The percentage of circulating CD34+ cells showing the respective chromosomal anomaly is decreasing in response to therapy and therefore can be used as an indicator for response. Fourteen pts receiving 5-Aza are included in our ongoing study as yet. Thirteen are male, 1 is female, the average age at the time of diagnosis is 67 years (49–81 years). Starting 5-Aza 7 pts were diagnosed as AML, 3 as RAEB-2, 1 as RAEB-1, 1 as RCMD, 1 as CMML, 1 as RA. In all pts initial karyotyping of bone marrow cells was performed. There was 1 pt with normal karyotype, 5 pts (35.7%) showed a complex aberrant karyotype. The other pts had 1–2 chromosomal anomalies (+8, −7/7q-, 20q-, 5q-). All pts with clonal anomalies (13/14) were informative for further FISH-analyses. In average pts received 4 cycles 5-Aza (range 2–8 cycles). Before every therapy cycle circulating CD34+ cells from peripheral blood were enriched by immunomagnetic cell sorting (MACS) and analysed by FISH. Modified IWG-criteria were used to evaluate hematologic response. At present 10 pts could be analysed completely. All of them showed hematologic and/or cytogenetic response to therapy: 4 pts achieved complete remission (CR), 2 of them after allogeneic bone marrow transplantation. Remarkably, 3 out of 4 evaluable pts with complex aberrant karyotypes responded to 5-Aza. One further pt reached partial remission (PR), one stayed in stable disease (SD). Two pts showed at least a marrow response (PR/CR) without developing other improvements. Two further pts achieved at least hematologic improvement (HI). Remarkably, 9 out of 10 pts showed chromosomal aberrations suitable for FISH. Seven (77.8%) developed partial cytogenetic remission (cyPR) measured in circulating CD34+ cells. Six out of 7 pts with cyPR reached HI and/or CR. One was lost for follw up. CyPR preceded hematologic response as well as progression by 4–8 weeks. We conclude that 5-Aza leads to good results even in high-risk MDS-pts with complex aberrant karyotypes. Our data thus imply that analysing circulating CD34+ cells from peripheral blood by FISH techniques is a suitable method to monitor and to predict response to therapy with 5-Aza and possibly other agents in MDS-pts. It might be the more sensitive method to measure the aberrant cell clone than analysing non-enriched bone marrow cells. It could preserve pts from repeated bmb and allows a very frequent monitoring of therapy response.

Philadelphia chromosome mosaicism at diagnosis in chronic myeloid leukemia: Clinical correlates and effect on imatinib mesylate treatment outcome

In chemotherapy-treated patients with chronic myeloid leukemia (CML), the karyotypic detection of Philadelphia chromosome (Ph)-negative metaphases at diagnosis (i.e. Ph mosaicism) is not considered significant as a prognostic factor for survival. In the current retrospective study, clinical correlates and prognostic relevance of Ph mosaicism were evaluated in 63 Ph-positive patients with CML, including 59 in chronic phase and 4 in accelerated phase, receiving imatinib mesylate as either first (n = 46) or second (n = 17) line therapy. Thirteen patients (21%) displayed Ph-negative metaphases at diagnosis and, compared to the other 50 patients with 100% Ph-positive metaphases, presented with significantly lower leukocyte count (p = 0.0004), circulating blast percentage (p = 0.02), and incidence of palpable splenomegaly (p = 0.02). Ph mosaicism did not correlate with other CML-pertinent prognostic factors including Sokal score (p = 0.4) or the presence of additional chromosome changes (p = 0.96) found in 10 patients (16%). Neither Ph mosaicism nor the presence of additional chromosome changes affected complete or partial cytogenetic remission rates to IM. Multivariable analysis identified Ph mosaicism as a risk factor for shortened survival. Due to the small sample size, the current preliminary observations require validation in a larger group of patients.

Additional Cytogenetic Abnormalities and/or Philadelphia Chromosome Metaphase Mosaicism Might Adversely Influence Survival and Imatinib Response in Chronic Myeloid Leukemia.

Background: Chronic myeloid leukemia (CML) is invariably associated with the reciprocal translocation of BCR and ABL to form the Philadelphia chromosome (Ph), t(9;22)(q34;q11). At diagnosis, a small proportion of patients display additional cytogenetic abnormalities and/or a variable proportion of cytogenetically normal metaphases that coexist with Ph-positive metaphases. The objective of the current study was to examine the prognostic relevance of these two scenarios.Methods: The study population consisted of 65 (47.7% female) consecutive, newly diagnosed CML patients seen at the Mayo Clinic with a median age of 59 years. 44.5 percent were initially treated with interferon alpha (IFN) resulting in complete cytogenetic remission in 10.7 percent for a median length of 5.83 years, partial cytogenetic remission in 3.57 percent for a median length of 2 years. 55.5 percent were treated with imatinib (Gleevec) resulting in complete cytogenetic remission in 54.1 percent for a median length of 3.08 years, partial cytogenetic remission in 13.5% for a median length of 1.08 years.Results: Survival at five years through Kaplan-Meier analysis was approximately 92 percent in patients demonstrating only the Ph chromosome (n = 53), versus 60 percent in patients with additional chromosomal abnormalities (n = 12; p < 0.0001 by both Logrank and Breslow-Gehan-Wilcox analysis). Furthermore, patients with additional chromosomal abnormalities demonstrated lower rates of either complete or partial cytogenetic remission with imatinib therapy. Similarly, five year survival of patients with less than 90% Ph-positive metaphases at diagnosis (n = 4) was approximately 60% compared to 92% in patients with greater than 90% Ph-positive metaphases (n = 61; p = 0.01 by Logrank and p = 0.001 by Breslow-Gehan-Wilcox analysis). Additionally, the former group of patients was significantly less responsive to imatinib. Finally, there was significant correlation between Ph-positive metaphase mosaicism and the presence of additional chromosomal abnormalities (p = 0.05).Conclusion: Although it is tempting to speculate the possibility that Ph-chromosome mosaicism and/or additional cytogenetic abnormalities at presentation of CML is a surrogate for the presence of Ph-negative imatinib-resistant clones, the preliminary results from the current study require validation from a larger study. [Display omitted] [Display omitted]

Ph'-Positive (Ph'+) Chronic Myeloid Leukemia (CML) Presenting as Ph'+ T-Lymphoblastic Lymphoma (LL) Resistant to High-Dose Chemotherapy and Gleevec.

BACKGROUNDExtramedullary blast crisis (BC) of Ph'+ CML is infrequent and commonly affects bone, lymphoid tissue, skin and soft tissues and central or peripheral nervous system. Most of nodal lymphoid tumors occuring in the setting of CML derive from T-cell precursors and represent the evolution of CML to a lymphoid nodal BC.CASE HISTORYIn February 2004, a 52 year-old patient underwent diagnostic wide biopsy of a nasopharingeal mass that caused severe acute respiratory symptoms. At the histological examination, the nasopharyngeal mucosa exhibited a diffuse pattern of infiltration by neoplastic cells with a characteristic single-file arrangement. The cells showed typical convoluted nuclei with one or two nucleoli and abundant cytoplasm (L2, lymphoblasts). The malignant cell population expressed a preT-cell immunophenotype: cytoplasmic CD3(+), CD43 (+), TdT (+/−), CD34(+), CD4(−), CD8(−). Thus, the diagnosis of T-cell LL was formulated. Whole-body CT scan revealed nasopharingeal mass, retropharyngeal, laterocervical, axillary, inguinal enlarged nodes and splenomegaly. Laboratory tests indicated leukocytosis (58,000/μl) with a differential WBC count typical of CML in chronic phase (myeloblasts <1%). This diagnosis was confirmed by a low alkaline phosphatase value (score: 1), histological features of bone marrow (BM) biopsy, classical cytogenetics (presence of the Ph' in 100% of 25 metaphases analysed and absence of other cytogenetic abnormalities) and FISH evaluation (D-FISH bcr-abl in interphase and metaphase, Oncor probe). The nested RT-PCR (JQ Guo et al., Leukemia ; 2002, 15:2447) disclosed the presence of the hybrid protein p210 (b2,a2), but not that of p190. A laterocervical node was excised to perform cytogenetic and molecular analyses in order to determine whether the T-cell LL was an unrelated disease or the expression of an extramedullary BC. The histology confirmed the presence of a uniform population of T-lymphoblasts in which classical cytogenetic analysis disclosed the following kariotype: 49,XY,t(9;22)(q34;q11),+9,+19,+der(22)t(9;22)(q34;q11) and 50–52,XY t(9;22)(q34;q11),+6+9,+9q+,+19,+20,+22q−. The FISH analysis showed the presence of bcr-abl gene in all cells analyzed and the presence of multiple copies of this gene as well as of double Ph' chromosome. Nested RT-PCR showed the presence of both p210 and p190 transcripts. These findings indicated that the T-cell LL was an extramedullary BC of a CML simultaneously diagnosed in chronic phase in BM and peripheral blood. The patient was initially treated with vincristine, daunomicine, asparaginase and prednisone combined with Gleevec (800mg/die). Consolidation therapy, consisting of high-dose Ara-C (4g/m2 for 4 days) and mithoxantrone (10mg/m2 for 2 days), followed by a mieloablative course with mithoxantrone (60 mg/m2) and melphalan (180 mg/m2) and autologous stem cell support (ASCT) was administered after an initial clinical response. Gleevec was given during the entire treatment period. A fugacious complete clinical-hematological and a partial cytogenetic (FISH: 12% of cells bcr-abl+) and molecular (number of bcr-abl/104 ABL copies= 184, real-time quantitative RT-PCR: J Gabert et al., Leukemia ; 2003:1) remission was documented after ASCT. The patient refused further consolidation treatment with Gleevec and died, due to progressive disease, in December 2004.

Immunological Effects of Interferon-alpha on Chronic Myelogenous Leukemia

Treatment with interferon-α is effective for chronic myelogenous leukemia in the chronic phase (CML-CP), but the immunological mechanisms of the antileukemic effect of this substance are still unclear. The objective of this study was to investigate the immunological effects of interferon-α in CML patients. Markers of cellular activation and apoptosis, natural killer (NK) cell cytotoxicity and production of intracellular cytokines (IFN-γ, IL-2 and IL-4) were determined by flow cytometry in the peripheral blood mononuclear cells (PBMC) of 26 CML-CP patients before and 3, 6 and 9 months after IFN-α treatment. The results were correlated with the hematological response. In the whole group of patients, IFN-α use was followed by a significant increase of lymphocytes producing IL-2 and IFN-γ, an increase in NK activity and a decrease in the number of CD34+ cells. Out of 26 CML patients, 15 achieved hematological remission and 7 achieved partial cytogenetic remission after 9 months of IFN-α treatment. There was an increase in the percentage of CD8/FasL+, DR/CD3+, DQ/CD3+, CD34/Fas+, DR/CD56+, CD56/FasL+ cells and of IFN-γ- and IL-2-producing lymphocytes and an increase in NK cytotoxicity only in the group of patients who achieved complete hematological remission. Our results indicate that IFN-α use in CML-CP reduces the number of CD34+ cells, activates T cells, enhances stem cell apoptotic markers and increases the production of intracellular IFN-γ and IL-2 by lymphocytes. Taken together, these results indicate that the therapeutic effect of IFN-α in CML-CP is mediated at least in part by immunological mechanisms.

Combination therapy with interferon-alpha-2b and hydroxyurea in patients with chronic myelogenous leukemia

Background: Interferon-α-2b (INF-α) has been able to achieve hematological and cytogenetic remission in chronic myelogenous leukemia (CML) patients and has a beneficial effect in patients who have relapsed after bone marrow transplantation (BMT). Hydroxyurea (HU) is the optimal conventional drug for treatment of CML in the chronic phase. IFN-α-based combinations have attempted to improve the results of single agent trials. Methods: Thirty previously untreated patients (16 males and 14 females, 26–70 years old), newly diagnosed as having CML (Philadelphia chromosome-positive) from a single hematology center, were treated with a combination of IFN-α and HU. The initial dose of IFN-α was 3×10 6 daily (not corrected for body surface). HU was administered at a dose of 40 mg/kg daily. The maintenance dosage was adjusted to the white blood cell (WBC) count. Results: The median observation time was 5.2 years. Twenty-seven patients maintained the prescribed dose of IFN-α, while three patients received IFN-α at a dose of 5×10 6 due to persistent elevated WBC count. Complete hematological remission was achieved in 20 patients (66.6%) and partial remission in the other 10 patients (33.3%). Two patients achieved complete cytogenetic remission and three patients partial cytogenetic remission. During the study period, 20 patients died (median survival 5.1 years). The youngest patient (26 years old) died during BMT. Ten patients are still alive (median survival 5.37 years). Side effects from IFN-α included fever (100%) and flu-like syndrome (80%); two patients had tumor cell lysis syndrome. Conclusions: Combination INF-α and HU therapy is well tolerated and effective for the treatment of CML patients, resulting in a rapid control of the disease with only minor side effects.