Dose and drug changes in chronic lymphocytic leukemia cell response in vitro: A comparison of standard therapy regimens with two novel cyclin­dependent kinase inhibitors.

Chronic lymphocytic leukemia (CLL) treatment is improving; however, some patients do not respond to therapy. Due to the high heterogeneity in disease development, there is an urgent need for personalization of therapy. In the present study, the response of leukemic mononuclear cells to anticancer drugs used for CLL treatment (cladribine + mafosfamide; CM or CM combined with rituximab; RCM) was compared with the response to new cyclin­dependent kinase (CDK) inhibitors: BP14 and BP30. Viable apoptotic and necrotic cells were quantified by flow cytometry using propidium iodide and Yo­Pro stains. CDK inhibitors were studied in several doses to determine the reduction of necrosis and simultaneous increase of apoptosis in leukemic cell incubations with anticancer agents. The distinct cell response to applied doses/anticancer agents was observed. Results obtained in the current manuscript confirmed that modulation of doses is important. This was particularly indicated in results obtained at 24 h of cells incubation with anticancer agent. While an important time for analysis of anticancer response efficacy (monitoring of apoptosis induction potential) seems to be 48 h of cells exposition to anticancer agents. High variability in response to the drugs revealed that both the nature and the dose of the anticancer agents could be important in the final effect of the therapy. The present findings support the thesis that personalized medicine, before drug administration in the clinic, could be important to avoid the application of ineffective therapy.

Bendamustine alone or with rituximab modifies expression of apoptosis-regulating genes and proteins of CLL cells, depending on IGVH mutational status.

We studied whether bendamustine (BENDA) alone or with rituximab (RIT) modifies in vitro expression of apoptosis-involved genes and proteins of chronic lymphocytic leukemia (CLL) cells depending on IGVH mutational status. Circulating lymphocytes from 34 untreated patients (18 IGVH-MUT and 16 IGVH-UNMUT) were incubated with above drugs to evaluate proteins expression. Microarray analysis of 93 genes was performed in 14 patients. BENDA and BENDA + RIT increased expression of BAX and BBC3 in IGVH-MUT and IGVH-UNMUT groups, and significant differences in expression of above genes after BENDA + RIT were observed between both groups. Additionally, BENDA + RIT decreased NFκB and BCL-2 genes in IGVH-UNMUT patients and increased expression of P53, BAX and PUMA proteins in IGVH-MUT and UNMUT subjects. However, no significant differences were found between these groups. In conclusion, BENDA + RIT modified gene expression profile in CLL cells and affected expression of some apoptosis-regulating proteins in vitro. Expression of BAX and BBC3 depends on action of drugs and IGVH mutational status.

Personalized therapy tests for the monitoring of chronic lymphocytic leukemia development.

There is individual variation in the course of disease development and response to therapy of patients with chronic lymphocytic leukemia (CLL). Novel treatment options for CLL include a new generation of purine analogs, antibodies and inhibitors of specific cell signaling pathways, which typically induce apoptosis or necrosis. A prospective analysis of patient blood samples revealed that a combination of four tests allowed the most appropriate and effective type of treatment to be selected prior to drug administration, and for the analysis of leukemic cell sensitivity to anticancer drug(s) during disease development. The comparative analysis of blood from the stable and progressive form of CLL in an individual patient revealed diversity in the response to anticancer agents. CLL peripheral blood mononuclear cells were incubated with cladribine + mafosfamide (CM), fludarabine + mafosfamide, CM + rituximab, rituximab alone (Rit) or kinetin riboside (RK). A combination of cell viability, differential scanning calorimetry (DSC) profiles of nuclear preparations and poly(ADP-ribose) polymerase 1 (PARP-1) protein expression analysis of the leukemic cells was performed to evaluate the anticancer effects of the tested agents during CLL development. The results of the present study indicate that such studies are effective in determining the most appropriate anticancer drug and could monitor disease progression on an individual level. In addition, the results of the current study suggest that CLL progression leads to diversification of the cellular drug response. The most efficient apoptosis inducer for the patient was purine analog RK when the disease was stable, while the CM combination was the most effective agent for the progressive form of disease.

Health-related quality of life and patient-reported outcomes of ofatumumab plus fludarabine and cyclophosphamide versus fludarabine and cyclophosphamide in the COMPLEMENT 2 trial of patients with relapsed CLL.

Chronic lymphocytic leukemia (CLL) is an incurable disease. Quality of life during treatment and periods of subsequent remission is therefore vital. Health-related quality of life (HRQoL) was compared in relapsed CLL during and after treatment with ofatumumab combined with fludarabine and cyclophosphamide versus fludarabine and cyclophosphamide alone. The European Organisation for Research and Treatment of Cancer (EORTC) QLQ-C30 v3 and QLQ-CLL16 were used to assess HRQoL in this open-label, phase 3 study. Improvements in prespecified domains of patient-reported outcomes (Global Health Status [GHS]/HRQoL and B symptom scores) were recorded in both treatment arms after three cycles and were sustained after 18 months of follow-up. The two treatment arms were not significantly different at the nominal 0.05 level for GHS/HRQoL (p = .7278) or B symptoms (p = .5968). Small improvements in quality of life were maintained after therapy. The addition of ofatumumab was without any adverse impact on HRQoL (NCT00824265).

Ofatumumab plus fludarabine and cyclophosphamide in relapsed chronic lymphocytic leukemia: results from the COMPLEMENT 2 trial.

In this multicenter, open-label, phase III study, patients with relapsed chronic lymphocytic leukemia (CLL) were randomized (1:1) to receive ofatumumab plus fludarabine and cyclophosphamide (OFA + FC) or FC alone; the primary endpoint being progression-free survival (PFS) assessed by an independent review committee (IRC). Between March 2009 and January 2012, 365 patients were randomized (OFA + FC: n = 183; FC: n = 182). Median IRC-assessed PFS was 28.9 months with OFA + FC versus 18.8 months with FC (hazard ratio = 0.67; 95% confidence interval, 0.51-0.88; p = .0032). Grade ≥3 adverse events (≤60 days after last dose) were reported in 134 (74%) OFA + FC-treated patients compared with 123 (69%) FC-treated patients. Of these, neutropenia was the most common (89 [49%] vs. 64 [36%]). OFA + FC improved PFS with manageable safety for patients with relapsed CLL compared with FC alone, thus providing an alternative treatment option for patients with relapsed CLL. TRIAL REGISTRATION: www.clinicaltrials.gov (NCT00824265).

In vitro antileukemic activity of novel adenosine derivatives bearing boron cluster modification.

A series of adenosine derivatives bearing a boron cluster were synthesized and evaluated for their cytotoxicity against primary peripheral mononuclear cells from the blood of 17 patients with leukemias (16 CLL and 1 very rare PLL), as well as from 5 healthy donors used as a control. Among the tested agents, two, i.e., compounds 1 and 2, displayed high in vitro cytotoxicity and proapoptotic potential on leukemic cells, with only scarce activity being seen against control cells. Biological tests related to apoptosis revealed the activation of the main execution apoptotic enzyme, procaspase-3, in CLL and PLL cells exposed to compounds 1 and 2. Moreover, the above compounds indicated high activity in the proteolysis of the apoptotic markers PARP-1 and lamin B1, fragmentation of DNA, and the induction of some changes in the expression of the Mcl-1, protein apoptosis regulator in comparison with control cells.

Antibody therapy alone and in combination with targeted drugs in chronic lymphocytic leukemia.

The development of non-chemotherapeutic agents, including monoclonal antibodies (mAbs) and other targeted drugs, makes chemotherapy-free treatment an attractive option for chronic lymphocytic leukemia (CLL). The classical mAb, rituximab, has been authorized for use in both first-line and second-line therapy for CLL. New mAbs directed against CD20, ofatumumab, and obinutuzumab (GA-101) have also been approved for the treatment of this disease. Recently, several new mAbs with potential benefits over the approved anti-CD20 antibodies have been developed for use in CLL. Anti-CD37, anti-CD19, and anti-CD40 mAbs are in early clinical trials and show promise in treating CLL. In addition, the combination of mAbs with B-cell receptor signaling pathway inhibitors and immunomodulatory drugs makes the chemotherapy-free option a reality today. Combinations of antibodies with targeted drugs like ibrutinib, idelalisib, or lenalidomide are expected to replace chemotherapy-based combinations for treating CLL in the near future. However, phase III trials should confirm the benefit of these new treatment strategies and establish their exact place in the therapeutic armamentarium for CLL.

Prognostic value of thymidine kinase activity in patients with chronic lymphocytic leukemia.

Thymidine kinase (TK) activity is a marker of biological activity that allows the indolent and aggressive forms of chronic lymphocytic leukemia (CLL) to be distinguished. The aims of the study were to determine the relationship between TK activity and clinical status and prognosis, as well as to compare its activity with that of other prognostic and predictive factors. TK activity was measured in patient sera at the time of diagnosis using the DiviTum method, with the mean value being 439 Du/L. A correlation was found between TK activity and risk of disease progression (p=0.045). The optimal discriminative value of TK activity in the prediction of CLL progression was found to be 600 Du/L. TK activity significantly differed between the patients who achieved complete remission and those who only partially responded to therapy. In 93% of patients without any response to treatment and 18 out of 20 patients with progressive disease, TK activity over 600 Du/L was noted. In addition, all of the 10 patients with 17p13 deletion displayed TK activity of over 600 Du/L (p=0.0004). High TK activity also correlated with elevated levels of LDH (p=0.001) and ß2­microglobulin (p=0.03) in the study group. The results of the study indicated the importance of TK activity as a prognostic factor in patients with CLL.

Relationship between in vitro drug sensitivity and clinical response of patients to treatment in chronic lymphocytic leukemia.

To improve the efficacy of therapeutic options in chronic lymphocytic leukemia (CLL) an in vitro system to determine the response of mononuclear blood cells from blood of patients was elaborated. The study combines four approaches, i.e., cell viability, apoptosis rate, differential scanning calorimetry (DSC), and immunoblotting to develop personalized therapy protocols based on the cell sensitivity to drug exposure of individual CLL patients. The complementary analyses were performed on 28 peripheral blood samples from previously untreated CLL patients before therapy. The induction and progress of apoptosis in CLL cells exposed in vitro to purine analogs combined with mafosfamide, i.e., cladribine + mafosfamide (CM) and fludarabine + mafosfamide (FM) were assessed using the above approaches. The changes in thermal profiles (decrease/loss of transition at 95±5˚C) coincided with an accumulation of apoptotic cells, a decrease in the number of viable cells, and differences in the expression of the apoptosis­related protein PARP­1. No significant changes were observed in the thermal profiles of nuclei isolated from CLL cells resistant to the treatment. The complementary assays revealed a strong relationship between both the in vitro sensitivity of leukemia cells to drugs and the clinical response of the patients, determined usually after the sixth course of treatment (after ~6 months of therapy). As a summary of studies followed by complementary tests, our findings demonstrate the value of in vitro exposure of CLL cell samples to drugs intended to treat CLL patients, before their administration in order to recommend the most suitable and effective therapy for individual patients.

Cytotoxic and apoptosis-inducing effects of bendamustine used alone and in combination with rituximab on chronic lymphocytic leukemia cells in vitro.

AIM: The aim of our study was to compare the cytotoxic effects of bendamustine (BENDA) and rituximab (RIT) used either alone or in combination and to evaluate the influence of the above mentioned drugs on apoptosis measured as changes in mitochondrial transmembrane potential (Δψm), expression of caspases and selected apoptosis-regulating proteins in freshly isolated peripheral blood mononuclear cells of chronic lymphocytic leukemia (CLL) patients. MATERIALS/METHODS: Cytotoxic effect of tested drugs, as well as induction of apoptosis, drop in Δψm and expression of selected proteins involved in regulation of apoptosis were assessed in 48 hour cultures containing autologous serum (AS) using flow cytometry. BENDA was used at the concentration of 40 µg/ml and RIT at the concentration of 10 µg/ml. Control cultures were incubated without drugs. RESULTS: BENDA used either alone or in combination with RIT strongly induced apoptosis as well as enhanced expression of selected apoptotic proteins, especially those involved in the intrinsic apoptotic pathway: P53, PUMA and BAX, which cause mitochondrial transmembrane potential changes leading to activation of caspase-9 and -3. CONCLUSIONS: Our results indicate that both BENDA and RIT participate in the induction of apoptosis of CLL lymphocytes in vitro in the presence of AS in the culture medium. The drug-induced apoptosis occurs mainly via intrinsic pathway and activation of P53 and PUMA proteins, however the extrinsic pathway is likely to be involved as well. We also found that the combination of these drugs induces the expression of P53, caspase-8 and -9 more potently than either of them used separately.

Long-term results of the Polish Adult Leukemia Group PALG-CLL2 phase III randomized study comparing cladribine-based combinations in chronic lymphocytic leukemia.

Long-term outcomes following newer therapies for chronic lymphocytic leukemia (CLL) have rarely been reported. This article presents the results of the final analysis of the Polish Adult Leukemia Group PALG-CLL2 study performed 10 years from final patient enrollment. With the extended follow-up time, it was found that cladribine (2-CdA)-based combinations CMC (2-CdA, cyclophosphamide, mitoxantrone) and CC (2-CdA, cyclophosphamide) administered as first-line treatment of progressive CLL resulted in significantly longer progression-free survival, but similar overall survival compared to 2-CdA monotherapy. Furthermore, the risk of potentially fatal late adverse events including infections, autoimmune complications and, particularly, secondary neoplasms was comparable among patients treated with CMC, CC or 2-CdA. The results of our analysis support the importance of long-term outcome monitoring of randomized trials in CLL.

Clonal evolution in CLL patients as detected by FISH versus chromosome banding analysis, and its clinical significance.

The acquisition of new aberrations during the course of chronic lymphocytic leukemia (CLL) named clonal evolution (CE) is usually detected by one of the two methods: chromosome banding analysis (CBA) and interphase fluorescence in situ hybridization (I-FISH). The purpose of this study was to compare the usefulness of FISH and CBA for detecting CE and to evaluate its influence on clinical outcome. FISH and CBA were performed at two time points: baseline and follow-up. Thirty-eight previously untreated patients with CLL were included in this study. CBA and I-FISH revealed CE in 15 (39.5%) and 10 (26.3%) patients, respectively. High-risk CE was detected in six cases by CBA and in five cases by I-FISH. In four cases with CE-dependent 17p abnormalities detected by CBA, metaphase FISH was needed for the confirmation of 17p13.1 deletion. Time from first-line to second-line treatment (TTST) and overall survival (OS) did not differ between patients with and without CE, irrespective of the CE-detecting method used. However, shorter OS (P = 0.043) and TTST (P = 0.006) were observed for the patients with potentially relevant CE (rCE) detected by CBA, in which acquired aberrations were present in at least 20% of undivided cells and/or changed baseline karyotype to abnormal or complex and were not resulting from 13q deletion. Our results suggest that some, but not all, CE-dependent aberrations detected by CBA influence clinical outcome. Moreover, I-FISH, which was aimed at detecting aberrations of prognostic significance, was found to be more precise than CBA in their detection, especially TP53 deletion.

In vivo and ex vivo responses of CLL cells to purine analogs combined with alkylating agent.

BACKGROUND: The heterogeneity of chronic lymphocytic leukemia (CLL) is thought to be due to differences in the expression of factors that regulate apoptosis and cell cycle, giving rise to diverse apoptotic disturbances and tumor properties. Therefore, the primary goal in CLL treatment is to overcome resistance to apoptosis and efficiently trigger this process in leukemic cells. METHODS: Mononuclear cells were obtained from the blood of CLL patients by Histopaque-1077 sedimentation. CLL cell samples from the blood of drug treated patients, (cladribine or fludarabine with cyclophosphamide; CC or FC), as well as the cell samples of untreated patients exposed to the used drug combinations (CM, FM) or mafosfamide alone for 48 h were fractionated into nuclear and cytoplasmic fractions or were lysed. DNA fragmentation was evaluated by agarose electrophoresis and also cytometrically as sub-G1 population. The expression of apoptosis related proteins and H1.2 histone translocation were evaluated in lysates and nuclear and cytoplasmic fractions, respectively with appropriate antibodies. RESULTS: Cladribine (C) and fludarabine (F) combined with cyclophosphamide/mafosfamide in vivo, as well as ex vivo trigger apoptosis in CLL cells. These drug combinations (CC; FC/CM; FM) induce leukemic cell apoptosis confirmed by DNA fragmentation, sub-G1 cell number, down-regulation of anti-apoptotic proteins (Mcl-1, Bcl-2), and H1.2 histone translocation in comparison with appropriate control cells, however, to a different degree. CONCLUSIONS: The kinetics and rate of drug-induced apoptosis in leukemic cells under ex vivo experiments differ between patients, mirroring the differences noticed during in vivo treatment. Individual model cell samples indicate comparable susceptibility to the used drug combinations under in vivo and ex vivo conditions.

Immune thrombocytopenia in patients with chronic lymphocytic leukemia treated with cladribine-based regiments or chlorambucil--follow-up of PALG-CLL randomized trials.

OBJECTIVES: The relationship between treatments of chronic lymphocytic leukemia (CLL) with cladribine (2-CdA) or chlorambucil and immune thrombocytopenia (IT) has not been yet determined. METHODS: The records of 777 patients in two randomized Polish Adult Leukemia Group (PALG)-CLL programs treated with these agents were retrospectively analyzed. RESULTS: Immune thrombocytopenia occurred in 55 of 777 (7.1%) patients. No significant differences in IT prevalence were seen between patients on chlorambucil or 2-CdA-based regiments (P = 0.33). IT developed at a median time of 0.499 yr (0.06-4.8) from the start of CLL therapy. This time was significantly longer in patients treated with chlorambucil (2.03 yr, 95% CI: 0.06-4.22) in relation to patients treated with 2-CdA-based regiments (0.52 yr, 95%CI: 0.34-0.69, P = 0.049). Overall survival (OS) of patients with IT and those without IT were similar (2.65 yr vs. 3.2 yr P = 0.23) but the severity of bleeding was more pronounced in the 2-CdA group. The responses to IT therapy were 35%, 54% and 75% for steroids, chemotherapy and splenectomy, respectively. CONCLUSIONS: In this study, an unexpectedly high percentage of IT incidence was demonstrated in patients with CLL requiring chemotherapy. Although no marked differences were seen in IT frequency in patients treated with 2-CdA-based regiments compared to chlorambucil regimen, the clinical course of hemorrhagic diathesis was more severe in 2-CdA group. Also, the time elapsed from study screening to IT diagnosis was significantly shorter in the 2-CdA group than in the chlorambucil group suggesting a causative relationship. The appearance of IT did not influence the median time of OS.

Promising anti-leukemic activity of atorvastatin.

There is a current need for novel therapeutic strategies for the treatment of chronic lymphocytic leukemia (CLL), a still incurable hematological cancer involving mainly deregulated apoptosis. The purpose of the present study was to determine ex vivo the effect of the synthetic statin, atorvastatin, a known cholesterol-lowering drug, on peripheral blood mononuclear cells obtained from CLL patients. Using flow cytometry, we investigated the viability and induction of apoptosis in leukemic cells exposed to statin by the Vybrant apoptosis assay kit #4, compared with untreated control cells. We also examined the expression levels of apoptosis-regulatory proteins (Mcl-1, Bcl-2 and Bax), as well as products of the expression/proteolysis of lamin B, poly(ADP-ribose) polymerase­1 (PARP­1) and p27Kip1 by western blot analysis. Moreover, the number of sub-G1 cells and DNA fragmentation in atorvastatin-treated leukemic cells were examined by flow cytometry and agarose gel electrophoresis, respectively. The obtained results indicated that CLL cells ex vivo were extremely sensitive to atorvastatin. The cytotoxic effect of this statin was caused by the induction of apoptosis in the leukemic cells. The induction of apoptosis in the drug-treated model cells was confirmed by the reduction or proteolysis of apoptotic markers, such as PARP-1, lamin B and p27Kip1, the increase in the number of sub-G1 cells and DNA ladder formation. During atorvastatin-triggered apoptosis, changes in the expression levels of mitochondrial outer membrane permeability regulatory proteins of the Bcl-2 family were also observed. Ex vivo promising data indicate the strong cytotoxic and pro-apoptotic potential of atorvastatin against leukemic cells, but not normal cells. The obtained data suggest that atorvastatin be considered as a therapeutic option for the treatment of CLL.

Toward personalized therapy for chronic lymphocytic leukemia: DSC and cDNA microarray assessment of two cases.

The differences in clinical course of chronic lymphocytic leukemia could have an impact on variations in a patient's response to therapy. Our published results revealed that thermal transition (95 ± 5°C) in differential scanning calorimetry profiles appear to be characteristic for the advanced stage of CLL. Moreover, a decrease/loss of this transition in nuclei from leukemic cells exposed to drugs ex vivo could indicate their diverse efficacy. It seems that the lack of changes in thermal profile could predict patient's drug resistance. In this study, we demonstrate the results obtained after drug treatment of leukemic cells by calorimetry, apoptosis-related parameters involved in expression of genes using cDNA microarray and western blot. These data were compared with the patients' clinical parameters before and after RCC therapy (rituximab + cladribine + cyclophosphamide). The complementary analysis of studied cases with opposite clinical response (CR or NR) revealed a strong relationship between clinical data, differences in thermal scans and apoptosis-related gene expression. We quantified expression of eight of apoptosis-related 89 genes, i.e., NOXA, PUMA, APAF1, ESRRBL1, CASP3, BCL2, BCL2A1 and MCL1. Particular differences in NOXA and BCL2 expression were revealed. NOXA expression in cells of patients who achieved a complete response to RCC therapy was 0.44 times higher in comparison to control ones. Interestingly, in the case of patients who did not respond to immunotherapy, NOXA expression was highly downregulated (RQ = 4.39) as compared with untreated cells. These results were confirmed by distinct cell viability, protein expression as well as by differences in calorimetry profiles.

Apoptotic gene expression under influence of fludarabine and cladribine in chronic lymphocytic leukemia-microarray study.

BACKGROUND: A deep insight into gene expression profiling (GEP) is a key to understanding the background of disease. It can lead to identification of diagnostic and prognostic factors and then to a selection of the most appropriate therapy. The aim of this study was to evaluate differences in apoptotic gene expression in chronic lymphocytic leukemia (CLL) cells influenced by fludarabine (FA) or cladribine (2-CdA). METHODS: GEP was performed in cells obtained from 10 untreated CLL patients and cultured in vitro with FA or 2-CdA. Ninety-three selected apoptotic genes were analyzed using 384 TaqMan® Low Density Arrays in pooled RNA. RESULTS: Relevant results were found in a set of 27 genes, however, the most striking differences between FA and 2-CdA were observed in the following 5 genes: BAD, TNFRSF21, DAPK1, CARD 6 and CARD 9. CONCLUSION: We have found some differences in apoptotic gene expression between FAand 2-CdA. These findings give prominence to genes qualifying for further studies currently conducted in our Department.

Changes in the apoptotic gene expression profile in CLL patients treated with rituximab combined with cladribine and cyclophosphamide-preliminary results.

The study was aimed to investigate modifications of apoptotic gene expression profile by microarray technique in 10 patients with chronic lymphocytic leukemia by treatment with rituximab, cladribine and cyclophosphamide (RCC) according to IGHV mutational status. The TaqMan Low Density Array for 96 gene transcripts was used. Those modifications followed two distinctive patterns largely overlapping the IGHV mutational status. In the IGHV-mutated group, the expression of many proapoptotic genes increased after treatment as compared to initial value. Our results suggest that RCC drugs may act through influence on the expression of some apoptosis-involved genes dependently on the IGVH mutational status.

Can ex vivo evaluation (testing) predict the sensitivity of CLL cells to therapy with purine analogs in conjunction with an alkylating agent? A comparison of in vivo and ex vivo responses to treatment.

Malfunctions in the regulation of apoptosis cause the accumulation of malignant, long-lived B CD19+/CD5+ cells in chronic lymphocytic leukemia (CLL). The primary goal in CLL therapy is to overcome resistance to apoptosis and efficiently trigger programmed cell death in leukemic cells. This study demonstrated that the in vivo responses of malignant cells from CLL patients after administration of purine analogs (cladribine/fludarabine) with cyclophosphamide vary significantly. For comparative purposes, the sensitivity of leukemic cells obtained from the same CLL patients to conventional purine analogs and the selective CDK inhibitor R-roscovitine (ROSC) was determined, with and without the addition of an alkylating agent, prior to the onset of in vivo therapy. The kinetics and rate of spontaneous and drug-induced apoptosis of CLL cells under ex vivo conditions differed significantly between patients, mirroring the variability observed during in vivo treatment. Interestingly, individual patients' leukemic cells were comparably sensitive to the drugs under both conditions. Of the drugs examined, ROSC exerted the highest therapeutic efficacy under ex vivo conditions. Our results indicate that ex vivo testing might be useful for identifying the most potent first-line therapeutic regimen for specific CLL patients and possibly for the design of therapies tailored for individual CLL patients.

Chromosomal aberrations in chronic lymphocytic leukemia detected by conventional cytogenetics with DSP30 as a single agent: comparison with FISH.

The aim of our study was to estimate the usefulness for conventional cytogenetics (CC) of DSP30 as a single agent (CC-DSP30) for detecting the most important chromosomal aberrations revealed in CLL by FISH and to find other abnormalities possibly existing but undetected by FISH with standard probes. Using CC-DSP30, the metaphases suitable for analysis were obtained in 90% of patients. CC-DSP30 and FISH were similarly efficacious for detecting del(11)(q22) and trisomy 12, whereas FISH was more sensitive for del(13)(q14). Sole del(13)(q14) detected by FISH, in 50% of patients was associated with other aberrations revealed by CC-DSP30. Additionally, the most recurrent anomaly detected by CC-DSP30 were structural aberrations of chromosome 2.