The Parthenolide Derivative LC-1 Is An Effective Single Agent and Is Highly Synergistic with Existing Therapies in Multiple Myeloma.

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 1708-1708
Author(s):  
Elisabeth J Walsby ◽  
Saman Hewamana ◽  
Alan Burnett ◽  
Steven Knapper ◽  
Chris Fegan ◽  
...  
Keyword(s):  
Multiple Myeloma ◽  
Bone Marrow ◽  
Cell Lines ◽  
Nuclear Localization ◽  
Plasma Cells ◽  
Bone Marrow Cells ◽  
Binding Capacity ◽  
Single Agent ◽  
Normal Bone Marrow ◽  
Normal Bone

Abstract Multiple myeloma (MM) remains incurable with conventional therapeutic agents and has a median survival of only 3–5 years. Therefore, there is clearly a need for novel treatment strategies that can change the natural pathology of this condition. The nuclear factor κB (NF-κB) family of transcription factors is constitutively activated in MM cell lines and the majority of MM patients. Since NF-κB has known oncogenic activity in a number of human malignancies, targeted inhibition of this family of proteins may be useful in the treatment of MM. We and others have recently shown that the parthenolide derivative LC-1 has activity in acute myeloid leukaemia (AML) and chronic lymphocytic leukaemia (CLL) cells. Unusually, it induces apoptosis via the activation of both the intrinsic and extrinsic pathways and apoptosis is preceded by marked inhibition of NF-κB. Importantly, LC-1 is more potent against primary AML blasts and CLL lymphocytes than normal bone marrow progenitors and normal B-cells and T-cells. In this study we set out to evaluate LC-1 in MM cell lines and plasma cells derived from MM patients. LC-1 was cytotoxic to MM cell lines H929, U266 and JJN3 and induced apoptosis in a dosedependent manner resulting in an overall LD50 of 3.6mM (±1.8) after 48 hours in culture. Primary myeloma plasma cells, identified by CD38 and CD138 positivity, had a mean LD50 for LC-1 of 5.4mM (±1.6) after 48 hours of in vitro culture. Normal bone marrow cells were significantly less sensitive to the effects of LC-1 under the same conditions (P = 0.0007). Treatment of MM cell lines with LC-1 resulted in a decrease in the nuclear localization of NF-κB, as evidenced by a dose-dependent decrease in the DNA binding capacity of the NF-κB subunit RelA after 4 hours of treatment. To demonstrate whether synergy exists between LC-1 and existing MM therapies, the H929 cell line was treated for 48 hours with LC-1 and doxorubicin (32:1), melphalan (1:1) or bortezimib (1:500) and the combination indices (CI) calculated using the median effect method. A combination index of less than 1 denotes synergy. LC-1 did not show synergy with doxorubicin (CI >1) but was synergistic with melphalan and bortezimib (CI values of 0.53 and 0.59 respectively). Taken together our data clearly demonstrate that LC-1 has activity in MM cell lines and primary MM cells. Its ability to inhibit the nuclear localization of NF-κB is important to its cytotoxic effects. Furthermore, it may also provide an explanation for the synergy demonstrated with melphalan and bortezimib. These results provide a rationale for exploring the potential of LC-1 in clinical studies.

Over Expression of YY1 and RKIP in Multiple Myeloma (Cell Lines and MM Tissues).

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 5035-5035
Author(s):  
Stavroula Baritaki ◽  
Sara Huerta-Yepez ◽  
Alina Katsman ◽  
Kam C. Yeung ◽  
Devasis Chatterjee ◽  
...  
Keyword(s):  
Multiple Myeloma ◽  
Bone Marrow ◽  
Cell Lines ◽  
Bone Marrow Cells ◽  
Myeloma Cell ◽  
Gene Products ◽  
Normal Bone Marrow ◽  
Over Expression ◽  
Normal Bone ◽  
Marrow Cells

Abstract There is a need to identify underlying molecular mechanisms and gene products (targets for intervention and biomarkers) involved in MM chemo- or immuno-resistance. Recent findings from our laboratory have reported high expression of the transcription factor Yin Yang 1 (YY1) in several tumor types and have demonstrated its fundamental role in tumor cell chemo/immuno-resistance (Gordon, S., et al., Oncogene25:1125, 2006). In contrast, low levels of the Raf kinase inhibitor protein (RKIP) has been shown to play a pivotal role in the negative regulation of cell survival signaling pathways, and has been considered a metastasis tumor suppressor. The overall objective of our studies is to examine whether MM cell lines and tissues derived from MM patients present deregulated expression patterns of RKIP and YY1, and to demonstrate the roles of YY1 and RKIP in MM pathogenesis and response to various therapies. In the present study, the myeloma cell lines RPMI-8226 and MM-1S were screened for RKIP and YY1 expression at the protein and m-RNA levels. In addition, fresh tissues from MM patients were also examined for RKIP and YY1 expression by IHC and their patterns compared with those observed in normal bone marrow cells. The findings demonstrate that both MM cell lines express remarkably higher levels of RKIP protein and transcripts compared to other tumor cell lines examined (lymphomas and prostate cancer), as assessed by western, RT-PCR and IHC. There was significantly higher expression of RKIP in MM patient’s samples compared to normal bone marrow cells. YY1 protein and m-RNA levels were detected in the MM cell lines; however, they were lower compared to Ramos and PC-3 cells. YY1 expression in MM tissues was significantly elevated compared to normal bone marrow cells. RKIP was basically present in the cytoplasm while YY1 was mainly accumulated in the nucleus. In contrast, normal bone marrow cells presented primarily cytoplasmic YY1 and RKIP expression. The YY1 expressing population was the CD38+/CD138− cell subset, which has been reported to be more susceptible to apoptosis (Mitsiadis, S.C., et al., Blood 98:795, 2001). Overall, the present findings support the potential involvement of two new gene products, such as YY1 and RKIP, in MM pathogenesis and their implication in regulating MM resistance to conventional therapeutics. Moreover, these gene products suggest their potential use as new therapeutic targets and/or biomarkers in multiple myeloma. Future studies with large cohorts of tissues will elucidate the importance of the above findings and will give new insights in the unexpected RKIP over expression and activity in multiple myeloma.

High Frequency of Molecular Remissions (m-CR) in Multiple Myeloma Patients Achieving a Hematologic Complete Remission (CR) on Total Therapy II (TT-2).

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 933-933
Author(s):  
Guido Tricot ◽  
Matteo Carrabba ◽  
Bart Barlogie ◽  
Joshua Epstein ◽  
John Shaughnessy ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Plasma Cells ◽  
Bone Marrow Cells ◽  
Residual Disease ◽  
Detection Threshold ◽  
M Protein ◽  
Pcr Analysis ◽  
Normal Bone Marrow ◽  
Normal Bone ◽  
Marrow Cells

Abstract TT-2 represents a very intensive treatment approach for newly diagnosed myeloma patients consisting of hematopoietic growth factor-dependent induction therapy (VAD, DCEP, CAD, DCEP); followed by melphalan 200 mg/m2-based tandem autotransplant; 4 quarterly consolidation chemotherapy cycles with D-PACE; and interferon alpha maintenance with added dexamethasone pulsing during the first year. A stringently defined hematologic CR (immunofixation negative, normal bone marrow aspirate and biopsy, and absence of a light-chain restricted aneuploid population of bone marrow plasma cells by DNA/cIg flow cytometry) was obtained in 48% of the first 446 patients enrolled on TT-2. A CR defined as such probably reflects a detection threshold of 108–109 tumor cells. CDR3-PCR to evaluate minimal residual disease (MRD) in myeloma has revealed m-CR in 50% of 40 reported allotransplant cases compared to approximately 15% among 75 reported autotransplants myeloma. We evaluated the frequency of m-CR in 20 TT-2 patients with a qualitative PCR for CDR3, which can detect 1 myeloma cell in 105–106 normal bone marrow cells (Corradini et al. J Clin Oncol 1999, 17: 208). CDR3 probes were generated from RNA of CD138 immunogenetic bead-purified plasma cells obtained at diagnosis. Light-density (<1.077 g/cm3) bone marrow cells served as the source of DNA post-therapy to assess MRD. The quality of DNA was determined by amplification of the p53 exon 6 sequence on all CDR3-PCR negative cases. At the time of CDR3-PCR analysis, 13 patients were in hematologic CR, 1 in near CR (immunofixation positivity as the only evidence of disease), 4 in partial remission (PR) (bone marrow < 5% plasma cells, decrease in serum M protein ≥ 75% and in urine M protein ≥ 90%) and two had less than a PR. Four patients were tested for MRD prior to the first transplant (2 < PR, 1 PR, 1 CR), two prior to the second transplant (2 PR) and 14 at a median of 21 months (range 6 months to 2 years) after the second transplant (12 CR, 1 near CR, 1 PR). Six of the 13 CR patients had abnormal metaphase cytogenetics at diagnosis, including 5 with deletion of chromosome 13. A m-CR was observed in 10/13 (77%) hematologic CR patients, including 4/6 with abnormal cytogenetics and 6/7 with normal cytogenetics. P53 exon 6 amplification was seen in all patients who were CDR3-PCR negative. Not unexpectedly, all seven patients failing to achieve a stringently defined hematologic CR had persistent disease as assessed by CDR3-PCR. Our results suggest that high intensity therapy as applied in TT-2 can indeed achieve a more marked tumor cytoreduction as reflected by a higher m-CR rate than previously reported with autotransplantation also in patients with baseline abnormal metaphase cytogenetics, and that the m-CR rate appears to be at least equivalent to that observed after allotransplantation. It remains to be shown whether achieving a m-CR will result in prolonged survival and a potential of cure in myeloma as has been shown convincingly for other hematologic malignancies. An additional 35 hematologic CR patients have samples available for CDR3-PCR analysis so that, at the time of presentation, a more definitive assessment of TT-2’s potential to effect a m-CR can be presented.

scholarly journals Effects of hoechst 33342 on survival and growth of two tumor cell lines and on hematopoietically normal bone marrow cells

Cytometry ◽  
1982 ◽  
Vol 3 (1) ◽  
pp. 42-47 ◽  
Author(s):  
Jerrold Fried ◽  
Jeffrey Doblin ◽  
Shigeru Takamoto ◽  
Amaury Perez ◽  
Herbert Hansen ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Tumor Cell ◽  
Cell Lines ◽  
Bone Marrow Cells ◽  
Normal Bone Marrow ◽  
Tumor Cell Lines ◽  
Hoechst 33342 ◽  
Normal Bone ◽  
Survival And Growth ◽  
Marrow Cells

A CD123-Targeting Antibody-Drug Conjugate (ADC), IMGN632, Designed to Eradicate Acute Myeloid Leukemia (AML) Cells While Sparing Normal Bone Marrow Cells

Blood ◽  
2016 ◽  
Vol 128 (22) ◽  
pp. 768-768 ◽  
Author(s):  
Yelena Kovtun ◽  
Gregory Jones ◽  
Charlene Audette ◽  
Lauren Harvey ◽  
Baudouin Gerard ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Flow Cytometry ◽  
Cell Lines ◽  
Bone Marrow Cells ◽  
Cell Populations ◽  
Normal Bone Marrow ◽  
Normal Bone ◽  
Healthy Donors ◽  
Marrow Cells ◽  
Myeloid Progenitors

Abstract Current AML therapies are effective in a subset of patients, but often lead to prolonged myelosuppression. CD123 is an attractive AML target due to its elevated expression on AML compared to normal bone marrow cells. Still, severe myelosuppression and myeloablation have been reported in preclinical studies for some CD123-targeted therapies. Here, we present a novel ADC which selectively kills CD123-positive AML cells over normal bone marrow cells. A novel humanized anti-CD123 antibody with two engineered cysteines for payload conjugation was generated. Indolinobenzodiazepine dimers, termed IGNs, were chosen as payload molecules for the antibody due to their high potency against AML cells. The IGN dimers containing mono-imines alkylate DNA, whereas the di-imine containing IGNs can both alkylate and crosslink DNA. To select an optimal IGN payload, we compared the cytotoxicity of an ADC with a mono-imine IGN (A-ADC) to one with a di-imine IGN (C-ADC) on AML cells, as well as normal bone marrow cells in vitro. Potency of the ADCs was evaluated using AML cell lines that have CD123 levels similar to patient cells and carry markers of poor prognosis (FLT3-ITD , MDR1, EVI1, DNMT3A and TP53), as well as on samples from 11 AML patients. AML cells exposed to either ADC displayed markers of DNA damage, cell cycle arrest and apoptotic cell death by flow cytometry. Both ADCs were highly cytotoxic, generating IC50 values between 0.4 to 60 pM on the cell lines in WST-8 assays and killing 90 percent of progenitors from AML patients between 2 to 46 pM in CFU assays. The C-ADC was, on average, two-fold more active than the A-ADC. The cytotoxicity of both ADCs was CD123 dependent, since masking CD123 with a competing anti-CD123 antibody reduced the potency by more than 100-fold. Toxicity of the ADCs was assessed using bone marrow cells from a healthy human donor. The cells were exposed to the ADCs at 100 pM (a concentration highly potent against all AML samples) for 72 hours, and then markers of apoptosis were detected in different cell populations by flow cytometry. Neither ADC affected the viability of monocytes, lymphocytes and multipotential progenitors, consistent with low CD123 levels in these cell populations. In contrast, an apoptotic signal was detected in myeloid progenitors, the population with the highest CD123 level, following exposure to the C-ADC, but not to the A-ADC. The toxicity of the ADCs was also tested in CFU assays on bone marrow cells from 7 healthy donors, as the assays have been reported to predict clinical myelosuppression. Surprisingly, the C-ADC was, on average, 50-fold more cytotoxic to normal myeloid progenitors than the A-ADC (40 pM vs 2,000 pM IC90 values, respectively) (Figure 1). Finally, we compared CD123 independent toxicity of the ADCs in CD-1 mice. The C-ADC showed significantly reduced tolerability, and unlike the A-ADC, was associated with delayed toxicity manifested by weight loss 30 days after administration. Based on its potent yet highly selective toxicity to AML cells and more favorable tolerability profile, the A-ADC was selected for further study, and renamed as IMGN632. To compare IMGN632 to an ADC previously approved for the treatment of AML, the potency of IMGN632 and gemtuzumab ozogamicin (GO) was tested on bone marrow cells from 11 healthy donors and 17 AML patients, including 4 relapsed/refractory and 8 with strong multidrug resistance (Figure 1). Only 6 of 17 AML samples were sensitive to GO at concentrations that did not impact normal progenitors. In contrast, AML progenitors from all 17 patients were highly sensitive to IMGN632. Importantly, normal progenitors were only affected by IMGN632 at 150-fold higher concentrations. The pronounced difference between AML and normal progenitors in their sensitivity to IMGN632 likely reflects both higher CD123 levels on AML progenitors and the lower sensitivity of normal progenitors to the mono-imine IGN payload we observed in CFU assays. In conclusion, through use of a mono-imine IGN payload, IMGN632 demonstrated potent activity in all tested AML samples at concentrations far below levels that impact normal bone marrow cells, suggesting the potential for efficacy in AML patients in the absence of or with limited myelosuppression. These findings together with strong efficacy in multiple AML xenograft models (Kovtun et al., 21st EHA congress, 2016; Adams et al., 58th ASH annual meeting, 2016) support advancing IMGN632 into clinical trials. Disclosures Kovtun: ImmunoGen, Inc.: Employment. Jones:ImmunoGen, Inc.: Employment. Audette:ImmunoGen, Inc.: Employment. Harvey:ImmunoGen, Inc.: Employment. Gerard:ImmunoGen, Inc.: Employment. Wilhelm:ImmunoGen, Inc.: Employment. Bai:ImmunoGen, Inc.: Employment. Adams:ImmunoGen, Inc.: Employment. Goldmacher:ImmunoGen, Inc.: Employment. Chari:ImmunoGen: Employment. Chittenden:ImmunoGen, Inc.: Employment.

scholarly journals Specific toxicity of 2-chlorodeoxyadenosine toward resting and proliferating human lymphocytes

Blood ◽  
1983 ◽  
Vol 62 (4) ◽  
pp. 737-743 ◽  
Author(s):  
DA Carson ◽  
DB Wasson ◽  
R Taetle ◽  
A Yu
Keyword(s):  
Bone Marrow ◽  
Cell Lines ◽  
Bone Marrow Cells ◽  
Deoxycytidine Kinase ◽  
Thymidine Uptake ◽  
Lymphoblastoid Cell Lines ◽  
Normal Bone Marrow ◽  
Normal Peripheral Blood ◽  
Normal Bone ◽  
Marrow Cells

2-Chlorodeoxyadenosine (CdA), an adenosine-deaminase-resistant purine deoxynucleoside, is markedly toxic toward human T-lymphoblastoid cell lines in vitro and is an effective agent against L1210 leukemia in vivo. The present studies have examined the toxicity, and in some cases, metabolism, of CdA in (1) multiple established human cell lines of varying phenotype, (2) leukemia and lymphoma cells taken directly from patients, (3) normal bone marrow cells, and (4) normal peripheral blood lymphocytes. Nanomolar concentrations of CdA blocked the proliferation of lymphoblastoid cell lines with a high ratio of deoxycytidine kinase to deoxynucleotidase. The drug had virtually no effect on the growth of cell lines derived from solid tissues. The CdA inhibited the spontaneous uptake of tritiated thymidine by many T and non-T, non-B acute lymphoblastic leukemia cell specimens at concentrations less than or equal to 5 nM. The same concentrations did not impair either thymidine uptake or granulocyte-monocyte colony formation by normal bone marrow cells. In common with deoxyadenosine, but unlike several other agents affecting purine and purine metabolism, CdA was lethal to resting normal T lymphocytes and to slowly dividing malignant T cells. In both resting and proliferating lymphocytes, the CdA was phosphorylated by deoxycytidine kinase and entered a rapidly turning over nucleotide pool. Dividing lymphocytes also incorporated abundant CdA into DNA. The selective toxicity of CdA toward both dividing and resting lymphocytes may render the drug useful as an immunosuppressive or antileukemic agent.

CD52 Expression Patterns in Normal and Neoplastic B-Cells: Myeloma Is an Unlikely Target for Single Agent Alemtuzumab Therapy.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 4883-4883
Author(s):  
A. C. Rawstron ◽  
G. Laycock-Brown ◽  
F. E. Davies ◽  
R. G. Owen ◽  
P. Hillmen ◽  
...  
Keyword(s):  
Monoclonal Antibody ◽  
Bone Marrow ◽  
B Cells ◽  
B Cell ◽  
Plasma Cell ◽  
Plasma Cells ◽  
Single Agent ◽  
Normal Bone Marrow ◽  
Normal Bone ◽  
Normal Controls

Abstract Alemtuzumab is a highly effective monoclonal antibody therapy for some B-cell disorders, and has been suggested as a possible therapeutic agent for treatment of myeloma. Monoclonal antibody therapeutic efficacy is closely associated with the expression level of the therapeutic target, as demonstrated by the lack of efficacy of single-agent rituximab in CLL. However, there are conflicting reports about the expression levels of CD52, the target for alemtuzumab, in plasma cell disorders. The aim of this study is to assess a large series of cases of plasma cell and B-cell disorders, utilising a standard approach to allow comparison of the target molecule. Plasma cells were assessed from patients with myeloma at presentation or relapse (n=106), monoclonal gammopathy of undetermined significance (MGUS, n=34), and from normal controls (n=19). In addition, B-cells were assessed from patients with chronic lymphocytic leukaemia (CLL, n=87), diffuse large B-cell lymphoma (DLBCL, n=10), follicular lymphoma (FCL, n=9), Waldenstroms macroglobulinaemia (WM, n=20), and also from normal bone marrow (n=37). Normal and neoplastic B-cells showed expression of CD52 (&gt;20% of cells above the CD3-control levels) in all patients except for 1/10 DLBL. B-CLL and WM are known to show responses to single-agent alemtuzumab therapy, and these two disorders had the highest levels of expression. In contrast, B-progenitor cells in normal bone marrow are unaffected by alemtuzumab, and proliferate during alemtuzumab treatment in CLL patients. The levels of CD52 expression by normal B-progenitors were 3-fold lower than CLL/WM. In DLBL and FCL, the B-cells showed very similar levels of CD52 expression to normal B-progenitors, on average 2.8-fold lower than CLL. All plasma cells, whether neoplastic (CD19− or CD19+56+) or normal (CD19+56−), showed much lower levels of expression than normal and neoplastic B-cells. Plasma cell CD52 expression was detectable in 68% of normal controls (13/19), 50% of MGUS patients (17/34), and only 43% of myeloma patients (46/106). Expression was uni-modal in all cases. There was significantly lower expression of CD52 by myeloma plasma cells than by their normal counterparts (median 2.4-fold decrease, P=0.03). Neoplastic plasma cell CD52 expression showed a high degree of inter-patient variation, but fewer than 10% of myeloma patients (7/106) had CD52 expression at a similar level to CLL cells. Neoplastic plasma cell CD52 expression was approximately 6-fold lower than that of normal B-progenitors, and nearly 20-fold lower than that of CLL cells. In summary, CD52 expression is not detectable above control levels in a significant proportion of myeloma patients. In cases with detectable CD52 expression, the antigen is at a much lower level than is present on normal B-progenitors, which actively proliferate during alemtuzumab therapy. The risk of immunosuppression due to depletion of residual normal B/T-cells must also be considered. As alemtuzumab efficacy appears to correlate with CD52 expression levels, myeloma is highly unlikely to respond to alemtuzumab as a single agent except in rare cases. However, alemtuzumab is more likely to be effective in the IgM immunosecretory disorders which show strong CD52 expression.

Prognostication by miRNome-Profiling in Multiple Myeloma

Blood ◽  
2011 ◽  
Vol 118 (21) ◽  
pp. 1822-1822
Author(s):  
Anja Seckinger ◽  
Tobias Meißner ◽  
Vladimir Benes ◽  
Sabine Schmidt ◽  
Jonathan Blake ◽  
...  
Keyword(s):  
Gene Expression ◽  
Multiple Myeloma ◽  
Bone Marrow ◽  
Plasma Cells ◽  
Proliferation Index ◽  
Normal Bone Marrow ◽  
Global Test ◽  
Myeloma Cells ◽  
Normal Bone ◽  
Bone Marrow Plasma

Abstract Abstract 1822 INTRODUCTION. MicroRNAs are an abundant class of small non-protein-coding RNAs that function as negative gene regulators in diverse biological processes including cancer by affecting the stability and translation of mRNAs. METHODS. We determined expression of 559 miRNAs by miChip (Exiqon LNA Array probes V9.2) in CD138-purified myeloma cells from previously untreated patients (n=69), normal bone marrow plasma cells of healthy donors (pooled to n=3), and human myeloma cell lines (n=20). For normalization, an invariant-based method was applied. Gene expression profiling was performed using Affymetrix U133 2.0 DNA-microarrays. RESULTS. We found 29 miRNAs to be significantly up- and 35 down-regulated in myeloma cells vs. normal bone marrow plasma cells, respectively. Expression of 18 miRNAs was simultaneously significantly (P<.01) associated with event-free survival (EFS) and overall survival (OS), and allowed the delineation of prognostic groups. Of these, miRNAs miR-659 (located at 22q12.1) was significantly lower expressed in myeloma cells compared to normal bone marrow plasma cells. For this miRNA, low expression delineates a group with inferior EFS (median 19.7 months vs. not reached, P<.001) and OS (median 52.9 months vs. not reached, P<.001). This group shows a significantly higher gene expression based proliferation index. In contrast, high miR-590 expression (located at 7q11.23) delineates a group with inferior EFS (median 12.4 vs. 36 months, P<0.001) and OS (median 29.4 months vs. not reached, P<.001). By using Goeman's global test, a significant association of the predicted target gene signatures with survival could be found for both, EFS (miR-590-5p, P<.001; miR-659, P<.001) and OS (P=.009; P=.002). CONCLUSION. In conclusion, we demonstrate the prognostic relevance of miRNome profiling in multiple myeloma. Disclosures: No relevant conflicts of interest to declare.

scholarly journals Specific toxicity of 2-chlorodeoxyadenosine toward resting and proliferating human lymphocytes

Blood ◽  
1983 ◽  
Vol 62 (4) ◽  
pp. 737-743 ◽  
Author(s):  
DA Carson ◽  
DB Wasson ◽  
R Taetle ◽  
A Yu
Keyword(s):  
Bone Marrow ◽  
Cell Lines ◽  
Bone Marrow Cells ◽  
Deoxycytidine Kinase ◽  
Thymidine Uptake ◽  
Lymphoblastoid Cell Lines ◽  
Normal Bone Marrow ◽  
Normal Peripheral Blood ◽  
Normal Bone ◽  
Marrow Cells

Abstract 2-Chlorodeoxyadenosine (CdA), an adenosine-deaminase-resistant purine deoxynucleoside, is markedly toxic toward human T-lymphoblastoid cell lines in vitro and is an effective agent against L1210 leukemia in vivo. The present studies have examined the toxicity, and in some cases, metabolism, of CdA in (1) multiple established human cell lines of varying phenotype, (2) leukemia and lymphoma cells taken directly from patients, (3) normal bone marrow cells, and (4) normal peripheral blood lymphocytes. Nanomolar concentrations of CdA blocked the proliferation of lymphoblastoid cell lines with a high ratio of deoxycytidine kinase to deoxynucleotidase. The drug had virtually no effect on the growth of cell lines derived from solid tissues. The CdA inhibited the spontaneous uptake of tritiated thymidine by many T and non-T, non-B acute lymphoblastic leukemia cell specimens at concentrations less than or equal to 5 nM. The same concentrations did not impair either thymidine uptake or granulocyte-monocyte colony formation by normal bone marrow cells. In common with deoxyadenosine, but unlike several other agents affecting purine and purine metabolism, CdA was lethal to resting normal T lymphocytes and to slowly dividing malignant T cells. In both resting and proliferating lymphocytes, the CdA was phosphorylated by deoxycytidine kinase and entered a rapidly turning over nucleotide pool. Dividing lymphocytes also incorporated abundant CdA into DNA. The selective toxicity of CdA toward both dividing and resting lymphocytes may render the drug useful as an immunosuppressive or antileukemic agent.

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