Activation of primary human chronic lymphocytic leukemia (cll) cells by interleukin-2 (il-2)

implications for cancer vaccine development

Michael Robertson, Kristen Ganjoo, Lynette Timmons

Research output: Contribution to journalArticle

Abstract

CLL, the most common leukemia in Western societies, is incurable by conventional therapy and new treatment approaches are needed. The cancer vaccine strategy attempts to stimulate an effective immune response against autologous tumor. Primary CLL cells are known to be poorly immunogenic. Nevertheless, CLL cells can be readily isolated from peripheral blood samples and manipulated in vitro to enhance their immunogenicity. Samples of peripheral blood were obtained from 12 patients with typical CD19+/CD5+ CLL who gave written informed consent. Mononuclear cells were isolated by density gradient centrifugation and cultured for 5 days in medium alone or with various stimuli. Tritiated thymidine was added during the last 18 hours of culture and thymidine incorporation was measured by liquid scintillation counting. rhIL-2 at 10 to 1000 picomolar (pM) induced dose-dependent proliferation of primary CLL cells (Table). (Values are mean ±SE of CPM from 12 different CLL samples. P values are from paired t-test comparing IL-2 conditions to medium alone.) Medium IL-2 10 pM IL-2 100 pM IL-2 1000 pM 970 ± 208 2784 ± 710 4952± 1397 6371 ± 1748 N.A. P ≤ 0.01 P ≤ 0.01 P ≤ 0.005 In contrast, significant proliferation was not detected in response to IL-4 at doses of 1, 10, or 100 ng/ml (not shown). Immunophenotyping of CLL cell cultures incubated with 1000 pM IL-2 for 2, 5, and 7 days confirmed that proliferation was not due to growth of contaminating normal cells. CLL cells in these cultures were strongly activated, as indicated by increases in their forward- and side- light scattering properties. Moreover, the apoptosis receptor CD95 ( APO-1 /Fas) was substantially upregulated on CLL cells incubated in 1000 pM IL-2 for 5 days: 99 ±0.3% (mean ±SE of 3 separate samples) of these cells expressed CD95, compared to 12 ±3% of cells in medium alone (P < 0.0005) or 11 < 3% of cells in 10 ng/ml of GM-CSF (P<0.0005). These results indicate that IL-2 by itself can induce the activation and proliferation of primary human CLL cells. Ongoing studies will determine whether IL-2-activated CLL cells are immunogenic and whether they are susceptible to retrovirally-mediated gene transfer in attempts to further enhance immunogenicity.

Original languageEnglish
JournalBlood
Volume96
Issue number11 PART II
StatePublished - 2000

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Cancer Vaccines
B-Cell Chronic Lymphocytic Leukemia
Interleukin-2
Chemical activation
Cell culture
Thymidine
Blood
Gene transfer
Centrifugation
Cell Culture Techniques
Scintillation
Granulocyte-Macrophage Colony-Stimulating Factor
Interleukin-4
Light scattering
Scintillation Counting
Immunophenotyping
Tumors
Density Gradient Centrifugation
Interleukin-5
Cells

ASJC Scopus subject areas

  • Hematology

Cite this

Activation of primary human chronic lymphocytic leukemia (cll) cells by interleukin-2 (il-2) : implications for cancer vaccine development. / Robertson, Michael; Ganjoo, Kristen; Timmons, Lynette.

In: Blood, Vol. 96, No. 11 PART II, 2000.

Research output: Contribution to journalArticle

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title = "Activation of primary human chronic lymphocytic leukemia (cll) cells by interleukin-2 (il-2): implications for cancer vaccine development",
abstract = "CLL, the most common leukemia in Western societies, is incurable by conventional therapy and new treatment approaches are needed. The cancer vaccine strategy attempts to stimulate an effective immune response against autologous tumor. Primary CLL cells are known to be poorly immunogenic. Nevertheless, CLL cells can be readily isolated from peripheral blood samples and manipulated in vitro to enhance their immunogenicity. Samples of peripheral blood were obtained from 12 patients with typical CD19+/CD5+ CLL who gave written informed consent. Mononuclear cells were isolated by density gradient centrifugation and cultured for 5 days in medium alone or with various stimuli. Tritiated thymidine was added during the last 18 hours of culture and thymidine incorporation was measured by liquid scintillation counting. rhIL-2 at 10 to 1000 picomolar (pM) induced dose-dependent proliferation of primary CLL cells (Table). (Values are mean ±SE of CPM from 12 different CLL samples. P values are from paired t-test comparing IL-2 conditions to medium alone.) Medium IL-2 10 pM IL-2 100 pM IL-2 1000 pM 970 ± 208 2784 ± 710 4952± 1397 6371 ± 1748 N.A. P ≤ 0.01 P ≤ 0.01 P ≤ 0.005 In contrast, significant proliferation was not detected in response to IL-4 at doses of 1, 10, or 100 ng/ml (not shown). Immunophenotyping of CLL cell cultures incubated with 1000 pM IL-2 for 2, 5, and 7 days confirmed that proliferation was not due to growth of contaminating normal cells. CLL cells in these cultures were strongly activated, as indicated by increases in their forward- and side- light scattering properties. Moreover, the apoptosis receptor CD95 ( APO-1 /Fas) was substantially upregulated on CLL cells incubated in 1000 pM IL-2 for 5 days: 99 ±0.3{\%} (mean ±SE of 3 separate samples) of these cells expressed CD95, compared to 12 ±3{\%} of cells in medium alone (P < 0.0005) or 11 < 3{\%} of cells in 10 ng/ml of GM-CSF (P<0.0005). These results indicate that IL-2 by itself can induce the activation and proliferation of primary human CLL cells. Ongoing studies will determine whether IL-2-activated CLL cells are immunogenic and whether they are susceptible to retrovirally-mediated gene transfer in attempts to further enhance immunogenicity.",
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AU - Ganjoo, Kristen

AU - Timmons, Lynette

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N2 - CLL, the most common leukemia in Western societies, is incurable by conventional therapy and new treatment approaches are needed. The cancer vaccine strategy attempts to stimulate an effective immune response against autologous tumor. Primary CLL cells are known to be poorly immunogenic. Nevertheless, CLL cells can be readily isolated from peripheral blood samples and manipulated in vitro to enhance their immunogenicity. Samples of peripheral blood were obtained from 12 patients with typical CD19+/CD5+ CLL who gave written informed consent. Mononuclear cells were isolated by density gradient centrifugation and cultured for 5 days in medium alone or with various stimuli. Tritiated thymidine was added during the last 18 hours of culture and thymidine incorporation was measured by liquid scintillation counting. rhIL-2 at 10 to 1000 picomolar (pM) induced dose-dependent proliferation of primary CLL cells (Table). (Values are mean ±SE of CPM from 12 different CLL samples. P values are from paired t-test comparing IL-2 conditions to medium alone.) Medium IL-2 10 pM IL-2 100 pM IL-2 1000 pM 970 ± 208 2784 ± 710 4952± 1397 6371 ± 1748 N.A. P ≤ 0.01 P ≤ 0.01 P ≤ 0.005 In contrast, significant proliferation was not detected in response to IL-4 at doses of 1, 10, or 100 ng/ml (not shown). Immunophenotyping of CLL cell cultures incubated with 1000 pM IL-2 for 2, 5, and 7 days confirmed that proliferation was not due to growth of contaminating normal cells. CLL cells in these cultures were strongly activated, as indicated by increases in their forward- and side- light scattering properties. Moreover, the apoptosis receptor CD95 ( APO-1 /Fas) was substantially upregulated on CLL cells incubated in 1000 pM IL-2 for 5 days: 99 ±0.3% (mean ±SE of 3 separate samples) of these cells expressed CD95, compared to 12 ±3% of cells in medium alone (P < 0.0005) or 11 < 3% of cells in 10 ng/ml of GM-CSF (P<0.0005). These results indicate that IL-2 by itself can induce the activation and proliferation of primary human CLL cells. Ongoing studies will determine whether IL-2-activated CLL cells are immunogenic and whether they are susceptible to retrovirally-mediated gene transfer in attempts to further enhance immunogenicity.

AB - CLL, the most common leukemia in Western societies, is incurable by conventional therapy and new treatment approaches are needed. The cancer vaccine strategy attempts to stimulate an effective immune response against autologous tumor. Primary CLL cells are known to be poorly immunogenic. Nevertheless, CLL cells can be readily isolated from peripheral blood samples and manipulated in vitro to enhance their immunogenicity. Samples of peripheral blood were obtained from 12 patients with typical CD19+/CD5+ CLL who gave written informed consent. Mononuclear cells were isolated by density gradient centrifugation and cultured for 5 days in medium alone or with various stimuli. Tritiated thymidine was added during the last 18 hours of culture and thymidine incorporation was measured by liquid scintillation counting. rhIL-2 at 10 to 1000 picomolar (pM) induced dose-dependent proliferation of primary CLL cells (Table). (Values are mean ±SE of CPM from 12 different CLL samples. P values are from paired t-test comparing IL-2 conditions to medium alone.) Medium IL-2 10 pM IL-2 100 pM IL-2 1000 pM 970 ± 208 2784 ± 710 4952± 1397 6371 ± 1748 N.A. P ≤ 0.01 P ≤ 0.01 P ≤ 0.005 In contrast, significant proliferation was not detected in response to IL-4 at doses of 1, 10, or 100 ng/ml (not shown). Immunophenotyping of CLL cell cultures incubated with 1000 pM IL-2 for 2, 5, and 7 days confirmed that proliferation was not due to growth of contaminating normal cells. CLL cells in these cultures were strongly activated, as indicated by increases in their forward- and side- light scattering properties. Moreover, the apoptosis receptor CD95 ( APO-1 /Fas) was substantially upregulated on CLL cells incubated in 1000 pM IL-2 for 5 days: 99 ±0.3% (mean ±SE of 3 separate samples) of these cells expressed CD95, compared to 12 ±3% of cells in medium alone (P < 0.0005) or 11 < 3% of cells in 10 ng/ml of GM-CSF (P<0.0005). These results indicate that IL-2 by itself can induce the activation and proliferation of primary human CLL cells. Ongoing studies will determine whether IL-2-activated CLL cells are immunogenic and whether they are susceptible to retrovirally-mediated gene transfer in attempts to further enhance immunogenicity.

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