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Nilotinib treatment in mouse models of P190 Bcr/Abl lymphoblastic leukemia.

Kaur P, Feldhahn N, Zhang B, Trageser D, Müschen M, Pertz V, Groffen J, Heisterkamp N - Mol. Cancer (2007)

Bottom Line: In addition, culture of such cells ex vivo showed that they were as sensitive as the parental cell line to nilotinib but that the presence of stromal support allowed resistant cells to grow out.Visible lymphoma masses disappeared within six days of treatment and leukemic cell numbers in peripheral blood were significantly reduced.These results show that nilotinib has very impressive anti-leukemia activity but that lymphoblastic leukemia cells can become unresponsive to it both in vitro and in vivo through mechanisms that appear to be Bcr/Abl independent.

View Article: PubMed Central - HTML - PubMed

Affiliation: Section of Molecular Carcinogenesis, Division of Hematology/Oncology, Saban Research Institute, Childrens Hospital Los Angeles and the Keck School of Medicine, University of Southern California, Los Angeles, California, USA. pavinder@gmail.com

ABSTRACT

Background: Ph-positive leukemias are caused by the aberrant fusion of the BCR and ABL genes. Nilotinib is a selective Bcr/Abl tyrosine kinase inhibitor related to imatinib, which is widely used to treat chronic myelogenous leukemia. Because Ph-positive acute lymphoblastic leukemia only responds transiently to imatinib therapy, we have used mouse models to test the efficacy of nilotinib against lymphoblastic leukemia caused by the P190 form of Bcr/Abl.

Results: After transplant of 10,000 highly malignant leukemic cells into compatible recipients, untreated mice succumbed to leukemia within 21 days, whereas mice treated with 75 mg/kg nilotinib survived significantly longer. We examined cells from mice that developed leukemia while under treatment for Bcr/Abl kinase domain point mutations but these were not detected. In addition, culture of such cells ex vivo showed that they were as sensitive as the parental cell line to nilotinib but that the presence of stromal support allowed resistant cells to grow out. Nilotinib also exhibited impressive anti-leukemia activity in P190 Bcr/Abl transgenic mice that had developed overt leukemia/lymphoma masses and that otherwise would have been expected to die within 7 days. Visible lymphoma masses disappeared within six days of treatment and leukemic cell numbers in peripheral blood were significantly reduced. Treated mice survived more than 30 days.

Conclusion: These results show that nilotinib has very impressive anti-leukemia activity but that lymphoblastic leukemia cells can become unresponsive to it both in vitro and in vivo through mechanisms that appear to be Bcr/Abl independent.

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Effect of Jak inhibitor AG490 on survival of nilotinib-insensitive cells in the presence of stroma. 3 × 106 lymphoma cells were seeded in 6 well tissue culture plates with irradiated MEFs and further treated with 5, 50 or 100 μM AG490 (A) or 20 nM nilotinib (B, C), respectively. In (B) and (C), in addition to nilotinib, treatment with 5, 50 or 75 μM AG490 was initiated as indicated by an arrow. As a control for the effect of AG490, single treatment with DMSO only (A) or 20 nM nilotinib (B, C) was analyzed, respectively. Fresh drug was added (depending on proliferation of the leukemia cells) every second or third day with the change of medium. Viability was assessed by propidium iodide uptake using a FACScan. Each point represents the average of triplicate values ± SEM.
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Figure 6: Effect of Jak inhibitor AG490 on survival of nilotinib-insensitive cells in the presence of stroma. 3 × 106 lymphoma cells were seeded in 6 well tissue culture plates with irradiated MEFs and further treated with 5, 50 or 100 μM AG490 (A) or 20 nM nilotinib (B, C), respectively. In (B) and (C), in addition to nilotinib, treatment with 5, 50 or 75 μM AG490 was initiated as indicated by an arrow. As a control for the effect of AG490, single treatment with DMSO only (A) or 20 nM nilotinib (B, C) was analyzed, respectively. Fresh drug was added (depending on proliferation of the leukemia cells) every second or third day with the change of medium. Viability was assessed by propidium iodide uptake using a FACScan. Each point represents the average of triplicate values ± SEM.

Mentions: We next examined a possible mechanism leading to Bcr/Abl-independent resistance to nilotinib. Samantha et al [36] showed that Jak2 is an important target in CML, and the Jak inhibitor AG490 was able to induce apoptosis in cells that expressed imatinib-resistant mutants of Bcr/Abl. Very recently, Wang et al [37] further implicated Jak2 in Bcr/Abl-independent imatinib and nilotinib resistance caused by GM-CSF production by myeloid leukemic cells. Therefore, using the Jak inhibitor AG490, we investigated if Jak2, in addition to its involvement in drug resistance of myeloid leukemia cells, also contributes to resistance development of lymphoid leukemia cells. As shown in Fig. 6A, AG490 treatment significantly decreased the survival of the lymphoid leukemia cells in a dose-dependent manner when these cells were co-cultured with MEFs. Interestingly, AG490 treatment for 48 hours also affected normal function of the feeder layer cells, as the proliferation of non-irradiated MEFs was severely reduced (more than 4 times reduction in the presence of 50 μM AG490; results not shown) compared to treatment with the vehicle DMSO. Treatment of the Bcr/Abl lymphoblastic leukemia cells with AG490 during (Fig. 6B) and after (Fig. 6C) resistance development to nilotinib did not further affect the survival, as compared to its effect on non-resistant leukemia cells (Fig. 6A). Instead, in both experiments, nilotinib-resistant lymphoblastic leukemia cells seemed to also acquire additional resistance to AG490, though in a dose dependent manner, as evidenced by the resumption of growth after an initial drop in viability upon first addition of AG490 (Fig. 6C).


Nilotinib treatment in mouse models of P190 Bcr/Abl lymphoblastic leukemia.

Kaur P, Feldhahn N, Zhang B, Trageser D, Müschen M, Pertz V, Groffen J, Heisterkamp N - Mol. Cancer (2007)

Effect of Jak inhibitor AG490 on survival of nilotinib-insensitive cells in the presence of stroma. 3 × 106 lymphoma cells were seeded in 6 well tissue culture plates with irradiated MEFs and further treated with 5, 50 or 100 μM AG490 (A) or 20 nM nilotinib (B, C), respectively. In (B) and (C), in addition to nilotinib, treatment with 5, 50 or 75 μM AG490 was initiated as indicated by an arrow. As a control for the effect of AG490, single treatment with DMSO only (A) or 20 nM nilotinib (B, C) was analyzed, respectively. Fresh drug was added (depending on proliferation of the leukemia cells) every second or third day with the change of medium. Viability was assessed by propidium iodide uptake using a FACScan. Each point represents the average of triplicate values ± SEM.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2169263&req=5

Figure 6: Effect of Jak inhibitor AG490 on survival of nilotinib-insensitive cells in the presence of stroma. 3 × 106 lymphoma cells were seeded in 6 well tissue culture plates with irradiated MEFs and further treated with 5, 50 or 100 μM AG490 (A) or 20 nM nilotinib (B, C), respectively. In (B) and (C), in addition to nilotinib, treatment with 5, 50 or 75 μM AG490 was initiated as indicated by an arrow. As a control for the effect of AG490, single treatment with DMSO only (A) or 20 nM nilotinib (B, C) was analyzed, respectively. Fresh drug was added (depending on proliferation of the leukemia cells) every second or third day with the change of medium. Viability was assessed by propidium iodide uptake using a FACScan. Each point represents the average of triplicate values ± SEM.
Mentions: We next examined a possible mechanism leading to Bcr/Abl-independent resistance to nilotinib. Samantha et al [36] showed that Jak2 is an important target in CML, and the Jak inhibitor AG490 was able to induce apoptosis in cells that expressed imatinib-resistant mutants of Bcr/Abl. Very recently, Wang et al [37] further implicated Jak2 in Bcr/Abl-independent imatinib and nilotinib resistance caused by GM-CSF production by myeloid leukemic cells. Therefore, using the Jak inhibitor AG490, we investigated if Jak2, in addition to its involvement in drug resistance of myeloid leukemia cells, also contributes to resistance development of lymphoid leukemia cells. As shown in Fig. 6A, AG490 treatment significantly decreased the survival of the lymphoid leukemia cells in a dose-dependent manner when these cells were co-cultured with MEFs. Interestingly, AG490 treatment for 48 hours also affected normal function of the feeder layer cells, as the proliferation of non-irradiated MEFs was severely reduced (more than 4 times reduction in the presence of 50 μM AG490; results not shown) compared to treatment with the vehicle DMSO. Treatment of the Bcr/Abl lymphoblastic leukemia cells with AG490 during (Fig. 6B) and after (Fig. 6C) resistance development to nilotinib did not further affect the survival, as compared to its effect on non-resistant leukemia cells (Fig. 6A). Instead, in both experiments, nilotinib-resistant lymphoblastic leukemia cells seemed to also acquire additional resistance to AG490, though in a dose dependent manner, as evidenced by the resumption of growth after an initial drop in viability upon first addition of AG490 (Fig. 6C).

Bottom Line: In addition, culture of such cells ex vivo showed that they were as sensitive as the parental cell line to nilotinib but that the presence of stromal support allowed resistant cells to grow out.Visible lymphoma masses disappeared within six days of treatment and leukemic cell numbers in peripheral blood were significantly reduced.These results show that nilotinib has very impressive anti-leukemia activity but that lymphoblastic leukemia cells can become unresponsive to it both in vitro and in vivo through mechanisms that appear to be Bcr/Abl independent.

View Article: PubMed Central - HTML - PubMed

Affiliation: Section of Molecular Carcinogenesis, Division of Hematology/Oncology, Saban Research Institute, Childrens Hospital Los Angeles and the Keck School of Medicine, University of Southern California, Los Angeles, California, USA. pavinder@gmail.com

ABSTRACT

Background: Ph-positive leukemias are caused by the aberrant fusion of the BCR and ABL genes. Nilotinib is a selective Bcr/Abl tyrosine kinase inhibitor related to imatinib, which is widely used to treat chronic myelogenous leukemia. Because Ph-positive acute lymphoblastic leukemia only responds transiently to imatinib therapy, we have used mouse models to test the efficacy of nilotinib against lymphoblastic leukemia caused by the P190 form of Bcr/Abl.

Results: After transplant of 10,000 highly malignant leukemic cells into compatible recipients, untreated mice succumbed to leukemia within 21 days, whereas mice treated with 75 mg/kg nilotinib survived significantly longer. We examined cells from mice that developed leukemia while under treatment for Bcr/Abl kinase domain point mutations but these were not detected. In addition, culture of such cells ex vivo showed that they were as sensitive as the parental cell line to nilotinib but that the presence of stromal support allowed resistant cells to grow out. Nilotinib also exhibited impressive anti-leukemia activity in P190 Bcr/Abl transgenic mice that had developed overt leukemia/lymphoma masses and that otherwise would have been expected to die within 7 days. Visible lymphoma masses disappeared within six days of treatment and leukemic cell numbers in peripheral blood were significantly reduced. Treated mice survived more than 30 days.

Conclusion: These results show that nilotinib has very impressive anti-leukemia activity but that lymphoblastic leukemia cells can become unresponsive to it both in vitro and in vivo through mechanisms that appear to be Bcr/Abl independent.

Show MeSH
Related in: MedlinePlus