Contributions
Abstract: S1553
Type: Oral Presentation
Presentation during EHA23: On Sunday, June 17, 2018 from 08:45 - 09:00
Location: Room A2
Background
Treatment with tyrosine kinase inhibitors (TKIs) fails to eradicate leukemia stem cells (LSC) in CML patients necessitating life-long therapy. Our recent work has shown that both p53 and c-Myc control the survival and proliferation of primitive CML cells (Abraham et al, Nature 2016), and that modulating both p53 and c-Myc can target CML LSC. Clinically, modulators of p53 and/or c-Myc are likely to be given in combination with TKI.
Aims
As our ultimate aim is to introduce novel therapies into the clinic that eliminate CML LSC, we examined the effect the MDM2 antagonist, idasanutlin (‘Idasa’) has in combination with nilotinib (‘Nil’) on leukemia cell numbers and LSC in vitro and in vivo.
Methods
We performed cell counts, apoptosis and cell cycle assays to determine combination indices (CI) and therefore synergy for drug combinations; and confirmed effects on primitive cells by long-term culture-initiating cell (LTC-IC) assay in vitro. To further examine the clinical utility of these drugs in CML we utilised our preclinical murine models: a patient derived xenograft (PDX) model and our double transgenic (DTG) SCL-tTA/BCR-ABL model.
Results
At the empirically derived ratio of 30:1 (Nil:Idasa) in primary CML CD34+ cells, the combination was synergistic by both cell count and apoptosis. At this ratio, the Nil/Idasa combination resulted in a significant increase in apoptosis compared to no drug control (NDC) or TKI alone (p<0.05; n=3), while no significance was reached for Nil/Idasa against normal samples (n=3). Nil/Idasa also resulted in a significant decrease in colony forming cell (CFC) counts compared to NDC or Nil alone (p<0.05; n=3), while no significant difference was seen in normal CD34+ cells. Moreover, by LTC-IC assay, compared to NDC, Nil/Idasa resulted in a significant decrease in output (p<0.05, n=3), while no significance was reached with the individual treatments. Again, no significant effect was seen in normal cells. Following transplant and long-term engraftment (12 weeks) of CML CD34+ cells into immunocompromised mice (PDX model), treatment with Idasa and Nil, either alone or in combination, resulted in a decrease in the percentage of human cells engrafted compared to vehicle alone, including the more primitive CD34+38- cells. However, while the percentage of human cells engrafted in the Nil-treated animals increased following 4 weeks recovery without drug, the percentage in the Idasa and combination arms remained the same, or even decreased. This was particularly evident in the more primitive cells, where Nil/Idasa combination resulted in the percentage of CD34+38- cells being significantly lower following recovery than both the vehicle and nil alone arms (p=0.029 & p=0.0107 respectively; n=4 vehicle / combo, n=3 Nil). In addition, when we treated our DTG CML mouse model with MDM2 antagonist (RG7112) and nil for 4 weeks following leukemia induction, we saw a significant decrease in the percentage of LSK cells in both the bone marrow and spleen with the combination compared to Nil alone (p<0.01 for both).
Conclusion
Overall these data show that pharmacological modulation of p53 in combination with TKI can target the most primitive CML cells, potentially in a synergistic manner. Moreover, recovery data from our PDX model suggests that this combination may abrogate the stem cell expansion which is seen with TKI alone following drug withdrawal, giving hope for treatment-free survival in patients. A planned clinical trial will test the efficacy of idasanutlin in combination with TKI.
Session topic: 7. Chronic myeloid leukemia – Biology & Translational Research
Keyword(s): Chronic myeloid leukemia
Abstract: S1553
Type: Oral Presentation
Presentation during EHA23: On Sunday, June 17, 2018 from 08:45 - 09:00
Location: Room A2
Background
Treatment with tyrosine kinase inhibitors (TKIs) fails to eradicate leukemia stem cells (LSC) in CML patients necessitating life-long therapy. Our recent work has shown that both p53 and c-Myc control the survival and proliferation of primitive CML cells (Abraham et al, Nature 2016), and that modulating both p53 and c-Myc can target CML LSC. Clinically, modulators of p53 and/or c-Myc are likely to be given in combination with TKI.
Aims
As our ultimate aim is to introduce novel therapies into the clinic that eliminate CML LSC, we examined the effect the MDM2 antagonist, idasanutlin (‘Idasa’) has in combination with nilotinib (‘Nil’) on leukemia cell numbers and LSC in vitro and in vivo.
Methods
We performed cell counts, apoptosis and cell cycle assays to determine combination indices (CI) and therefore synergy for drug combinations; and confirmed effects on primitive cells by long-term culture-initiating cell (LTC-IC) assay in vitro. To further examine the clinical utility of these drugs in CML we utilised our preclinical murine models: a patient derived xenograft (PDX) model and our double transgenic (DTG) SCL-tTA/BCR-ABL model.
Results
At the empirically derived ratio of 30:1 (Nil:Idasa) in primary CML CD34+ cells, the combination was synergistic by both cell count and apoptosis. At this ratio, the Nil/Idasa combination resulted in a significant increase in apoptosis compared to no drug control (NDC) or TKI alone (p<0.05; n=3), while no significance was reached for Nil/Idasa against normal samples (n=3). Nil/Idasa also resulted in a significant decrease in colony forming cell (CFC) counts compared to NDC or Nil alone (p<0.05; n=3), while no significant difference was seen in normal CD34+ cells. Moreover, by LTC-IC assay, compared to NDC, Nil/Idasa resulted in a significant decrease in output (p<0.05, n=3), while no significance was reached with the individual treatments. Again, no significant effect was seen in normal cells. Following transplant and long-term engraftment (12 weeks) of CML CD34+ cells into immunocompromised mice (PDX model), treatment with Idasa and Nil, either alone or in combination, resulted in a decrease in the percentage of human cells engrafted compared to vehicle alone, including the more primitive CD34+38- cells. However, while the percentage of human cells engrafted in the Nil-treated animals increased following 4 weeks recovery without drug, the percentage in the Idasa and combination arms remained the same, or even decreased. This was particularly evident in the more primitive cells, where Nil/Idasa combination resulted in the percentage of CD34+38- cells being significantly lower following recovery than both the vehicle and nil alone arms (p=0.029 & p=0.0107 respectively; n=4 vehicle / combo, n=3 Nil). In addition, when we treated our DTG CML mouse model with MDM2 antagonist (RG7112) and nil for 4 weeks following leukemia induction, we saw a significant decrease in the percentage of LSK cells in both the bone marrow and spleen with the combination compared to Nil alone (p<0.01 for both).
Conclusion
Overall these data show that pharmacological modulation of p53 in combination with TKI can target the most primitive CML cells, potentially in a synergistic manner. Moreover, recovery data from our PDX model suggests that this combination may abrogate the stem cell expansion which is seen with TKI alone following drug withdrawal, giving hope for treatment-free survival in patients. A planned clinical trial will test the efficacy of idasanutlin in combination with TKI.
Session topic: 7. Chronic myeloid leukemia – Biology & Translational Research
Keyword(s): Chronic myeloid leukemia