TY - CONF
T1 - Potential therapeutic role of seliciclib in combination with ionizing radiation for human nasopharyngeal carcinoma
AU - Yue, Shijun
AU - Hui, Angela
AU - Alajez, Nehad
AU - Frame, Sheelagh
AU - Liu, Fei Fei
AU - Shi, Wei
AU - Green, Simon R.
PY - 2008
Y1 - 2008
N2 - Introduction:
Seliciclib is a small molecule CDK inhibitor, with anti-proliferative and anti-tumoral effects, which is currently undergoing evaluation in Phase II clinical trials. No preclinical studies have been undertaken in EBV-positive nasopharyngeal carcinoma (NPC), hence the focus of this work was to evaluate the therapeutic efficacy of seliciclib alone and in combination with ionizing radiation (IR), on the C666-1 NPC model.
Experimental Procedure:
The MTS assay was utilized to measure viability after C666-1 cells were treated with seliciclib + IR. Apoptosis and cell cycle effects were evaluated using flow cytometric analysis. The proportion of cells containing DNA double-strand breaks was assessed using γ-H2AX immunofluorescence. Downstream biochemical effects of seliciclib on Mcl-1 mRNA were assessed using quantitative real time PCR. Therapeutic efficacy of seliciclib + IR was evaluated on established C666-1 tumours implanted in SCID mice.
Results:
Seliciclib reduced viability of C666-1 cells in both a time and dose-dependent manner, with an IC25 of 20 µM. The optimal sequence when combined with IR (4 Gy) was seliciclib treatment first, followed 24 hrs later by IR, which reduced viability down to 20% by day 6. Cell cycle analysis demonstrated accumulation of cells in G2/M, along with a significant increase in the proportion of cells in sub-G1, from 4% under control conditions, up to 14% when treated with both seliciclib (20 µM) and IR (2 Gy), at 72 hrs. This was associated with a increase in caspase-2,-3, -8, -9 activation by 5, 4, 3 and 4-fold respectively, at 48 hrs. Likewise, the proportion of γH2AX-expressing cells post-IR increased to 20% after seliciclib, compared to 14% with IR alone. Quantitative RT-PCR confirmed that seliciclib treatment reduced Mcl-1 transcript levels down to 18% of control, at 72 hrs. In vivo, control mice reached the predefined end-point (tumour and leg diameter 15 mm) at day 15; seliciclib alone prolonged survival by an average of 11 days. Using IR, survival was 57 days, when seliciclib was combined with IR (6 Gy), survival extended to 79 days, with tumour and leg diameter reaching only ~12 mm.
Conclusion:
The NPC model was highly sensitive to seliciclib, with at least an additive interaction when combined with IR, efficiently inducing G2M arrest, apoptosis mediated by caspase activation, along with increased DNA double-strand breaks. Seliciclib combined with IR is also efficacious in vivo, with significant prolongation in survival of tumour-bearing mice, with no discernible toxicity. These data support the evaluation of this combinatorial strategy for human NPC.
AB - Introduction:
Seliciclib is a small molecule CDK inhibitor, with anti-proliferative and anti-tumoral effects, which is currently undergoing evaluation in Phase II clinical trials. No preclinical studies have been undertaken in EBV-positive nasopharyngeal carcinoma (NPC), hence the focus of this work was to evaluate the therapeutic efficacy of seliciclib alone and in combination with ionizing radiation (IR), on the C666-1 NPC model.
Experimental Procedure:
The MTS assay was utilized to measure viability after C666-1 cells were treated with seliciclib + IR. Apoptosis and cell cycle effects were evaluated using flow cytometric analysis. The proportion of cells containing DNA double-strand breaks was assessed using γ-H2AX immunofluorescence. Downstream biochemical effects of seliciclib on Mcl-1 mRNA were assessed using quantitative real time PCR. Therapeutic efficacy of seliciclib + IR was evaluated on established C666-1 tumours implanted in SCID mice.
Results:
Seliciclib reduced viability of C666-1 cells in both a time and dose-dependent manner, with an IC25 of 20 µM. The optimal sequence when combined with IR (4 Gy) was seliciclib treatment first, followed 24 hrs later by IR, which reduced viability down to 20% by day 6. Cell cycle analysis demonstrated accumulation of cells in G2/M, along with a significant increase in the proportion of cells in sub-G1, from 4% under control conditions, up to 14% when treated with both seliciclib (20 µM) and IR (2 Gy), at 72 hrs. This was associated with a increase in caspase-2,-3, -8, -9 activation by 5, 4, 3 and 4-fold respectively, at 48 hrs. Likewise, the proportion of γH2AX-expressing cells post-IR increased to 20% after seliciclib, compared to 14% with IR alone. Quantitative RT-PCR confirmed that seliciclib treatment reduced Mcl-1 transcript levels down to 18% of control, at 72 hrs. In vivo, control mice reached the predefined end-point (tumour and leg diameter 15 mm) at day 15; seliciclib alone prolonged survival by an average of 11 days. Using IR, survival was 57 days, when seliciclib was combined with IR (6 Gy), survival extended to 79 days, with tumour and leg diameter reaching only ~12 mm.
Conclusion:
The NPC model was highly sensitive to seliciclib, with at least an additive interaction when combined with IR, efficiently inducing G2M arrest, apoptosis mediated by caspase activation, along with increased DNA double-strand breaks. Seliciclib combined with IR is also efficacious in vivo, with significant prolongation in survival of tumour-bearing mice, with no discernible toxicity. These data support the evaluation of this combinatorial strategy for human NPC.
M3 - Paper
ER -