Frequent and perhaps obligatory genetic alteration affecting the cdk4/6-cyclin D-INK4-Rb growth regulatory axis in GBM is well documented (2–8; in the current report), and has been recently corroborated in two large-scale, multi-institutional genomic analyses of GBM (9
). The most common alteration of this pathway in GBM is homozygous deletion of CDKN2A/B, encoding p16INK4a
, present in greater than 50% of tumors. Other alterations include amplification and overexpression of CDK4 (15–20%) and homozygous deletion/mutation of RB1 (~10%). Amplification of CDK6 and individual D-type cyclins, and homozygous deletion of CDKN2C encoding p18INK4c
are less common. Of these alterations, only genetic inactivation of RB1 itself is thought to render a tumor resistant to inhibition of cdk4/6. Since genetic inactivation of RB1 occurs infrequently in GBM, a substantial majority of GBM patients are predicted to be candidates for therapies targeting cdk4/6.
Several previous studies have tested the effects of kinase inhibitors against GBM (e.g.
imatinib, erlotinib, flavopiridol), both in a preclinical setting and in clinical trials without significant efficacy (27
). While some of these compounds promiscuously inhibit cyclin-dependent kinases (e.g.
flavopiridol), none of the previously tested compounds display selectivity for cyclin-dependent kinases relative to other kinases that may or may not be activated in GBM. Our study for the first time tests a cdk-specific inhibitor in the treatment of GBM, demonstrating significant efficacy both in vitro
and in vivo
as a single agent and in combination with radiation therapy.
PD-0332991 is an orally available pyridopyrimidine derivative that selectively inhibits cyclin-dependent kinases 4 and 6 (12
), leading to a reduction in Rb phosphorylation and subsequent cell cycle arrest. The in vitro
and in vivo
results presented here further emphasize that Rb is the primary determinant of sensitivity to cdk4/6 inhibition. This aspect of PD-0332991 efficacy should be especially attractive with respect to clinical trial evaluation, since there are excellent reagents and protocols established for immunohistochemical detection of Rb in paraffin-embedded tissues (14
Though Rb expression is clearly the primary determinant of tumor cell response to PD-0332991, variable growth inhibition among Rb-proficient cell lines () suggests the existence of secondary factors that influence tumor cell sensitivity to PD-0332991, such as tumor CDK4/6 amplification vs. CDKN2A homozygous deletion. For the cell lines examined here, the single case with CDK6 amplification (CCF-STTG1) showed substantial sensitivity to PD-0332991 ().
Analysis of GBM response to PD-0332991 in vivo, using three different GBM tumor cell sources for establishing intracranial tumors (), yielded results entirely consistent with the in vitro data. PD-0332991 arrested the growth of xenografts generated from U87MG cells and GBM 39 (both Rb-proficient) and led to significantly improved survival (). The growth inhibitory effect of PD-0332991 was remarkably durable during the period of treatment.
Recent studies point to interest in investigating anti-tumor effects of PD-0332991 in combination with therapeutics that affect tumor properties other than cell cycling. For instance, it has been shown that PD-0332991, which by itself does not promote apoptotic response of cancer cells, markedly enhances myeloma cell killing by dexamethasone (31
) as well as by bortezomib (32
). Similarly, PD-0332991 has been shown to enhance breast cancer cell sensitivity to tamoxifen in vitro
). Here, our bioluminescence and survival analysis of mice with intracranial U87MG tumors indicate that the anti-tumor activity of PD-0332991 when used with radiation, either concurrently or sequentially, is superior to using either agent as a monotherapy (). These results, therefore, could help motivate clinical trial testing of PD-0332991 against newly-diagnosed GBM, for which the use of radiotherapy is the standard of care.
Finally, treatment of intracranial GBM tumors that had re-grown following initial therapy with temozolomide demonstrated that PD-0332991 has activity against recurrent GBM (). These results additionally suggest a general approach for pre-clinical animal model testing of therapies for patients with recurrent brain tumors, which is a mostly neglected area of neurooncology research.
In total, these findings provide strong support for evaluating the efficacy of PD-0332991 in treating GBM patients. For future investigation, it will be interesting to examine combination therapies involving PD-0332991 with small molecule inhibitors targeting activated gene products in other GBM core pathways, such as erlotinib for inhibiting EGFR. Determination of the full range of applications of this cdk4/6 inhibitor will undoubtedly prove informative, and will hopefully lead to improved treatment for this devastating cancer.