Epidermal growth factor

The EGFR pathway is an attractive target because it is frequently dysregulated in GBMs through overexpression, amplification, or activating mutations. EGFR dysregulation results in aberrant signaling which, in turn, is associated with increased proliferation, motility, and survival of tumor cells in vitro. The most common EGFR mutation, found in 20% to 30% of GBMs, is EGFRvIII, which causes constitutive activation and phosphorylation of the receptor without ligand binding. Several approaches exist for targeting EGFR including small-molecule tyrosine kinase inhibitors (TKIs), monoclonal antibodies (mAbs), and vaccines.

The most extensively studied TKIs in GBM are erlotinib and gefitinib. Both are oral, low molecular weight, reversible inhibitors of tyrosine kinase activity associated with EGFR. When combined with RT in newly diagnosed GBM patients, the addition of these agents did not prolong 6-month progression-free survival (6M-PFS) or overall survival (OS) in multicenter trials as compared with standard therapy. In several phase 2 clinical trials of recurrent high-grade gliomas, neither gefitinb nor erlotinib monotherapy demonstrated any significant survival benefit as compared with historical controls. Clinical activity was modest in the GBM subset, with radiographic responses (RR) ranging from 0% to 13% for gefitinib and 0% to 26% for erlotinib, and with 6M-PFS ranging from 9% to 13% for gefitinib and 0% to 26% for erlotinib. Small subgroups of patients had durable responses. Both TKIs were generally well tolerated with rash, diarrhea, and fatigue as the most common side effects. The combination of EGFR inhibitors with other therapies is discussed later in this article.

Lapatinib, which inhibits ErbB2/HER2 (human epidermal growth factor receptor 2) as well as EGFR, has also been evaluated in recurrent glioblastomas. This agent achieves therapeutic concentrations in gliomas but has limited activity.

Although subsets of GBM patients have sustained responses to reversible TKIs that target EGFR, the studies to date have been largely disappointing. Potential reasons for lack of response include poor blood-brain barrier penetration, insufficient local tumor concentrations, coactivation of multiple TKIs, redundant signaling pathways, and resistance. Irreversible EGFR inhibitors, such as BIBW 2992 and PF-00299804, could have better efficacy in GBM than gefitinib or elotinib due to increased potency and in the case of PF-00299804, better brain concentration. This newer class of EGFR inhibitors has been shown to circumvent mechanisms of response to gefitinib or erlotinib in non–small cell lung cancer cells.

Attempts to define subsets of erlotinib or gefitinib responders have also been met with limited success. The activating mutations found in EGFR in non–small cell lung cancer that increase response to EGFR inhibitors are not present in GBM. Initial molecular studies suggested that gefitinib and erlotinib were more effective in tumors with EGFRvIII mutations and intact PTEN. However, recent studies failed to validate these results.

Monoclonal antibodies (mAb) as well as vaccines that target EGFR are currently under investigation in GBM. These therapies have been well tolerated in early clinical trials. Results from a phase 1/2 study combining nimotuzumab (a humanized anti-EGFR mAb) with radiotherapy in patients were favorable ( Table 2 ). Both nimotuzumab and cetuximab, a chimeric anti-EGFR human-mouse mAb, are now being studied in combination with radiation and temozolomide as upfront GBM therapies. Preliminary results from a phase 2 clinical trial suggest that the addition of CDX-110, a peptide based EGFRvIII vaccine, to standard therapy prolongs survival in patients with newly diagnosed GBM (see Table 2 ). However, patients were required to have gross total resections as well as EGFRvIII positivity by immunohistochemistry in order to be eligible for this trial.

Scatter factor/hepatocyte growth factor

Scatter factor/hepatocyte growth factor (SF/HGF) and its tyrosine kinase receptor c-Met play a role in cell growth, cell motility, morphogenesis, and angiogenesis. HGF and c-Met are overexpressed in gliomas, and expression levels correlate with grade and the degree of malignancy. AMG-102 is a fully human monoclonal antibody that selectively targets SF/HGF. A phase 2 study of AMG-102 in recurrent GBM was recently completed but failed to produce any benefit. It is unclear whether the lack of activity was related to the inability of AMG-102 to pass through the blood-tumor barrier or whether inhibiting HGF alone was insufficient. A recent study suggests that the combination of EGFR inhibitors and Met inhibitors may be more effective than either agent alone in PTEN-null GBM tumors. Met TKIs such as XL184 are in clinical trial in GBM.

Platelet-derived growth factor

Platelet-derived growth factors (PDGFs) are a pleiotropic family of peptides that signal through PDGFR to stimulate cellular functions including growth, proliferation, and differentiation. PDGF and PDGFR are overexpressed in gliomas and are associated with more malignant tumors. Blocking PDGF-mediated phosphorylation causes either apoptosis or growth inhibition in glioma cell lines. PDGF-C also induces angiogenic activity via activation of PDGFR-α and -β receptors as well as upregulation of vascular endothelial growth factor (VEGF).

Imatinib mesylate, an inhibitor of PDGFR-α, PDGFR-β, Bcr-Abl, c-Fms, and c-Kit tyrosine kinases, demonstrated activity in preclinical models of glioma. However, in two multicenter phase 2 clinical trials, imatinib monotherapy was not associated with clinically useful activity in GBM, with RR of 3% to 13% and 6M-PFS of 3% to 16%. Results were also disappointing in a multicenter phase 3 trial of imatinib plus hydroxyurea (a ribonucleoside diphosphate reductase inhibitor) versus hydroxyurea monotherapy in progressive GBM. Radiographic response rates by central review and 6M-PFS were similar in both arms. One explanation for the lack of efficacy is that imatinib is a substrate for the P-glycoprotein efflux pump, which limits its passage over an intact blood-brain barrier. Two second-generation PDGFR inhibitors with improved central nervous system penetration, tandutinib and dasatinib, are in clinical trials for recurrent high-grade glioma, as is IMC-3G3, an antibody against PDGFR-α.

Epidermal growth factor

The EGFR pathway is an attractive target because it is frequently dysregulated in GBMs through overexpression, amplification, or activating mutations. EGFR dysregulation results in aberrant signaling which, in turn, is associated with increased proliferation, motility, and survival of tumor cells in vitro. The most common EGFR mutation, found in 20% to 30% of GBMs, is EGFRvIII, which causes constitutive activation and phosphorylation of the receptor without ligand binding. Several approaches exist for targeting EGFR including small-molecule tyrosine kinase inhibitors (TKIs), monoclonal antibodies (mAbs), and vaccines.

The most extensively studied TKIs in GBM are erlotinib and gefitinib. Both are oral, low molecular weight, reversible inhibitors of tyrosine kinase activity associated with EGFR. When combined with RT in newly diagnosed GBM patients, the addition of these agents did not prolong 6-month progression-free survival (6M-PFS) or overall survival (OS) in multicenter trials as compared with standard therapy. In several phase 2 clinical trials of recurrent high-grade gliomas, neither gefitinb nor erlotinib monotherapy demonstrated any significant survival benefit as compared with historical controls. Clinical activity was modest in the GBM subset, with radiographic responses (RR) ranging from 0% to 13% for gefitinib and 0% to 26% for erlotinib, and with 6M-PFS ranging from 9% to 13% for gefitinib and 0% to 26% for erlotinib. Small subgroups of patients had durable responses. Both TKIs were generally well tolerated with rash, diarrhea, and fatigue as the most common side effects. The combination of EGFR inhibitors with other therapies is discussed later in this article.

Lapatinib, which inhibits ErbB2/HER2 (human epidermal growth factor receptor 2) as well as EGFR, has also been evaluated in recurrent glioblastomas. This agent achieves therapeutic concentrations in gliomas but has limited activity.

Although subsets of GBM patients have sustained responses to reversible TKIs that target EGFR, the studies to date have been largely disappointing. Potential reasons for lack of response include poor blood-brain barrier penetration, insufficient local tumor concentrations, coactivation of multiple TKIs, redundant signaling pathways, and resistance. Irreversible EGFR inhibitors, such as BIBW 2992 and PF-00299804, could have better efficacy in GBM than gefitinib or elotinib due to increased potency and in the case of PF-00299804, better brain concentration. This newer class of EGFR inhibitors has been shown to circumvent mechanisms of response to gefitinib or erlotinib in non–small cell lung cancer cells.

Attempts to define subsets of erlotinib or gefitinib responders have also been met with limited success. The activating mutations found in EGFR in non–small cell lung cancer that increase response to EGFR inhibitors are not present in GBM. Initial molecular studies suggested that gefitinib and erlotinib were more effective in tumors with EGFRvIII mutations and intact PTEN. However, recent studies failed to validate these results.

Monoclonal antibodies (mAb) as well as vaccines that target EGFR are currently under investigation in GBM. These therapies have been well tolerated in early clinical trials. Results from a phase 1/2 study combining nimotuzumab (a humanized anti-EGFR mAb) with radiotherapy in patients were favorable ( Table 2 ). Both nimotuzumab and cetuximab, a chimeric anti-EGFR human-mouse mAb, are now being studied in combination with radiation and temozolomide as upfront GBM therapies. Preliminary results from a phase 2 clinical trial suggest that the addition of CDX-110, a peptide based EGFRvIII vaccine, to standard therapy prolongs survival in patients with newly diagnosed GBM (see Table 2 ). However, patients were required to have gross total resections as well as EGFRvIII positivity by immunohistochemistry in order to be eligible for this trial.

Scatter factor/hepatocyte growth factor

Scatter factor/hepatocyte growth factor (SF/HGF) and its tyrosine kinase receptor c-Met play a role in cell growth, cell motility, morphogenesis, and angiogenesis. HGF and c-Met are overexpressed in gliomas, and expression levels correlate with grade and the degree of malignancy. AMG-102 is a fully human monoclonal antibody that selectively targets SF/HGF. A phase 2 study of AMG-102 in recurrent GBM was recently completed but failed to produce any benefit. It is unclear whether the lack of activity was related to the inability of AMG-102 to pass through the blood-tumor barrier or whether inhibiting HGF alone was insufficient. A recent study suggests that the combination of EGFR inhibitors and Met inhibitors may be more effective than either agent alone in PTEN-null GBM tumors. Met TKIs such as XL184 are in clinical trial in GBM.

Platelet-derived growth factor

Platelet-derived growth factors (PDGFs) are a pleiotropic family of peptides that signal through PDGFR to stimulate cellular functions including growth, proliferation, and differentiation. PDGF and PDGFR are overexpressed in gliomas and are associated with more malignant tumors. Blocking PDGF-mediated phosphorylation causes either apoptosis or growth inhibition in glioma cell lines. PDGF-C also induces angiogenic activity via activation of PDGFR-α and -β receptors as well as upregulation of vascular endothelial growth factor (VEGF).

Imatinib mesylate, an inhibitor of PDGFR-α, PDGFR-β, Bcr-Abl, c-Fms, and c-Kit tyrosine kinases, demonstrated activity in preclinical models of glioma. However, in two multicenter phase 2 clinical trials, imatinib monotherapy was not associated with clinically useful activity in GBM, with RR of 3% to 13% and 6M-PFS of 3% to 16%. Results were also disappointing in a multicenter phase 3 trial of imatinib plus hydroxyurea (a ribonucleoside diphosphate reductase inhibitor) versus hydroxyurea monotherapy in progressive GBM. Radiographic response rates by central review and 6M-PFS were similar in both arms. One explanation for the lack of efficacy is that imatinib is a substrate for the P-glycoprotein efflux pump, which limits its passage over an intact blood-brain barrier. Two second-generation PDGFR inhibitors with improved central nervous system penetration, tandutinib and dasatinib, are in clinical trials for recurrent high-grade glioma, as is IMC-3G3, an antibody against PDGFR-α.

PI3K/Akt pathway inhibitors

Phosphatidylinositol 3-kinase/AKT (PI3K/Akt) is a signaling pathway important for cell growth and survival. The pathway is activated by several RTKs, including EGFR and PDGFR, and is inhibited by PTEN. Activation of the PI3K-Akt pathway is associated with poor prognosis in glioma patients and with more aggressive tumors. In vivo studies suggest that PI3K inhibitors sensitize GBM cells to apoptosis. Several PI3K and Akt inhibitors are in development or early clinical trials. XL765, an inhibitor of PI3K and mTOR, is currently in a phase 1 clinical trial in combination with temozolomide for patients with high-grade glioma. Akt inhibitors undergoing evaluation in high-grade glioma include perifosine and MK2206.

PI3K/Akt signaling leads to downstream activation of several targets including the mammalian target of rapamycin (mTOR). The mTOR inhibitor sirolimus (rapamycin) and its analogues temsirolimus, everolimus, and ridaforolimus are the most clinically advanced PI3K/Akt pathway inhibitors. Despite promising results from preclinical studies, temsirolimus monotherapy was not clinically active in recurrent GBM in two multicenter phase 2 clinical trials. Radiographic response rates were 0% and 5% while 6M-PFS rates were 2% and 8%. One possible explanation for the lack of efficacy is that rapamycin inhibition of TOR complex 1 (TORC1) disrupts the negative feedback loop on the Akt pathway, which may lead paradoxically to Akt activation. An Akt inhibitor, such as perifosine or MK2206, or a combined PI3K/mTOR inhibitor, such as XL765, may ultimately prove more favorable. Studies combining mTOR inhibitors with other targeted agents are discussed later in this article.

Ras-mitogen activated protein kinase pathway inhibitors

The mitogen-activated protein kinase (MAPK) pathway is involved in important signaling cascades that regulate normal cell proliferation, survival, and differentiation. The pathway is triggered by several RTKs, including EGFR and vascular endothelial growth factor receptor (VEGFR), and results in downstream activation of Ras, a guanine nucleotide triphosphatase. Ras then transmits signals to MAPK, PI3K, and other signaling pathways.

Several posttranslational modifications of Ras are necessary for proper functioning. One of these modifications is the addition of a farnesyl group, which enables Ras to translocate from the cytoplasm to the cell membrane. Farnesyl transferase inhibitors (FTIs), such as tipifarnib and lonafarnib, are designed to block farnesylation and ultimately prevent proper functioning of Ras. These agents demonstrated activity in human glioma cell lines, but results from clinical trials have been disappointing. Both tipifarnib and lonafarnib are now being studied in combination with TMZ for high-grade gliomas.

The Raf serine/threonine kinases are the main downstream effectors of Ras in the MAPK pathway. Inhibiting at the level of Raf may be more efficacious because it is a key element situated at the convergence of several redundant signaling pathways. Activating Raf mutations have been found in several malignancies, including GBM. Sorafenib is a potent inhibitor of Raf kinase but also inhibits proangiogenic RTKs including VEGFR-2, VGFRR-3, PDGFR-β, Flt-3, c-Kit, and fibroblast growth factor receptor 1. Several trials of sorafenib in high-grade glioma are underway.

Protein kinase C inhibitors

Protein kinase C (PKC) is a family of serine/threonine kinases that are involved in cell proliferation, differentiation, angiogenesis, and apoptosis. PKCs are activated by RTKs and G-protein–coupled receptors and help regulate several signaling pathways including PI3K/Akt and MAPK. Several PKC isoforms are overexpressed in glioma cell lines. Enzastaurin is an oral serine/threonine kinase inhibitor that targets the PKC and Akt pathways. In a multicenter phase 2 open-label trial, patients with recurrent GBM were randomized to either enzastaurin or lomustine. Unfortunately, enrollment was stopped after interim futility analysis due to a lack of difference in PFS between the 2 arms. Other studies of enzastaurin in combination with radiation and/or chemotherapeutic agents are in progress.