**10. Novel therapeutic approaches**

## **10.1. Molecularly targeted agents**

#### *10.1.1. RTK-RAS-PI3K pathway*

Given DIPG tumors show aberrant activation of growth factor receptor-mediated signal transduction pathways, using drugs targeting these pathways is a rational approach. *In-vitro* studies confirmed the efficacy of tyrosine kinase inhibitors (TKI) like dasatinib in reducing tumor proliferation and inhibition of *PDGFRA* activity [154]. Phase I studies of PDGFR pathway inhibition by imatinib [155] and dasatinib [156], VEGFR2 inhibition by vandetinib [156, 157], EGFR inhibition by gefitinib [158] and erlotinib [49] revealed the safety of using these drugs in children and provided the doses for phase II studies. Temsirolimus is an mTOR inhibitor [159] and its combination with perifosine, an Akt inhibitor, was shown to be safe and feasible in a phase I study in children with recurrent/refractory solid tumors including DIPG [160]. This study tested the hypothesis of dual targeting of PI3K-Akt–mTOR pathway. Phase II studies of multi-tyrosine kinase inhibitor sunitinib [161] and farnesyl transferase inhibitor tipifarnib, which inhibits farnesylation of Ras, [162] did not show activity. BIOMEDE, a phase II study, is stratifying patients based on overexpression of EGFR and/or loss of PTEN following diagnostic biopsy. The patients are randomized or assigned to different treatment arms with erlotinib, everolimus and dasatinib [163].

#### **10.2. Epigenetic modifying agents**

the tumor microenvironment. Similarly, patient-derived orthotopic xenografts fail to grow in

PTN is a growth factor secreted by neural precursor cells in the lateral ventricle subventricular zone (SVZ). It has an important role in normal neurodevelopment, plasticity and regeneration. PTN acts as a glioma chemoattractant. Through autocrine/paracrine actions, it activates Rho/Rho kinase (ROCK) signaling pathway enabling migration of DIPG cells to the SVZ [153].

Proteomic, methylation and mRNA analyses have identified interesting subgroups of DIPG. PDGFRA amplification is seen in association with H3.3 mutation and ACVR1 mutation is

Mesenchymal transition with stem cell-like phenotype and oligodendroglial differentiation

Given DIPG tumors show aberrant activation of growth factor receptor-mediated signal transduction pathways, using drugs targeting these pathways is a rational approach. *In-vitro* studies confirmed the efficacy of tyrosine kinase inhibitors (TKI) like dasatinib in reducing tumor proliferation and inhibition of *PDGFRA* activity [154]. Phase I studies of PDGFR pathway inhibition by imatinib [155] and dasatinib [156], VEGFR2 inhibition by vandetinib [156, 157], EGFR inhibition by gefitinib [158] and erlotinib [49] revealed the safety of using these drugs in children and provided the doses for phase II studies. Temsirolimus is an mTOR inhibitor [159] and its combination with perifosine, an Akt inhibitor, was shown to be safe and feasible in a phase I study in children with recurrent/refractory solid tumors including DIPG [160]. This study tested the hypothesis of dual targeting of PI3K-Akt–mTOR pathway. Phase II studies of multi-tyrosine kinase inhibitor sunitinib [161] and farnesyl transferase inhibitor tipifarnib, which inhibits farnesylation of Ras, [162] did not show activity. BIOMEDE, a phase II study, is stratifying patients based on overexpression of EGFR and/or loss of PTEN following diagnostic biopsy. The patients are randomized or assigned to different treatment arms

*Nlgn3* knockout mice [152].

52 Brain Tumors - An Update

*8.7.2. Pleiotrophin (PTN) role*

**9. DIPG subgroups**

mainly seen in H3.1 mutant tumors [87, 115].

and PDGFRA amplification/mutation.

**10. Novel therapeutic approaches**

with erlotinib, everolimus and dasatinib [163].

**10.1. Molecularly targeted agents**

*10.1.1. RTK-RAS-PI3K pathway*

Upregulation of N-Myc and Hh signaling pathways [92].

Upregulation of N-Myc (H3 wild-type), silent genome and H3K27 M [115].

Histone deacetylases (HDAC) regulate the histone acetylation in nucleosomes, which mediates changes in chromatin conformation, leading to gene expression regulation [164]. HDAC gene mutations, downregulation and altered expression are linked to tumorigenesis. Histone deacetylase (HDAC) inhibitors modify histone activity to increase expression of previously silenced genes thereby leading to cell death [165]. Panobinostat is a multi-HDAC inhibitor which increases global H3 acetylation and H3K27 M methylation and reduces oncogene expression. Panobinostat-induced polyacetylation of the H3 N-terminal tail has been shown to reverse PRC2 inhibition caused by K27 M and rescue the H3K27 hypomethylation phenotype [166]. Panobinostat has shown promising results in *in vitro* [164] and some *in vivo* [167] studies although a narrow therapeutic index causing dose limiting toxicities at the required antitumor concentrations being the likely reason for lack of efficacy [164]. Its efficacy was further enhanced by histone demethylase inhibitor GSK-J4 which increase H3K27me3 [168]. DIPG patients showed no objective responses to single agent vorinostat in a phase I clinical trial [169]. Vorinostat is currently being investigated in combination with temsirolimus and RT in DIPG in a phase I trial [170]. Pediatric Brain Tumor Consortium (PBTC) is undertaking a phase I trial of single agent Panobinostat in patients with DIPG [171].

#### **10.3. Targeting cell cycle regulation**

DIPG tumors contain cell cycle regulatory gene abnormalities like amplification of D-type cyclins and CDK4/6 or loss of Ink4a-ARF resulting in cellular proliferation [172]. Targeting the cyclin/CDK/RB pathway was investigated in a preclinical trial. PD-0332991 (PD), a CDK4/6 inhibitor, was used in a genetically engineered, PDGF-B overexpressed, Ink4a-ARF and p53 deficient brainstem glioma mouse model. PD induced cell cycle arrest both *in vitro* and *in vivo*. The survival of mice treated with PD alone or in combination with RT was significantly more than untreated or mice treated with only RT [172]. Inhibition WEE1 kinase, which is overexpressed in DIPG, was shown to increase the RT response in vitro and in vivo [73].

#### **10.4. Immunotherapy**

Interferon, IL-12 and anti-glioma monoclonal antibody have shown efficacy in mouse models of non-brainstem malignancies [173]. Recently, subcutaneous vaccinations with glioma-associated antigen epitope peptides (EphA2, IL13-Rα2 and survivin) were investigated in children with DIPG. The vaccine was tolerated well and the results were encouraging for future trials [174]. IL13-Rα2 is of particular significance as it is highly and differentially expressed in DIPG [175, 176] compared to the normal brain tissue which makes it a suitable immunotherapy target.

#### **10.5. Human neural and mesenchymal stem cells**

Neural stem cells (NSC) have been utilized as the vehicle for therapeutic agents in gene therapy for experimental malignant brain tumors [177, 178]. NSC have robust tumor tropism and this property can be used to deliver therapeutic agents to the inoperable DIPG tumors. NSC are difficult to harvest and use due to their inherent immunogenicity [179]. Mesenchymal stem cells (MSC) have been found to be a suitable alternative as they have all the properties of NSC and are more practical to use [180]. In a rat model, brainstem glioma directed migratory capacities of NSC and MSC from different sources was found to be similar *in vitro* and *in vivo*. This study showed promise as 20–30% of stem cells migrated from the site of injection in the right forebrain to the brainstem glioma cells. In addition, the group treated with genetically engineered NSC encoding cytosine deaminase (CD) suicide gene and IFNβ proinflammatory cytokine and systemic 5-fluorouracil resulted in 59% reduction in the tumor size [181].

*10.6.2. Nanoparticle delivery*

*10.6.3. Cell-mediated delivery*

including DIPG [204].

**11. Conclusion**

**Author details**

**References**

*10.6.4. BBB disruption techniques*

systemically delivered drugs [28].

and may help in identifying new targeted therapies.

\*Address all correspondence to: santosh.valvi@health.wa.gov.au

Santosh Valvi\* and Nicholas G. Gottardo

Perth Children's Hospital, Perth, Australia

Lippincott Williams & Wilkins; 2011. p. 2

the volume of distribution [28, 201].

Another alternative drug delivery technique involves encapsulation of cationic substances into capsules termed micro- and nanoparticles. This reduces their tissue affinity and increases

Diffuse Intrinsic Pontine Glioma

55

http://dx.doi.org/10.5772/intechopen.78578

Bacterial cell-derived vehicles to transport chemotherapy agents across the BBB was utilized with bi-specific antibodies recognizing cell wall and EGFR moieties [202]. Similar EGFR-targeted vehicles loaded with doxorubicin resulted in significant tumor regression in canine brain cancer models [203]. ECREST (A Study of Intravenous EEDVsMit in Children With Recurrent/Refractory Solid or CNS Tumors Expressing EGFR) is a phase I study using mitoxantrone loaded nanocells in EGFR positive relapsed/refractory solid and CNS tumors

BBB can be temporarily disrupted by ultrasound methods. This results in enhanced uptake of

DIPG remains the main cause of death in pediatric patients with brain tumors. Despite many clinical trials, minimal improvement has been achieved as compared with RT. Most of the clinical trials in the past did not have sound biological bases due to a lack of biopsy specimens. In the last few years, international collaborative research has helped to identify new molecular aberrations in DIPG. These have provided insight into the pathogenesis of DIPG

[1] Scheurer ME, Bondy ML, Gurney JG. Epidemiology of childhood Cancer. In: Pizzo PA, Poplack DG, editors. Principles and Practice of Pediatric Oncology. 6th ed. Philadelphia:

#### **10.6. Delivery methods to overcome blood-brain barrier (BBB)**

The vast majority of brain tumors are characterized by the loss of BBB integrity due to the disordered and highly permeable tumor neovasculature [182]. In DIPG, however, the BBB integrity is normal and the drug permeability is reduced as the tumor makes use of the existing brain vasculature [183]. Traditional cytotoxic drugs have shown good efficacy against DIPG cells *in vitro* but they likely fail to penetrate the BBB which may explain the contrasting results with clinical trials [28]. To overcome the disappointing results associated with systemic therapy, methods utilizing localized and targeted drug delivery have been investigated.

#### *10.6.1. Convection-enhanced delivery (CED)*

CED is a relatively novel delivery modality which utilizes the properties of bulk flow to achieve homogeneously distributed infusions into the brain parenchyma [184]. It is a neurosurgical technique in which one to several catheters are stereotactically placed within or around the tumor mass and drug is delivered locally, bypassing the BBB and reducing systemic toxicity [185, 186]. The local concentrations achieved are significantly higher than the systemic administration and the drug distribution occurs along the patterns of the glioma cell invasion along the white matter tracts [187]. Many *in vivo* feasibility studies have confirmed that drugs can be safely delivered into the rodent brainstem [188]. CED of carmustine [74, 189], carboplatin [190, 191], temozolomide [192], small-molecule kinase inhibitors [193], cisplatin [194] and gemcitabine [190, 195] have been evaluated for safety and distribution parameters. Topotecan CED in two pediatric patients with DIPG showed initial reduction in tumor size [196]. MRI guided and robotically placed catheters were used for CED of carboplatin in DIPG patients. Three out of eight patients survived beyond 15 months and the procedure was tolerated well [197, 198]. Administration of 124I-8H9, a radioactively labeled antibody and a chimeric toxin with B7-H3 specificity, is currently being explored in a phase I clinical trial for DIPG patients [199]. A patient with DIPG received CED infusion of the recombinant cytotoxin IL13-PE38QQR with GD-diethylenetriamine but the tumor progresses within a few weeks [200]. IL13-PE38QQR is a recombinant *Pseudomonas aeruginosa* toxin and some early phase clinical trials investigating its CED efficacy are being conducted or recently completed in patients with DIPG and HGG [185]. These studies have established the feasibility of CED. However, more data is required to prove safety and efficacy and CED models are undergoing rigorous investigations into the physical properties of the catheters and infusion rates [173]. CED targets the primary tumor only and not the metastatic sites. In DIPG, local and distant metastases in the brain or spine are seen in 13–17% patients. Therefore, in situations where disseminated disease is present, CED therapy alone, directed toward the primary tumor site, will be inadequate and should be complemented with therapies for the secondaries [28, 32].

#### *10.6.2. Nanoparticle delivery*

stem cells (MSC) have been found to be a suitable alternative as they have all the properties of NSC and are more practical to use [180]. In a rat model, brainstem glioma directed migratory capacities of NSC and MSC from different sources was found to be similar *in vitro* and *in vivo*. This study showed promise as 20–30% of stem cells migrated from the site of injection in the right forebrain to the brainstem glioma cells. In addition, the group treated with genetically engineered NSC encoding cytosine deaminase (CD) suicide gene and IFNβ proinflammatory cytokine and systemic 5-fluorouracil resulted in 59% reduction in the tumor size [181].

The vast majority of brain tumors are characterized by the loss of BBB integrity due to the disordered and highly permeable tumor neovasculature [182]. In DIPG, however, the BBB integrity is normal and the drug permeability is reduced as the tumor makes use of the existing brain vasculature [183]. Traditional cytotoxic drugs have shown good efficacy against DIPG cells *in vitro* but they likely fail to penetrate the BBB which may explain the contrasting results with clinical trials [28]. To overcome the disappointing results associated with systemic therapy, methods utilizing localized and targeted drug delivery have been investigated.

CED is a relatively novel delivery modality which utilizes the properties of bulk flow to achieve homogeneously distributed infusions into the brain parenchyma [184]. It is a neurosurgical technique in which one to several catheters are stereotactically placed within or around the tumor mass and drug is delivered locally, bypassing the BBB and reducing systemic toxicity [185, 186]. The local concentrations achieved are significantly higher than the systemic administration and the drug distribution occurs along the patterns of the glioma cell invasion along the white matter tracts [187]. Many *in vivo* feasibility studies have confirmed that drugs can be safely delivered into the rodent brainstem [188]. CED of carmustine [74, 189], carboplatin [190, 191], temozolomide [192], small-molecule kinase inhibitors [193], cisplatin [194] and gemcitabine [190, 195] have been evaluated for safety and distribution parameters. Topotecan CED in two pediatric patients with DIPG showed initial reduction in tumor size [196]. MRI guided and robotically placed catheters were used for CED of carboplatin in DIPG patients. Three out of eight patients survived beyond 15 months and the procedure was tolerated well [197, 198]. Administration of 124I-8H9, a radioactively labeled antibody and a chimeric toxin with B7-H3 specificity, is currently being explored in a phase I clinical trial for DIPG patients [199]. A patient with DIPG received CED infusion of the recombinant cytotoxin IL13-PE38QQR with GD-diethylenetriamine but the tumor progresses within a few weeks [200]. IL13-PE38QQR is a recombinant *Pseudomonas aeruginosa* toxin and some early phase clinical trials investigating its CED efficacy are being conducted or recently completed in patients with DIPG and HGG [185]. These studies have established the feasibility of CED. However, more data is required to prove safety and efficacy and CED models are undergoing rigorous investigations into the physical properties of the catheters and infusion rates [173]. CED targets the primary tumor only and not the metastatic sites. In DIPG, local and distant metastases in the brain or spine are seen in 13–17% patients. Therefore, in situations where disseminated disease is present, CED therapy alone, directed toward the primary tumor site, will be inadequate and should be

**10.6. Delivery methods to overcome blood-brain barrier (BBB)**

*10.6.1. Convection-enhanced delivery (CED)*

54 Brain Tumors - An Update

complemented with therapies for the secondaries [28, 32].

Another alternative drug delivery technique involves encapsulation of cationic substances into capsules termed micro- and nanoparticles. This reduces their tissue affinity and increases the volume of distribution [28, 201].

#### *10.6.3. Cell-mediated delivery*

Bacterial cell-derived vehicles to transport chemotherapy agents across the BBB was utilized with bi-specific antibodies recognizing cell wall and EGFR moieties [202]. Similar EGFR-targeted vehicles loaded with doxorubicin resulted in significant tumor regression in canine brain cancer models [203]. ECREST (A Study of Intravenous EEDVsMit in Children With Recurrent/Refractory Solid or CNS Tumors Expressing EGFR) is a phase I study using mitoxantrone loaded nanocells in EGFR positive relapsed/refractory solid and CNS tumors including DIPG [204].

#### *10.6.4. BBB disruption techniques*

BBB can be temporarily disrupted by ultrasound methods. This results in enhanced uptake of systemically delivered drugs [28].
