*8.4.3.5. Poly (ADP-ribose) polymerase (PARP)-1 abnormalities*

PARP-1 is a nuclear protein involved in the DNA damage repair processes [144]. PARP-1 activity provides an escape mechanism for cancer cells to avoid apoptosis and its overexpression may be associated with temozolomide and radiation resistance [47]. Gain of *PARP-1* is seen in DIPG tumors and provides a potentially targetable therapeutic option [47].

## **8.5. Chromosomal number abnormalities**

Copy number abnormalities (CNAs) reported in DIPG include gain in chromosomes 1q, 2q, 8q, 9q, 7p/7q and loss in chromosomes 16q, 17p, 20p, 21q, 10q and 4q [47, 48, 122, 123, 139, 145]. The CNAs may represent the initial mutations responsible for DIPG tumorigenesis as well as the treatment effect.

#### **8.6. Immune checkpoint abnormalities**

#### *8.6.1. B7-H3 abnormalities*

*8.4.3. Abnormalities of cell cycle regulation pathways*

The TP53 pathway is a complex network of genes which respond to diverse internal and external stress signals and have an impact on the normal cellular homeostasis [129]. The p53 protein is activated by stress signals transmitted as post-translational modifications leading to apoptosis [130]. In addition, the TP53 pathway produces proteins which aid directly in DNA repair processes and alter cellular environment enabling inter-cellular communication [131]. In the critical role of safeguarding the genomic integrity, it functions as a tumor suppression pathway [132]. *TP53* is the most commonly mutated gene found in a broad variety of human

*TP53* mutations are commonly found in DIPG with the reported incidence between 9 and 77% [45, 99]. They are more common in higher grade histology tumors (grades III and IV) [53]. About 50% of *TP53* wild-type grade II DIPG show presence of *PPM1D* mutations [134]. *PPM1D* is an oncogene associated with cancers like neuroblastoma [135] and lung cancer [136] which codes for wild-type p53-induced phosphatase 1D (WIP1). WIP1 is a negative regulator of *TP53* as it inactivates p53 and promotes termination of stress-induced responses. So *PPM1D* mutations have the same functional significance as *TP53* mutations [137]. *PPM1D* and *TP53* mutations are mutually

exclusive and may ultimately lead to dysregulated homeostasis and tumorigenesis [134].

*TP53* mutations more commonly co-segregate with H3.3 K27 M than H3.1 K27 M [112] and

The abnormalities involving these regulators observed in DIPG include cyclin-dependent kinase inhibitor 2A or 2B (*CDKN2A* or *CDKN2B*) deletions [48, 122] and *CDK4*, *CDK6* or cyclin

Aurora kinase family include three highly homologous serine/threonine kinases required during mitosis and which are linked to many cancers [140]. AURKB forms the catalytic component of the chromosomal passenger complex (CPC) which plays a critical role during mito-

sis [141]. Almost 70% of DIPG have demonstrated overexpression of *AURKB* [142].

/S transition of the cell cycle [138].

checkpoint. DIPG cells, unlike normal cells, have aber-

checkpoint, including *TP53*, *MDM2*, *CDKN2A*, and *ATM*

*8.4.3.1. TP53 pathway*

50 Brain Tumors - An Update

cancers [129, 133].

*8.4.3.1.1. TP53 mutations and DIPG*

*8.4.3.1.2. TP53 co-mutations in DIPG*

*8.4.3.2. The RB pathway*

*8.4.3.3. Aurora kinase pathway*

*8.4.3.4. WEE1 kinase pathway*

WEE1 kinase is an important part of G2

rations in genes regulating the G1

frequently occur in the setting of *PDGFRA* aberrations [124].

Cyclins and cyclin-dependent kinases (CDKs) control the G1

D1 (*CCND1*), *CCND2,* and *CCND3* amplifications [48, 122, 139].

B7-H3 or CD276, a member of the B7-CD28 family, is a type I transmembrane glycoprotein [146]. Many malignant neuroectodermal tumors including adult HGG over-express B7-H3. B7-H3 was found to be overexpressed in a small panel of DIPG samples obtained at autopsy [147]. A monoclonal antibody 8H9 recognizes it and binds specifically to the tumor cells [148, 149] enabling therapeutic cell selectivity. B7H3 was targeted safely in the salvage therapy of stage IV neuroblastoma using intrathecal 131I-8H9 [150].

#### **8.7. Tumor microenvironment abnormalities**

Therapies targeted at intrinsic cellular pathways have yielded poor results in DIPG. Tumor microenvironment plays a vital role in tumorigenesis and progression, so studies have looked into investigating microenvironment alteration for better results.

#### *8.7.1. Neuroligin-3 (NLGN3) role*

Neuroligin-3 (NLGN3) is a synaptic adhesion molecule which is cleaved from neurons and oligodendrocyte precursor cells via the ADAM10 sheddase and released into the tumor microenvironment. This important neuronal activity promotes many types of brain cancers including DIPG, pediatric and adult HGG and anaplastic oligodendroglioma. NLGN3 release activates oncogenic pathways like focal adhesion kinase activation resulting in the downstream PI3K-mTOR pathway induction. This in turn causes upregulation of several synapse-related genes resulting in the proliferation of glioma cells [151]. HGG glioma growth in xenograft models was blocked by ADAM10 inhibitors by preventing NLGN3 release into the tumor microenvironment. Similarly, patient-derived orthotopic xenografts fail to grow in *Nlgn3* knockout mice [152].

**10.2. Epigenetic modifying agents**

**10.3. Targeting cell cycle regulation**

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

**10.4. Immunotherapy**

target.

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

Diffuse Intrinsic Pontine Glioma

53

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

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].

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

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

a phase I trial of single agent Panobinostat in patients with DIPG [171].

#### *8.7.2. Pleiotrophin (PTN) role*

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].
