A Trial of Poly-ICLC in the Management of Recurrent Pediatric Low Grade Gliomas

Background/Rationale The incidence of primary pediatric brain tumors in the United States is about 1500 per year. Brain tumors are the most common solid tumor diagnosed in childhood and thus account for significant childhood mortality in the United States. Low-grade astrocytomas and gliomas are the most common type of brain tumor of childhood (36% of childhood brain tumors). These tumors encompass a heterogeneous assortment of histological subtypes including: fibrillary, protoplasmic, gemistocytic, and mixed variants. Pilocytic astrocytomas, pleomorphic xanthoastrocytomas and subependymal giant cell astrocytomas are also included. Furthermore, in young children there are some unique rare entities that behave like low-grade tumors, including infantile desmoplastic gangliogliomas, and desmoplastic astrocytomas. Although children with low-grade astrocytomas often survive many years after conventional treatment with surgery and sometimes radiotherapy, some children will not fare as well. These tumors constitute a heterogeneous group because of differing locations within the brain and varying biological behavior of different subtypes. For those where total excision is possible, such as cerebellar astrocytomas, prognosis is excellent with over 90% ten-year survival rates with surgical excision alone. In contrast, survival rates in children with cerebral or diencephalic tumors are 40-70% at five years with irradiation, but decline to 11-50% at 10 years. Some tumors however may be unresectable/partially resectable, and radiation can have undesirable side effects in young children. While the most significant intellectual deficits occur in young children less than 5 years treated with cranial irradiation, the deficits recognized even in young adults warrant extending the age to 10 years for avoiding radiation. Chemotherapy regimens are used for high-risk patients (progressive tumor, residual tumor) as a means to avoid or delay radiation in young patients, but side effects of chemotherapy are frequently reported.

Newer forms of effective treatment that will have lesser side effects are much needed in childhood brain tumors especially low-grade gliomas. We propose to study the efficacy and toxicity of poly-ICLC, a biological response modifier in children with low-grade gliomas.

PROTEOMICS

Current diagnostic and therapeutic monitoring of brain tumor patients are significantly hindered due to limited understanding of brain tumor biology and response to therapy. The majority of central nervous system (CNS) tumors cannot be identified or followed by expression of serum or Cerebrospinal fluid (CSF) markers. However, if available, such markers would be highly desirable and could be used to:

• Detect minimal residual disease
• Predict response to specific targeted therapies
• Predict or anticipate tumor progression
• Distinguish tumor recurrence from post surgical changes or post-radiation changes on neuro-imaging
• Augment current histopathologic classification systems
• Improve current clinical and pathological treatment stratification schemata
• Assess efficacy of and tumor response to specific biologic targeted therapies that may not impact tumor size as a primary tumor endpoint (e.g., small molecule inhibitors or anti-angiogenic strategies) While such markers would be useful to prognosticate, monitor and treat all CNS tumors, its use in glial tumors including recurrent low grade astrocytomas is critical since these tumors are often biopsied at presentation, but not at recurrence. Often these tumors are not amenable complete resection or biopsy due to the eloquence of brain tissue they infiltrate (e.g., optic pathway, brainstem or hypothalamic gliomas), or the blood vessels that they encase.

CNS biologic material in CSF Glial tumors tend to disseminate locally along white matter tracts rather than through sub-arachnoid seeding. Dissemination of low grade gliomas along the sub-arachnoid space has been reported in children with low grade gliomas. Even focal tumors are frequently adjacent to CSF pathways (e.g., intrapeduncular fossa, third and fourth ventricles) resulting in direct contact between tumor tissue and spinal fluid. Yet examination of CSF cytology for these tumors is not standard. Given limitations of identifying tumor cells in the CSF, methodologies that could improve our understanding of CNS tumors of all types are needed. This would provide a significant improvement in currently available knowledge about the biology of these tumors, and could elucidate potential therapeutic avenues.

Proteomics, a relatively new area of research whereby total protein complement of a tissue compartment is analyzed, has successfully been used to identify novel biomarkers in solid tumors. Because proteins are effectors of all cellular functions, their measurement should represent the most direct means of cellular characterization and hence tumor biology. Because cells and their environment exist in an integrated state, it has been possible to interrogate the proteins of extra-cellular compartments to assess the presence and impact of tumor cells. This has been done primarily using serum or plasma to establish a method of screening for the presence of low stage tumors. An analogous extra-cellular compartment for use in brain tumors would be cerebrospinal fluid (CSF). It circulates throughout the CNS and exchanges proteins with the extra-cellular fluid of the brain and spinal cord.

CSF is continuously created and reabsorbed, providing a real time steady state proteome. Unlike serum, which contains a highly complex protein mixture ranging from very low abundance proteins in the 10-30 pg/mL range to very abundant proteins in the 35-55 mg/mL range, CSF contains a less complex protein mixture.

Therefore, the CSF is more likely to contain higher relative concentrations of tumor-specific proteins (higher signal to noise ratio) than serum. Taken together this makes CSF and attractive alternative to serum for detection of brain tumor related biomarkers. Unlike leukemia and many solid tumors outside the CNS, where serial biopsies are readily performed, tumors of the CNS are not easily accessible other than at the time of initial or repeat resection or biopsy. While studies on these samples provide important findings regarding tumor biology, serial analyses during treatment are not reasonable. By contrast, the CSF of tumor patients can be more readily sampled in most pediatric patients. With the development of proteomic technology, investigation of tumor related signals at the time of diagnosis through treatment, and then in remission and/or at the time of recurrence or progression is possible.

While CSF for seeding tumors is readily available and routinely obtained for cytology, the systematic evaluation of the proteins within these samples could be of considerable scientific importance. In addition to identifying potential makers of disease or response to therapy, the glycosylation and phosphorylation status of many proteins can also be evaluated. Studies in tumor tissue show that such information reveals activity of different enzymes that correlate with treatment response or progression of leptomeningeal metastases.

Proteomics CSF proteomics has been applied to many neurological disorders including Alzheimer's disease, amyotrophic lateral sclerosis, multiple sclerosis, acute brain injury and Creutzfeldt-Jakob disease.

Reports of its use in neuro-oncology are limited, but demonstrate the potential of this technology to effectively identify tumor biomarkers. One study used two dimensional polyacrylamide (2-D) gel electrophoresis to measure the relative quantities of two pre-selected markers, proto-oncogene protein (N-Myc) and Intracranial atherosclerotic disease (l-CaD), in the CSF of brain tumor patients. CSF proteomics using 2-D gel electrophoresis in combination with mass spectroscopy and cleavable isotope Coded Affinity Tag (cICAT) was used to evaluate 60 samples of CSF and tumor cyst fluid taken from adults with brain tumors and non-neoplastic controls. These techniques were used to find a panel of proteins differentially expressed in lower vs. higher-grade gliomas. Findings were confirmed using Western Blot analysis probing for eight selected proteins based on implied role in gliomagenesis and availability of antibodies. This report, which has been accepted for publication pending revisions, identified 21 potential CSF biomarkers for astrocytoma.

As mentioned above, there is evidence that gliomas disseminate through the subarachnoid space. Currently there are several consortia actively studying protein expression in the spinal fluid of children with malignant glial and embryonal tumors. Proteins interrogated in these protocols include those involved with angiogenesis and neovascularity, those involved in tumor growth and migration (Secreted protein with acidic and cysteine rich domains (SPARC), attractin). There is no consortium actively collecting spinal fluid sample in children with lower grade tumors. A secondary goal of this study is to examine these proteins in the CSF of children with low grade gliomas who have tumor progression. Comparison of CSF protein expression of in high grade and low grade tumors is likely to help identify biological markers specific for tumor progression, or for tumor pathology.