The Cooperative Group Bulletin Board Targeted Delivery in Primary and Metastatic Brain Tumors: Summary Report of the Seventh Annual Meeting of the Blood-Brain Barrier Disruption Consortium

The November 2000 NIH report of the Brain Tumor Progress Review Group identified delivering and targeting therapeutic agents as a priority in the treatment of malignant brain tumors. For this reason, the seventh annual Blood-Brain Barrier Disruption Consortium meeting, partially funded by an NIH R13 Grant, focused on recent advances in targeted delivery to the central nervous system, clinical trials for primary and metastatic brain tumors using enhanced chemotherapy delivery, and strategies to lessen the toxicities associated with dose intensive treatments, using thiols. Introduction Despite important advances in molecular characterization, malignant brain tumors remain notoriously difficult to treat with the standard tools of surgery, radiation, and chemotherapy, and outcomes remain discouraging. Recent studies using targeted delivery show promising results. Discussion topics at this meeting, which was held March 8–10, 2001 in Gleneden Beach, Oregon, included advances in immunotherapy and gene therapy in high-grade glioma, the role of chemotherapy in CNS metastases, delivery strategies, such as convection-enhanced delivery of immunotoxins and enhanced chemotherapy administered in conjunction with osmotic opening of the blood-brain barrier, the role of high-dose chemotherapy with autologous stem cell support in malignant brain tumors, and new molecular targets in childhood cancers. Advances in Malignant Glioma The Role of Surgery. Despite major technical advances in neurosurgery such as microsurgery and stereotaxy, malignant gliomas remain very difficult to treat, and the benefit of aggressive surgery versus biopsy on survival is unknown (1–3). Arguments in favor of radical resection include cytoreduction and the importance of decreasing tumor burden, relieving increased intracranial pressure, reversing progressive neurological deficits, including potentially eliminating seizures, obtaining large amounts of tissue for study and for therapies, such as vaccination strategies, and the limited efficacy of radiation on long-term survival. Arguments against radical resection include the infiltrative nature and multifocal or multilobular presentation of most gliomas, and the potential for new neurological deficits from technical difficulties encountered when resecting tumors involving eloquent areas of the brain. Whether or not extensive resection is planned and in view of its limited efficacy, malignant gliomas require adjunctive therapy, and it is clear that this therapy should be multimodal. Molecular Classification and Neuropathology. In view of the emphasis placed on modern genetic techniques in the detection and diagnosis of gliomas by the Brain Tumor Progress Review Group (4), molecular profiles of 160 snap-frozen oligodendrogliomas and astrocytomas via loss of heterozygosity analysis, multiplex polymerase chain reaction, or Southern blotting for oncogene amplification, single-strand conformational polymorphisms, and/or sequencing to detect gene mutations (mut), and comparative genomic hybridization were performed Received 7/11/01; revised 2/12/02; accepted 3/2/02. The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked advertisement in accordance with 18 U.S.C. Section 1734 solely to indicate this fact. 1 Supported by NIH Grant 1R13 CA 86959-02 through the National Cancer Institute, the National Institute of Neurological Disorders and Stroke, and the National Institute of Deafness and Other Communication Disorders. 2 To whom requests for reprints should be addressed, at Oregon Health and Science University, Department of Neurology, 3181 S.W. Sam Jackson Park Road-L603, Portland, OR 97201-3098. Phone: (503) 4945626; Fax: (503) 494-5627; E-mail: [email protected]. 3 The abbreviations used are: CNS, central nervous system; BR96-Dox, BR96-Doxorubicin; OHSU, Oregon Health and Science University; 4-HPR, fenretinide; i.a. intra-arterial. 1702 Vol. 8, 1702–1709, June 2002 Clinical Cancer Research Research. on April 20, 2017. © 2002 American Association for Cancer clincancerres.aacrjournals.org Downloaded from (Table 1). Loss of heterozygosity for 1p and 19q was seen in 34 of 45 oligodendrogliomas and 6 of 100 astrocytomas; however, comparative genomic hybridization revealed complete loss of 1p ( 1p) and 19q ( 19q) confined to the 34 of 45 oligodendrogliomas, whereas astrocytomas had either partial loss of 1p (n 4) and/or 19q (n 3) or total loss of 1p (n 2) and/or 19q (n 3). Forty-three astrocytomas and 9 oligodendrogliomas (as classified by light microscopy) revealed loss of heterozygosity for 17p and/or p53mut ( 17p/p53mut). Among these tumors none was found to have 1p/ 19q by comparative genomic hybridization concomitant with 17p/p53 mut. These data suggest comparative genomic hybridization provides strict, reproducible criteria for the identification of oligodendroglioma. The number of accumulated genetic defects in the astrocytomas revealed that patients with tumors with two or fewer abnormalities had a significantly longer survival than patients with tumors with three or more genetic defects. The survival of patients whose tumors exhibited the complete 1p/ 19q defects was significantly better than patients with other gliomas, including the group with low-grade astrocytomas with partial 1p/ 19q defects, regardless of therapeutic intervention. Although previous studies have suggested a therapeutic benefit to the presence of the 1p defect in gliomas (5, 6), these results may also be interpreted as demonstrating the inherent chemosensitivity of true oligodendrogliomas (7, 8). This interpretation is further supported by the insignificant survival effects of the 1p defect in tumors either microscopically diagnosed as glioblastoma or found to have three or more molecular defects. These findings lend support to the concept that molecular profiles will be useful in increasing specificity of categorization and grading gliomas. Immunotherapy. Recent discoveries in adoptive cellular immunotherapy, monoclonal antibodies to deliver local irradiation, cytokine gene therapy, dendritic cell vaccines, and targeted toxins offer much promise (9, 10). Anticancer vaccines stimulate the host immune system to recognize cancer as foreign. Because brain tumors do not elicit an effective immune response, brain tumor vaccines require that a tumor sample be exposed to an agent, usually a cytokine, such as granulocytemacrophage colony-stimulating factor, that allows antigenpresenting cells to stimulate an immune response. Most preliminary work in brain tumor immunotherapy has been performed in mice and rat animal models (11, 12). The most potent antigen-presenting cells are dendritic cells, which are capable of enticing cytotoxic T cells to attack tumor cells. Early work in a syngeneic rat flank tumor model of 9L gliosarcoma demonstrated complete tumor remission in treated animals that were vaccinated with irradiated tumor cells and a continuous infusion of granulocyte-macrophage colony-stimulating factor in the flank contralateral to the tumor (11). These animals were also resistant to rechallenge with peripheral or intracerebral gliosarcoma. In animals bearing an intracerebral 9L inoculation, peripheral challenge with irradiated 9L cells and granulocyte-macrophage colony-stimulating factor resulted in remission of the intracranial tumor in 40% of animals. In an intracerebral mouse GL261 glioma model, dendritic cell immunotherapy resulted in a cure rate of 43% (12). All treated animals that were rechallenged with intracranial GL261 glioma survived. Under the proper conditions, T cells can cross the bloodbrain barrier to attack brain tumors. Once dendritic cells have been exposed to tumor antigen in the presence of a cytokine, they can be reintroduced into an area of the body rich in lymph nodes, to generate a proliferative T-cell response to CNS tumor. Another method of stimulating an immune response is to inoculate the patient with nonviable irradiated tumor cells, which serve as a source of tumor antigen, in the presence of a cytokine. This method activates peripheral T cells to migrate into the CNS and attack the brain tumor. Numerous questions remain unanswered (13): (a) How often is vaccination necessary? (b) What is the best cytokine for T-cell activation? and (c) How long should vaccination continue? If the immune system has sufficient memory, a single inoculation should be all that is required. However, most protocols require multiple treatments because of the difficulty achieving a durable response. Gene Therapy. Gene transfer or “gene therapy” offers a potentially promising treatment strategy. Tumor-selective adenoviral vectors have been developed by using an E2F-responsive promoter to express different prodrug-activating genes (14), e.g., vectors expressing the thymidine kinase gene have been successfully used to eradicate gliomas both in vitro and in vivo after ganciclovir treatment with significantly less normal tissue toxicity, compared with standard thymidine kinase-expressing viral vectors. Similarly, gene transfer has been used to target tumorassociated angiogenesis (15). Both retroviral and adenoviral vectors carrying antiangiogenic genes (e.g., platelet factor 4 and angiostatin) have been constructed. These vectors mediate an antitumor effect in experimental glioma models. Finally, human and mouse endothelial progenitor cells were used as angiogen4 R. E. McLendon, A. Rasheed, R. Wiltshire, J. Herndon, A. Friedman, H. Friedman, D. Bigner, and S. Bigner, unpublished data, 2001. Table 1 Molecular profiles of oligodendrogliomas (n 45) and astrocytomas (n 100) Grade No. of tumors % Oligodendroglioma 1p/ 19q CGH II 16/23 70% III 18/22 82% 9p CGH II 2/23 9% III 11/22 50% P53 mutations II 3/23 13% III 3/22 14% Astrocytoma 17pLOH/p53 mutations I & II 7/19 37% III 16/27 59% IV 20/54 37% 10LOH I & II 0/19 0% III 14/27 52% IV 31/54 57% 7CGH/ 10CGH I & II 0/19 0% III 7/27 26% IV 32/54 59% Oncogenic amplification I & II 0/19 0% III 2/27 7% IV 20/54 37% a CGH, comparative genomic hybridization; LOH, loss of heterozygosity. 1703 Clinical Cancer Research Research. on April 20, 2017. © 2002 American Association for Cancer clincancerres.aacrjournals.org Downloaded from esis-selective, gene-targeting vectors. Endothelial progenitor cells were isolated, expanded, and genetically engineered ex vivo to express the galactosidase or thymidine kinase genes using retrovirus-mediated gene transfer. Genetically labeled endothelial progenitor cells were transplanted into wild-type and sublethally irradiated mice and found to migrate and incorporate into the angiogenic vasculature of growing tumors while maintaining transgene expression. Ganciclovir treatment resulted in significant tumor necrosis in animals previously administered thymidine kinase-expressing endothelial progenitor cells. These results demonstrate the potential of using genetically modified endothelial progenitor cells as angiogenesis-selective, genetargeting vectors and the potential of this approach to mediate nontoxic and systemic antitumor responses. Metastases to the CNS Is There a Role for Chemotherapy in CNS Metastases? Although CNS metastases occur 10-fold more frequently than primary brain tumors, available treatment options are limited (16), and chemotherapy in this setting has been poorly studied. The Brain Tumor Progress Review Group noted a dearth of NIH-funded grants to investigate CNS metastases. Possible roles for chemotherapy include: (a) treatment of established brain metastases, either before or after whole brain radiation therapy; (b) treatment of leptomeningeal metastases; (c) prevention of brain metastases; and (d) prediction of patients who could benefit from whole brain radiation therapy (17). Most trials of chemotherapy in brain metastases have been small, and almost all enrolled patients after radiation therapy. Two randomized trials of whole brain radiation therapy versus whole brain radiation therapy chemotherapy showed a higher response rate in the combined arms but equivalent survival attributable to progression of systemic disease (17, 18). Several recent trials have used temozolomide with encouraging results as a single agent or combined with whole brain radiation therapy (19–21). Data were presented at the meeting using i.a. carboplatin with i.v. etoposide for brain metastases. In 24 evaluable patients, 6 (25%) achieved a complete response with time to progression of 66 weeks; 11 (46%) achieved a partial response and time to progression of 39 weeks. Overall median time to progression for the entire cohort was 30 weeks. To move forward with chemotherapy for brain metastases, the following approaches were discussed: (a) patients should be stratified/selected according to prognostic classes defined by recursive partitioning analysis to study comparable groups (22); (b) patients with controlled systemic disease should be studied as a group to allow analysis of long-term efficacy; (c) preirradiation chemotherapy trials should be performed to allow better drug penetration and more accurate assessment of response; and (d) Phase III trials must include quality of life and neuropyschometric end points. This need was identified as a priority by the NIH Brain Tumor Progress Review Group. Table 2 lists suggested trial designs for patients with CNS metastases. Immunoconjugates in Primary and Metastatic Tumors. A great deal of research has surrounded the antitumor effects of monoclonal antibody Doxorubicin conjugate BR96-Dox (SGI15; Seattle Genetics, Inc.; Ref. 23). BR96 binds to an extended form of the Lewis Y antigen on the surface of cells from a variety of carcinomas and on antigen binding, rapidly internalizes into endosomes and lysosomes. The bond between the drug and the monoclonal antibody is acid labile and undergoes hydrolysis within these acidic vesicles, leading to the release of active drug. BR96-Dox has pronounced activities in several preclinical models and leads to cures of established tumors at well-tolerated doses (24). Pharmacokinetic studies show that the activities are attributable to high and sustained intratumoral drug concentrations exceeding those obtainable through systemic doxorubicin administration (25). Several approaches have been devised to produce more potent immunoconjugates, including branched linkers that increase drug multiplicity (26) and peptide linkers that lead to enhanced serum stability and rapid lysosomal drug release. These new generation conjugates exhibit improved in vitro and in vivo activities over BR96-Dox. Another approach for more effective immunoconjugates includes utilization of drugs that are significantly more potent than doxorubicin. DNA minor groove binding drugs related in structure to CC-1065, one of the prototypes for potent alkylating agents that bind to the minor groove of DNA (27, 28), and antimitotic drugs related in structure to dolastatin 10 were modified to facilitate attachment to BR96. As with BR96-Dox, the conjugates released the drug under slightly acidic conditions. The conjugates and released drugs were 50–1000-fold more potent than BR96-Dox and doxorubicin, respectively. Significant antitumor activities in preclinical in vivo models were obtained at low doses. The effects were immunologically specific, and the conjugates were well tolerated. These immunoconjugates are strong candidates for clinical development.