Bharat H. Joshi

Phase III randomized trial of CED of IL13-PE38QQR vs Gliadel wafers for recurrent glioblastoma †

Convection-enhanced delivery (CED) of cintredekin besudotox (CB) was compared with Gliadel wafers (GW) in adult patients with glioblastoma multiforme (GBM) at first recurrence. Patients were randomized 2:1 to receive CB or GW. CB (0.5 microg/mL; total flow rate 0.75 mL/h) was administered over 96 hours via 2-4 intraparenchymal catheters placed after tumor resection. GW (3.85%/7.7 mg carmustine per wafer; maximum 8 wafers) were placed immediately after tumor resection. The primary endpoint was overall survival from the time of randomization. Prestated interim analyses were built into the study design. Secondary and tertiary endpoints were safety and health-related quality-of-life assessments. From March 2004 to December 2005, 296 patients were enrolled at 52 centers. Demographic and baseline characteristics were balanced between the 2 treatment arms. Median survival was 36.4 weeks (9.1 months) for CB and 35.3 weeks (8.8 months) for GW (P = .476). For the efficacy evaluable population, the median survival was 45.3 weeks (11.3 months) for CB and 39.8 weeks (10 months) for GW (P = .310). The adverse-events profile was similar in both arms, except that pulmonary embolism was higher in the CB arm (8% vs 1%, P = .014). This is the first randomized phase III evaluation of an agent administered via CED and the first with an active comparator in GBM patients. There was no survival difference between CB administered via CED and GW. Drug distribution was not assessed and may be crucial for evaluating future CED-based therapeutics.

Specific Targeting of Human Interleukin (IL)-13 Receptor α2-Positive Cells with Lentiviral Vectors Displaying IL-13

The ability to selectively and efficiently target transgene delivery to specific cell types in vitro and in vivo remains one of the formidable challenges in gene therapy. Lentiviral vectors have several advantages that make them attractive as gene delivery vehicles and their tropism can be altered through pseudotyping, allowing transgene delivery to specific populations of cells. The human interleukin-13 receptor α2 (IL-13Rα2) is uniquely overexpressed in many different human tumors, making it an attractive target for cancer therapy. In this study, we examined whether IL-13Rα2-positive tumor cells can be specifically targeted with lentiviral vector pseudotypes containing a truncated fusion (F) protein derived from measles virus (MV) and a tail-truncated and receptor-blind MV hemagglutinin (H) protein bearing IL-13 at the C terminus. The retargeted lentiviral vector efficiently transduced cells that express high levels of IL-13Rα2, but not cells expressing low levels of IL-13Rα2 in vitro. In vivo, it specifically targeted IL-13Rα2-positive glioma cell xenografts in immunodeficient mice in the context of subcutaneous and intracranial glioma models. Similar lentiviral vectors may be developed for targeting other tumors expressing specific cell surface receptors.

Histone modification enhances the effectiveness of IL-13 receptor targeted immunotoxin in murine models of human pancreatic cancer

BackgroundInterleukin-13 Receptor α2 (IL-13Rα2) is a tumor-associated antigen and target for cancer therapy. Since IL-13Rα2 is heterogeneously overexpressed in a variety of human cancers, it would be highly desirable to uniformly upregulate IL-13Rα2 expression in tumors for optimal targeting.MethodsWe examined epigenetic regulation of IL-13Rα2 in a murine model of human pancreatic cancer by Bisulfite-PCR, sequencing for DNA methylation and chromatin immunoprecipitation for histone modification. Reverse transcription-PCR was performed for examining changes in IL-13Rα2 mRNA expression after treatment with histone deacetylase (HDAC) and c-jun inhibitors. In vitro cytotoxicity assays and in vivo testing in animal tumor models were performed to determine whether HDAC inhibitors could enhance anti-tumor effects of IL-13-PE in pancreatic cancer. Mice harboring subcutaneous tumors were treated with HDAC inhibitors systemically and IL-13-PE intratumorally.ResultsWe found that CpG sites in IL-13Rα2 promoter region were not methylated in all pancreatic cancer cell lines studied including IL-13Rα2-positive and IL-13Rα2-negative cell lines and normal cells. On the other hand, histones at IL-13Rα2 promoter region were highly-acetylated in IL-13Rα2-positive but much less in receptor-negative pancreatic cancer cell lines. When cells were treated with HDAC inhibitors, not only histone acetylation but also IL-13Rα2 expression was dramatically enhanced in receptor-negative pancreatic cancer cells. In contrast, HDAC inhibition did not increase IL-13Rα2 in normal cell lines. In addition, c-jun in IL-13Rα2-positive cells was expressed at higher level than in negative cells. Two types of c-jun inhibitors prevented increase of IL-13Rα2 by HDAC inhibitors. HDAC inhibitors dramatically sensitized cancer cells to immunotoxin in the cytotoxicity assay in vitro and increased IL-13Rα2 in the tumors subcutaneously implanted in the immunodeficient animals but not in normal mice tissues. Combination therapy with HDAC inhibitors and immunotoxin synergistically inhibited growth of not only IL-13Rα2-positive but also IL-13Rα2-negative tumors.ConclusionsWe have identified a novel function of histone modification in the regulation of IL-13Rα2 in pancreatic cancer cell lines in vitro and in vivo. HDAC inhibition provides a novel opportunity in designing combinatorial therapeutic approaches not only in combination with IL-13-PE but with other immunotoxins for therapy of pancreatic cancer and other cancers.

Histone deacetylase inhibitiors enhance sensitivity of murine sarcoma tumors to IL-13-PE immunotoxin-based cancer therapy by upregulating IL-13Rα2 expression in vitro and in vivo

Interleukin-13 Receptor alpha 2 (IL-13Rα2) is a tumor antigen and a potent target for cancer therapy. Previously, we have reported that histone deacetylase (HDAC) inhibitors upregulated the IL-13Rα2 expression and enhanced anti-cancer effects of IL-13-PE, an immunotoxin composed of interleukin-13 and truncated Pseudomonas exotoxin, in a mouse model of human pancreatic cancer. Herein, we have investigated whether HDAC inhibitors can enhance the effect of IL-13-PE in immunocompetent mouse tumor models by upregulating IL-13Rα2. We analyzed mRNA and protein levels of mouse IL-13Rα2 in four mouse tumor cell lines, MCA304 sarcoma, 4T1 breast cancer, GL261 glioma and D5 melanoma. By RT-PCR analysis, mRNA levels for IL-13Rα2 were high in MCA304, moderate in 4T1, low in GL261 and below the detection limit in D5 mouse tumor cell lines. These cell lines were treated with three types of HDAC inhibitors, trichostatin A, sodium butyrate and suberoylanilide hydroxamic acid (SAHA) to evaluate their effects on IL-13Rα2 expression in vitro. All HDAC inhibitors dramatically increased IL-13Rα2 mRNA expression in MCA304, 4T1, and GL261 cell lines. The D5 tumor cell line showed only a slight increase in mRNA levels of IL-13Rα2. Western blot analysis for IL-13Rα2 demonstrated increased levels of IL-13Rα2 protein in all of the HDAC inhibitor treated tumor cells. We also observed that all three HDAC inhibitors selectively enhanced cytotoxicity of IL-13-PE in MCA304 and 4T1 cell lines. TSA treatment resulted in maximum improvement of IL-13-PE induced cytotoxicity in MCA304, NaB in GL261, and SAHA in 4T1 cell lines. We next developed a subcutaneous tumor model of mouse sarcoma by implanting MCA304 tumor cells in C57BL/6 mice. Mice were treated with SAHA (an FDA licensed drug) i.p. at 25mg/kg/day dose from day 4 through 9 prior to IL-13-PE immunotoxin treatment. IL-13-PE was administered at a 250mg/kg dose intratumorally from day 5 through 9. Mice treated with vehicle control and SAHA showed tumor growth to ~1500 mm3 in ~22 days and were sacrificed because of ethical reasons. Treatment with IL-13-PE alone showed a significant reduction in tumor growth and mice survived longer. Tumor size reached ~750 mm3 in 35 days. However, when IL-13-PE was administered to SAHA pretreated mice, tumor size showed further reduction and reached only ~400 mm3 on day 35. Additional studies are ongoing to determine the mechanism of synergistic antitumor effects of HDAC inhibitors and IL-13-PE. These results suggest that HDAC inhibitors may upregulate tumor antigens in vivo and thus may be considered in combination with cancer vaccines and immunotherapy products for cancer therapy.

Abstract 3932: IL13Rα2 regulates cell invasion, and is a therapeutic target for adrenocortical carcinoma

Adrencortical neoplasms are one of the most common human neopolasms. Furthermore, it is difficult to distinguish localized benign from malignant adrenal tumors. Adrenocortical carcinoma (ACC) is a relatively rare but aggressive malignancy with limited therapeutic options for patients with advanced and metastatic disease. Several genome wide expression studies have shown overexpression of Interleukin-13 Receptor alpha2 (IL13Rα2) in some human malignancies but the function of this gene in cancer cells is poorly understood. We evaluated IL13Rα2 mRNA expression in normal (n=21), and in benign (n=78) and malignant (n=10) adrenocortical tumors. We found 11-fold higher expression in malignant tumors compared to benign (p Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 3932. doi:1538-7445.AM2012-3932