Novel treatment option for MUC16-positive malignancies with the targeted TRAIL-based fusion protein Meso-TR3

BMC Cancer

Gunjal Garg⁶  Jesse Gibbs⁴  Brian Belt⁴  Matthew A Powell⁵  David G Mutch⁵  Peter Goedegebuure⁵  Lynne Collins²  David Piwnica-Worms³  William G Hawkins⁵  Dirk Spitzer¹ 

[1] Department of Surgery, Washington University School of Medicine, 660 S. Euclid Avenue, Campus Box 8109, St. Louis, MO 63110, USA

[2] Molecular Imaging Center, Mallinckrodt Institute of Radiology, BRIGHT Institute, Departments of Cell Biology & Physiology, and Developmental Biology, Washington University School of Medicine, St. Louis, MO 63110, USA

[3] Current address: Cancer Systems Imaging Department, Division of Diagnostic Imaging, The University of Texas M.D. Anderson Cancer Center, T. Boone Pickens Academic Tower, 1400 Pressler Street, Unit 1479, Houston, TX 77030, USA

[4] Department of Surgery, Washington University School of Medicine, St. Louis, MO 63110, USA

[5] Siteman Cancer Center, Washington University School of Medicine, St. Louis, MO 63110, USA

[6] Department of Obstetrics and Gynecologic Oncology, Washington University School of Medicine, St. Louis, MO 63110, USA

关键词: Meso-TR3;    Fusion protein;    Biomarker;    MUC16;    Mesothelin;    TRAIL;   

Others  :  859121
DOI  :  10.1186/1471-2407-14-35

 received in 2013-09-19, accepted in 2014-01-18,  发布年份 2014

【 摘 要 】

Background

The targeted delivery of cancer therapeutics represents an ongoing challenge in the field of drug development. TRAIL is a promising cancer drug but its activity profile could benefit from a cancer-selective delivery mechanism, which would reduce potential side effects and increase treatment efficiencies. We recently developed the novel TRAIL-based drug platform TR3, a genetically fused trimer with the capacity for further molecular modifications such as the addition of tumor-directed targeting moieties. MUC16 (CA125) is a well characterized biomarker in several human malignancies including ovarian, pancreatic and breast cancer. Mesothelin is known to interact with MUC16 with high affinity. In order to deliver TR3 selectively to MUC16-expressing cancers, we investigated the possibility of targeted TR3 delivery employing the high affinity mesothelin/MUC16 ligand/receptor interaction.

Methods

Using genetic engineering, we designed the novel cancer drug Meso-TR3, a fusion protein between native mesothelin and TR3. The recombinant proteins were produced with mammalian HEK293T cells. Meso-TR3 was characterized for binding selectivity and killing efficacy against MUC16-positive cancer cells and controls that lack MUC16 expression. Drug efficacy experiments were performed in vitro and in vivo employing an intraperitoneal xenograft mouse model of ovarian cancer.

Results

Similar to soluble mesothelin itself, the strong MUC16 binding property was retained in the Meso-TR3 fusion protein. The high affinity ligand/receptor interaction was associated with a selective accumulation of the cancer drug on MUC16-expressing cancer targets and directly correlated with increased killing activity in vitro and in a xenograft mouse model of ovarian cancer. The relevance of the mesothelin/MUC16 interaction for attaching Meso-TR3 to the cancer cells was verified by competitive blocking experiments using soluble mesothelin. Mechanistic studies using soluble DR5-Fc and caspase blocking assays confirmed engagement of the extrinsic death receptor pathway. Compared to non-targeted TR3, Meso-TR3 displayed a much reduced killing potency on cells that lack MUC16.

Conclusions

Soluble Meso-TR3 targets the cancer biomarker MUC16 in vitro and in vivo. Following attachment to the tumor via surface bound MUC16, Meso-TR3 acquires full activation with superior killing profiles compared to non-targeted TR3, while its bioactivity is substantially reduced on cells that lack the tumor marker. This prodrug phenomenon represents a highly desirable property because it has the potential to enhance cancer killing with fewer side-effects than non-targeted TRAIL-based therapeutics. Thus, further exploration of this novel fusion protein is warranted as a possible therapeutic for patients with MUC16-positive malignancies.

【 授权许可】

   
2014 Garg et al.; licensee BioMed Central Ltd.

【 图 表 】

【 参考文献 】

• [1]Wiley SR, Schooley K, Smolak PJ, Din WS, Huang CP, Nicholl JK, Sutherland GR, Smith TD, Rauch C, Smith CA, et al.: Identification and characterization of a new member of the TNF family that induces apoptosis. Immunity 1995, 3:673-682.
• [2]Pitti RM, Marsters SA, Ruppert S, Donahue CJ, Moore A, Ashkenazi A: Induction of apoptosis by Apo-2 ligand, a new member of the tumor necrosis factor cytokine family. J Biol Chem 1996, 271:12687-12690.
• [3]LeBlanc HN, Ashkenazi A: Apo2L/TRAIL and its death and decoy receptors. Cell Death Differ 2003, 10:66-75.
• [4]Walczak H, Miller RE, Ariail K, Gliniak B, Griffith TS, Kubin M, Chin W, Jones J, Woodward A, Le T, et al.: Tumoricidal activity of tumor necrosis factor-related apoptosis-inducing ligand in vivo. Nat Med 1999, 5:157-163.
• [5]Ashkenazi A, Pai RC, Fong S, Leung S, Lawrence DA, Marsters SA, Blackie C, Chang L, McMurtrey AE, Hebert A, et al.: Safety and antitumor activity of recombinant soluble Apo2 ligand. J Clin Invest 1999, 104:155-162.
• [6]Bretz JD, Mezosi E, Giordano TJ, Gauger PG, Thompson NW, Baker JR Jr: Inflammatory cytokine regulation of TRAIL-mediated apoptosis in thyroid epithelial cells. Cell Death Differ 2002, 9:274-286.
• [7]Ichikawa K, Liu W, Fleck M, Zhang H, Zhao L, Ohtsuka T, Wang Z, Liu D, Mountz JD, Ohtsuki M, et al.: TRAIL-R2 (DR5) mediates apoptosis of synovial fibroblasts in rheumatoid arthritis. J Immunol 2003, 171:1061-1069.
• [8]Strater J, Walczak H, Pukrop T, Von Muller L, Hasel C, Kornmann M, Mertens T, Moller P: TRAIL and its receptors in the colonic epithelium: a putative role in the defense of viral infections. Gastroenterology 2002, 122:659-666.
• [9]Walczak H, Krammer PH: The CD95 (APO-1/Fas) and the TRAIL (APO-2L) apoptosis systems. Exp Cell Res 2000, 256:58-66.
• [10]Wajant H, Moosmayer D, Wuest T, Bartke T, Gerlach E, Schonherr U, Peters N, Scheurich P, Pfizenmaier K: Differential activation of TRAIL-R1 and −2 by soluble and membrane TRAIL allows selective surface antigen-directed activation of TRAIL-R2 by a soluble TRAIL derivative. Oncogene 2001, 20:4101-4106.
• [11]Bremer E, Kuijlen J, Samplonius D, Walczak H, de Leij L, Helfrich W: Target cell-restricted and -enhanced apoptosis induction by a scFv:sTRAIL fusion protein with specificity for the pancarcinoma-associated antigen EGP2. Int J Cancer 2004, 109:281-290.
• [12]Spitzer D, McDunn JE, Plambeck-Suess S, Goedegebuure PS, Hotchkiss RS, Hawkins WG: A genetically encoded multifunctional TRAIL trimer facilitates cell-specific targeting and tumor cell killing. Mol Cancer Ther 2010, 9:2142-2151.
• [13]Gubbels JA, Belisle J, Onda M, Rancourt C, Migneault M, Ho M, Bera TK, Connor J, Sathyanarayana BK, Lee B, et al.: Mesothelin-MUC16 binding is a high affinity, N-glycan dependent interaction that facilitates peritoneal metastasis of ovarian tumors. Mol Cancer 2006, 5:50. BioMed Central Full Text
• [14]Haridas D, Chakraborty S, Ponnusamy MP, Lakshmanan I, Rachagani S, Cruz E, Kumar S, Das S, Lele SM, Anderson JM, et al.: Pathobiological implications of MUC16 expression in pancreatic cancer. PLoS One 2011, 6:e26839.
• [15]Wu YM, Nowack DD, Omenn GS, Haab BB: Mucin glycosylation is altered by pro-inflammatory signaling in pancreatic-cancer cells. J Proteome Res 2009, 8:1876-1886.
• [16]Moritani S, Ichihara S, Hasegawa M, Endo T, Oiwa M, Yoshikawa K, Sato Y, Aoyama H, Hayashi T, Kushima R: Serous papillary adenocarcinoma of the female genital organs and invasive micropapillary carcinoma of the breast. Are WT1, CA125, and GCDFP-15 useful in differential diagnosis? Hum Pathol 2008, 39:666-671.
• [17]Spitzer D, Unsinger J, Bessler M, Atkinson JP: ScFv-mediated in vivo targeting of DAF to erythrocytes inhibits lysis by complement. Mol Immunol 2004, 40:911-919.
• [18]Gross S, Piwnica-Worms D: Real-time imaging of ligand-induced IKK activation in intact cells and in living mice. Nat Methods 2005, 2:607-614.
• [19]Hassan R, Bera T, Pastan I: Mesothelin: a new target for immunotherapy. Clin Cancer Res 2004, 10:3937-3942.
• [20]Chang K, Pastan I: Molecular cloning of mesothelin, a differentiation antigen present on mesothelium, mesotheliomas, and ovarian cancers. Proc Natl Acad Sci USA 1996, 93:136-140.
• [21]Xiang X, Feng M, Felder M, Connor JP, Man YG, Patankar MS, Ho M: HN125: a novel immunoadhesin targeting MUC16 with potential for cancer therapy. J Cancer 2011, 2:280-291.
• [22]Lane D, Cartier A, L’Esperance S, Cote M, Rancourt C, Piche A: Differential induction of apoptosis by tumor necrosis factor-related apoptosis-inducing ligand in human ovarian carcinoma cells. Gynecol Oncol 2004, 93:594-604.
• [23]Lane D, Cote M, Grondin R, Couture MC, Piche A: Acquired resistance to TRAIL-induced apoptosis in human ovarian cancer cells is conferred by increased turnover of mature caspase-3. Mol Cancer Ther 2006, 5:509-521.
• [24]Reis CR, van der Sloot AM, Natoni A, Szegezdi E, Setroikromo R, Meijer M, Sjollema K, Stricher F, Cool RH, Samali A, et al.: Rapid and efficient cancer cell killing mediated by high-affinity death receptor homotrimerizing TRAIL variants. Cell Death Dis 2010, 1:e83.
• [25]Schneider P, Holler N, Bodmer JL, Hahne M, Frei K, Fontana A, Tschopp J: Conversion of membrane-bound Fas(CD95) ligand to its soluble form is associated with downregulation of its proapoptotic activity and loss of liver toxicity. J Exp Med 1998, 187:1205-1213.
• [26]Herbst RS, Eckhardt SG, Kurzrock R, Ebbinghaus S, O’Dwyer PJ, Gordon MS, Novotny W, Goldwasser MA, Tohnya TM, Lum BL, et al.: Phase I dose-escalation study of recombinant human Apo2L/TRAIL, a dual proapoptotic receptor agonist, in patients with advanced cancer. J Clin Oncol 2010, 28:2839-2846.
• [27]Mahalingam D, Szegezdi E, Keane M, de Jong S, Samali A: TRAIL receptor signalling and modulation: are we on the right TRAIL? Cancer Treat Rev 2009, 35:280-288.
• [28]ten Cate B, Bremer E, de Bruyn M, Bijma T, Samplonius D, Schwemmlein M, Huls G, Fey G, Helfrich W: A novel AML-selective TRAIL fusion protein that is superior to Gemtuzumab Ozogamicin in terms of in vitro selectivity, activity and stability. Leukemia 2009, 23:1389-1397.
• [29]Bodmer JL, Meier P, Tschopp J, Schneider P: Cysteine 230 is essential for the structure and activity of the cytotoxic ligand TRAIL. J Biol Chem 2000, 275:20632-20637.
• [30]Liu J, Moore K, Birrer M, Berlin S, Matulonis U, Infante J, Xi J, Kahn R, Wang Y, Wood K, et al.: Targeting MUC16 with the antibody-drug conjugate (ADC) DMUC5754A in patients with platinum-resistant ovarian cancer: a phase I study of safety and pharmacokinetics. Washington, DC: AACR Annual Meeting; 2013. Abstract LB-290
• [31]Muhlenbeck F, Schneider P, Bodmer JL, Schwenzer R, Hauser A, Schubert G, Scheurich P, Moosmayer D, Tschopp J, Wajant H: The tumor necrosis factor-related apoptosis- inducing ligand receptors TRAIL-R1 and TRAIL-R2 have distinct cross-linking requirements for initiation of apoptosis and are non-redundant in JNK activation. J Biol Chem 2000, 275:32208-32213.
• [32]Wilson NS, Yang A, Yang B, Couto S, Stern H, Gogineni A, Pitti R, Marsters S, Weimer RM, Singh M, Ashkenazi A: Proapoptotic activation of death receptor 5 on tumor endothelial cells disrupts the vasculature and reduces tumor growth. Cancer Cell 2012, 22:80-90.
• [33]Singh AP, Moniaux N, Chauhan SC, Meza JL, Batra SK: Inhibition of MUC4 expression suppresses pancreatic tumor cell growth and metastasis. Cancer Res 2004, 64:622-630.
• [34]Rump A, Morikawa Y, Tanaka M, Minami S, Umesaki N, Takeuchi M, Miyajima A: Binding of ovarian cancer antigen CA125/MUC16 to mesothelin mediates cell adhesion. J Biol Chem 2004, 279:9190-9198.
• [35]Scholler N, Garvik B, Hayden-Ledbetter M, Kline T, Urban N: Development of a CA125-mesothelin cell adhesion assay as a screening tool for biologics discovery. Cancer Lett 2007, 247:130-136.
• [36]Bergan L, Gross JA, Nevin B, Urban N, Scholler N: Development and in vitro validation of anti-mesothelin biobodies that prevent CA125/Mesothelin-dependent cell attachment. Cancer Lett 2007, 255:263-274.
• [37]Evdokiou A, Bouralexis S, Atkins GJ, Chai F, Hay S, Clayer M, Findlay DM: Chemotherapeutic agents sensitize osteogenic sarcoma cells, but not normal human bone cells, to Apo2L/TRAIL-induced apoptosis. Int J Cancer 2002, 99:491-504.
• [38]El-Zawahry A, McKillop J, Voelkel-Johnson C: Doxorubicin increases the effectiveness of Apo2L/TRAIL for tumor growth inhibition of prostate cancer xenografts. BMC Cancer 2005, 5:2. BioMed Central Full Text
• [39]Buchsbaum DJ, Zhou T, Grizzle WE, Oliver PG, Hammond CJ, Zhang S, Carpenter M, LoBuglio AF: Antitumor efficacy of TRA-8 anti-DR5 monoclonal antibody alone or in combination with chemotherapy and/or radiation therapy in a human breast cancer model. Clin Cancer Res 2003, 9:3731-3741.