Preclinical evaluation of a novel engineered recombinant human anti-CD44v6 antibody for potential use in radio-immunotherapy

Mortensen,Anja  C.; Spiegelberg,Diana; Haylock,Anna-Karin; Lundqvist,Hans; Nestor,Marika

doi:10.3892/ijo.2018.4364

Preclinical evaluation of a novel engineered recombinant human anti-CD44v6 antibody for potential use in radio-immunotherapy

Authors:
- Anja C. Mortensen
- Diana Spiegelberg
- Anna-Karin Haylock
- Hans Lundqvist
- Marika Nestor
View Affiliations

Affiliations: Department of Immunology, Genetics and Pathology, Uppsala University, SE-75185 Uppsala, Sweden, Unit of Otolaryngology and Head and Neck Surgery, Department of Surgical Sciences, Uppsala University, SE-75185 Uppsala, Sweden
Published online on: April 11, 2018 https://doi.org/10.3892/ijo.2018.4364
Pages: 1875-1885
Copyright: © Mortensen et al. This is an open access article distributed under the terms of Creative Commons Attribution License.

Metrics: Total Views: 0 (Spandidos Publications: | PMC Statistics: )

Metrics: Total PDF Downloads: 0 (Spandidos Publications: | PMC Statistics: )

Cited By (CrossRef): 0 citations Loading Articles...

Abstract

CD44v6 is overexpressed in a variety of cancers, rendering it a promising target for radio-immunotherapy (RIT). In this study, we have characterized a novel engineered recombinant monoclonal anti-CD44v6 antibody, AbN44v6, and assessed its potential for use in RIT using either 177Lu or 131I as therapeutic radionuclides. In vitro affinity and specificity assays characterized the binding of the antibody labeled with 177Lu, 125I or 131I. The therapeutic effects of 177Lu-AbN44v6 and 131I-AbN44v6 were investigated using two in vitro 3D tumor models with different CD44v6 expression. Finally, the normal tissue biodistribution and dosimetry for 177Lu-AbN44v6 and 125I-AbN44v6/131I-AbN44v6 were assessed in vivo using a mouse model. All AbN44v6 radioconjugates demonstrated CD44v6-specific binding in vitro. In the in vitro 3D tumor models, dose-dependent therapeutic effects were observed with both 177Lu-AbN44v6 and 131I-AbN44v6, with a greater significant therapeutic effect observed on the cells with a higher CD44v6 expression. Biodistribution experiments demonstrated a greater uptake of 177Lu-AbN44v6 in the liver, spleen and bone, compared to 125I-AbN44v6, whereas 125I-AbN44v6 demonstrated a longer circulation time. In dosimetric calculations, the critical organs for 177Lu-AbN44v6 were the liver and spleen, whereas the kidneys and red marrow were considered the critical organs for 131I-AbN44v6. The effective dose was in the order of 0.1 mSv/MBq for both labels. In conclusion, AbN44v6 bound specifically and with high affinity to CD44v6. Furthermore, in vitro RIT demonstrated growth inhibition in a CD44v6-specific activity-dependent manner for both radioconjugates, demonstrating that both 177Lu-AbN44v6 and 131I-AbN44v6 may be promising RIT candidates. Furthermore, biodistribution and dosimetric analysis supported the applicability of both conjugates for RIT. The CD44v6-specific therapeutic effects observed with radiolabeled AbN44v6 in the 3D tumor models in vitro, combined with the beneficial dosimetry in vivo, render AbN44v6 a potential candidate for RIT.

View Figures

View References

1	Schmidt F and Efferth T: Tumor heterogeneity, single-cell sequencing, and drug resistance. Pharmaceuticals (Basel). 9. pp. 332016, View Article : Google Scholar
2	Zukotynski K, Jadvar H, Capala J and Fahey F: Targeted radionuclide therapy: Practical applications and future prospects. Biomark Cancer. 8(Suppl 2): 35–38. 2016.PubMed/NCBI
3	Otte A: Neuroendocrine tumors: Peptide receptors radionuclide therapy (PRRT). Hell J Nucl Med. 19:1822016.PubMed/NCBI
4	Dash A, Knapp FF and Pillai MR: Targeted radionuclide therapy - an overview. Curr Radiopharm. 6:152–180. 2013. View Article : Google Scholar : PubMed/NCBI
5	Sneath RJ and Mangham DC: The normal structure and function of CD44 and its role in neoplasia. Mol Pathol. 51:191–200. 1998. View Article : Google Scholar
6	Todaro M, Gaggianesi M, Catalano V, Benfante A, Iovino F, Biffoni M, Apuzzo T, Sperduti I, Volpe S, Cocorullo G, et al: CD44v6 is a marker of constitutive and reprogrammed cancer stem cells driving colon cancer metastasis. Cell Stem Cell. 14:342–356. 2014. View Article : Google Scholar : PubMed/NCBI
7	Shi J, Zhou Z, Di W and Li N: Correlation of CD44v6 expression with ovarian cancer progression and recurrence. BMC Cancer. 13:1822013. View Article : Google Scholar : PubMed/NCBI
8	Tjhay F, Motohara T, Tayama S, Narantuya D, Fujimoto K, Guo J, Sakaguchi I, Honda R, Tashiro H and Katabuchi H: CD44 variant 6 is correlated with peritoneal dissemination and poor prognosis in patients with advanced epithelial ovarian cancer. Cancer Sci. 106:1421–1428. 2015. View Article : Google Scholar : PubMed/NCBI
9	Ni J, Cozzi PJ, Hao JL, Beretov J, Chang L, Duan W, Shigdar S, Delprado WJ, Graham PH, Bucci J, et al: CD44 variant 6 is associated with prostate cancer metastasis and chemo-/radioresistance. Prostate. 74:602–617. 2014. View Article : Google Scholar : PubMed/NCBI
10	Wu XJ, Li XD, Zhang H, Zhang X, Ning ZH, Yin YM and Tian Y: Clinical significance of CD44s, CD44v3 and CD44v6 in breast cancer. J Int Med Res. 43:173–179. 2015. View Article : Google Scholar
11	Afify A, Durbin-Johnson B, Virdi A and Jess H: The expression of CD44v6 in colon: From normal to malignant. Ann Diagn Pathol. 20:19–23. 2016. View Article : Google Scholar
12	Fox SB, Fawcett J, Jackson DG, Collins I, Gatter KC, Harris AL, Gearing A and Simmons DL: Normal human tissues, in addition to some tumors, express multiple different CD44 isoforms. Cancer Res. 54:4539–4546. 1994.PubMed/NCBI
13	Heider KH, Mulder JW, Ostermann E, Susani S, Patzelt E, Pals ST and Adolf GR: Splice variants of the cell surface glycoprotein CD44 associated with metastatic tumour cells are expressed in normal tissues of humans and cynomolgus monkeys. Eur J Cancer. 31A:2385–2391. 1995. View Article : Google Scholar : PubMed/NCBI
14	Lv L, Liu HG, Dong SY, Yang F, Wang QX, Guo GL, Pan YF and Zhang XH: Upregulation of CD44v6 contributes to acquired chemoresistance via the modulation of autophagy in colon cancer SW480 cells. Tumour Biol. 37:8811–8824. 2016. View Article : Google Scholar : PubMed/NCBI
15	Bellerby R, Smith C, Kyme S, Gee J, Günthert U, Green A, Rakha E, Barrett-Lee P and Hiscox S: Overexpression of specific CD44 isoforms is associated with aggressive cell features in acquired endocrine resistance. Front Oncol. 6:1452016. View Article : Google Scholar : PubMed/NCBI
16	Saito S, Okabe H, Watanabe M, Ishimoto T, Iwatsuki M, Baba Y, Tanaka Y, Kurashige J, Miyamoto Y and Baba H: CD44v6 expression is related to mesenchymal phenotype and poor prognosis in patients with colorectal cancer. Oncol Rep. 29:1570–1578. 2013. View Article : Google Scholar : PubMed/NCBI
17	Omran OM and Ata HS: CD44s and CD44v6 in diagnosis and prognosis of human bladder cancer. Ultrastruct Pathol. 36:145–152. 2012. View Article : Google Scholar : PubMed/NCBI
18	Shiozaki M, Ishiguro H, Kuwabara Y, Kimura M, Mitsui A, Naganawa Y, Shibata T, Fujii Y and Takeyama H: Expression of CD44v6 is an independent prognostic factor for poor survival in patients with esophageal squamous cell carcinoma. Oncol Lett. 2:429–434. 2011. View Article : Google Scholar : PubMed/NCBI
19	Motohara T, Fujimoto K, Tayama S, Narantuya D, Sakaguchi I, Tashiro H and Katabuchi H: CD44 variant 6 as a predictive biomarker for distant metastasis in patients with epithelial ovarian cancer. Obstet Gynecol. 127:1003–1011. 2016. View Article : Google Scholar : PubMed/NCBI
20	Yang Q, Liu Y, Huang Y, Huang D, Li Y, Wu J and Duan M: Expression of COX-2, CD44v6 and CD147 and relationship with invasion and lymph node metastasis in hypopharyngeal squamous cell carcinoma. PLoS One. 8:e710482013. View Article : Google Scholar : PubMed/NCBI
21	Liu HG, Lv L and Shen H: Intratumoral heterogeneity of CD44v6 in rectal cancer. Clin Transl Oncol. 19:425–431. 2017. View Article : Google Scholar
22	Spiegelberg D, Kuku G, Selvaraju R and Nestor M: Characterization of CD44 variant expression in head and neck squamous cell carcinomas. Tumour Biol. 35:2053–2062. 2014. View Article : Google Scholar :
23	Spiegelberg D and Nilvebrant J: CD44v6-targeted imaging of head and neck squamous cell carcinoma: Antibody-based approaches. Contrast Media Mol Imaging. 2017:27095472017. View Article : Google Scholar : PubMed/NCBI
24	Haylock AK, Spiegelberg D, Mortensen AC, Selvaraju RK, Nilvebrant J, Eriksson O, Tolmachev V and Nestor MV: Evaluation of a novel type of imaging probe based on a recombinant bivalent mini-antibody construct for detection of CD44v6-expressing squamous cell carcinoma. Int J Oncol. 48:461–470. 2016. View Article : Google Scholar :
25	Spiegelberg D, Mortensen AC, Selvaraju RK, Eriksson O, Stenerlöw B and Nestor M: Molecular imaging of EGFR and CD44v6 for prediction and response monitoring of HSP90 inhibition in an in vivo squamous cell carcinoma model. Eur J Nucl Med Mol Imaging. 43:974–982. 2016. View Article : Google Scholar :
26	Börjesson PK, Postema EJ, Roos JC, Colnot DR, Marres HA, van Schie MH, Stehle G, de Bree R, Snow GB, Oyen WJ, et al: Phase I therapy study with (186)Re-labeled humanized monoclonal antibody BIWA 4 (bivatuzumab) in patients with head and neck squamous cell carcinoma. Clin Cancer Res. 9:3961S–3972S. 2003.PubMed/NCBI
27	Jødal L: Beta emitters and radiation protection. Acta Oncol. 48:308–313. 2009. View Article : Google Scholar
28	Mujammami M, Hier MP, Payne RJ, Rochon L and Tamilia M: Long-term outcomes of patients with papillary thyroid cancer undergoing remnant ablation with 30 milliCuries radioiodine. Thyroid. 26:951–958. 2016. View Article : Google Scholar
29	Makis W, McCann K and McEwan AJ: Orbital metastases of neuroendocrine tumors treated with ¹⁷⁷Lu-DOTATATE PRRT or 131I-MIBG therapies. Clin Nucl Med. 41:137–141. 2016. View Article : Google Scholar
30	Bergsma H, Konijnenberg MW, van der Zwan WA, Kam BL, Teunissen JJ, Kooij PP, Mauff KA, Krenning EP and Kwekkeboom DJ: Nephrotoxicity after PRRT with (177)Lu-DOTA-octreotate. Eur J Nucl Med Mol Imaging. 43:1802–1811. 2016. View Article : Google Scholar : PubMed/NCBI
31	NuDat 2 [Internet]. Cited Mar 7, 2017. Available from: http://www.nndc.bnl.gov/nudat2/.
32	Hindié E, Zanotti-Fregonara P, Quinto MA, Morgat C and Champion C: Dose deposits from ⁹⁰Y, ¹⁷⁷Lu, ¹¹¹In, and ¹⁶¹Tb in micrometastases of various sizes: Implications for radiopharma-ceutical therapy. J Nucl Med. 57:759–764. 2016. View Article : Google Scholar
33	Wang G, Wu Z, Wang Y, Li X, Zhang G and Hou J: Therapy to target renal cell carcinoma using 131I-labeled B7-H3 monoclonal antibody. Oncotarget. 7:24888–24898. 2016.PubMed/NCBI
34	Long B, Yang M, Yang Z, Yi H and Li L: Assessment of radio-iodine therapy efficacy for treatment of differentiated thyroid cancer patients with pulmonary metastasis undetected by chest computed tomography. Oncol Lett. 11:965–968. 2016. View Article : Google Scholar : PubMed/NCBI
35	Steidl S, Ratsch O, Brocks B, Dürr M and Thomassen-Wolf E: In vitro affinity maturation of human GM-CSF antibodies by targeted CDR-diversification. Mol Immunol. 46:135–144. 2008. View Article : Google Scholar : PubMed/NCBI
36	Ostendorp R, Frisch C and Urban M: Generation, engineering and production of human antibodies using hucal. Antibodies. Subramanian G: Springer; US: pp. 13–52. 2004, View Article : Google Scholar
37	Cho MS, Yee H and Chan S: Establishment of a human somatic hybrid cell line for recombinant protein production. J Biomed Sci. 9:631–638. 2002. View Article : Google Scholar : PubMed/NCBI
38	Wolff J and Chaikoff IL: Plasma inorganic iodide as a homeo-static regulator of thyroid function. J Biol Chem. 174:555–564. 1948.PubMed/NCBI
39	Eng PH, Cardona GR, Fang SL, Previti M, Alex S, Carrasco N, Chin WW and Braverman LE: Escape from the acute Wolff-Chaikoff effect is associated with a decrease in thyroid sodium/iodide symporter messenger ribonucleic acid and protein. Endocrinology. 140:3404–3410. 1999. View Article : Google Scholar : PubMed/NCBI
40	Tolmachev V, Malmberg J, Estrada S, Eriksson O and Orlova A: Development of a 124I-labeled version of the anti-PSMA monoclonal antibody capromab for immunoPET staging of prostate cancer: Aspects of labeling chemistry and biodistribution. Int J Oncol. 44:1998–2008. 2014. View Article : Google Scholar : PubMed/NCBI
41	Sehlin D, Fang XT, Meier SR, Jansson M and Syvänen S: Pharmacokinetics, biodistribution and brain retention of a bispecific antibody-based PET radioligand for imaging of amyloid-β. Sci Rep. 7:172542017. View Article : Google Scholar
42	Haylock A-K, Spiegelberg D, Nilvebrant J, Sandström K and Nestor M: In vivo characterization of the novel CD44v6-targeting Fab fragment AbD15179 for molecular imaging of squamous cell carcinoma: A dual-isotope study. EJNMMI Res. 4:112014. View Article : Google Scholar : PubMed/NCBI
43	Steffen AC, Wikman M, Tolmachev V, Adams GP, Nilsson FY, Ståhl S and Carlsson J: In vitro characterization of a bivalent anti-HER-2 affibody with potential for radionuclide-based diagnostics. Cancer Biother Radiopharm. 20:239–248. 2005. View Article : Google Scholar : PubMed/NCBI
44	Brown BA, Comeau RD, Jones PL, Liberatore FA, Neacy WP, Sands H and Gallagher BM: Pharmacokinetics of the monoclonal antibody B72.3 and its fragments labeled with either ¹²⁵I or ¹¹¹In. Cancer Res. 47:1149–1154. 1987.PubMed/NCBI
45	Ahl PL, Bhatia SK, Meers P, Roberts P, Stevens R, Dause R, Perkins WR and Janoff AS: Enhancement of the in vivo circulation lifetime of L-alpha-distearoylphosphatidylcholine liposomes: Importance of liposomal aggregation versus complement opsonization. Biochim Biophys Acta. 1329:370–382. 1997. View Article : Google Scholar : PubMed/NCBI
46	Holechek MJ: Glomerular filtration: An overview. Nephrol Nurs J. 30:285–290; quiz 291-292. 2003.PubMed/NCBI
47	Blau M: Letter: Radiation dosimetry of 131-I-19-iodocholesterol: The pitfalls of using tissue concentration data. J Nucl Med. 16:247–249. 1975.PubMed/NCBI