A New Discovery in the diagnosis of HPV-associated cervical cancers

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Dr. Lifang Zhang a renowned scientist from Wenzhou Medical University, China has made a new discovery in the diagnosis of HPV-associated cervical cancers. According to her research on HPV-associated cervical cancers, Human papillomavirus HPV16, HPV18 is widely known as the one responsible for cervical cancers. Cervical cancer is one the world’s most common gynecologic malignancy in women nowadays.

Among medical researchers, it is a well-known fact that viral oncoproteins E6/E7 plays an important role in the formation of cancer tumors in HPV-associated cervical cancers. So, by discovering these two oncoproteins in the patient’s body can provide vital information regarding the diagnosis of HPV-associated cervical cancer in the patient’s body. Dr. Lifang Zhang and her team had found that affibody molecules will be very useful in molecular imaging probes for cervical cancer, and has proved to be a promising candidate for development as molecular imaging probes. Affibody molecules are nothing but small, robust proteins engineered to bind to a large number of target proteins or peptides with high affinity, imitating monoclonal antibodies, which comes under a member of the family of antibody mimetics.

Dr. Lifang Zhang and his team have generated two monomeric affibody molecules (ZHPV16E7 and ZHPV18E7) in their laboratory for their testing purposes. They have used a peptide linker (Gly4Ser)3 to link these two monomeric affibody molecules (ZHPV16E7 and ZHPV18E7) to create a novel heterodimeric affibody ZHPV16E7–(Gly4Ser)3–ZHPV18E7. After that, they have done biosensor and immunofluorescence assays to find out the influence of the newly created monomeric affibody molecules. Results have proved that the heterodimeric affibody has targeted both HPV16 and HPV18E7 proteins by binding to the viral oncoproteins. In vivo tumor-imaging experiments using the Dylight755-labeled heterodimeric affibody showed that strongly high-contrast tumor retention of the heterodimers occurred in both HPV16- and HPV18-derived tumors of nude mice 0.5 h post- injection. The above results were obtained after 48 hours of Dylight755-labeled heterodimeric affibody test. This has proved that using heterodimeric affibody molecule has a great potential in binding to HPV16 and HPV18 proteins, therefore it can be used in molecular imaging in vivo and diagnosis of HPV-associated cervical cancers.

Results:

Fig. 1  Schematic presentation, SDS-PAGE, Western blot analysis and tumor uptake of the dimeric affibody molecules. a Schematic presentation of the dimeric affibody molecules. ZHPV16E7 = HPV16E7-specific affibody molecule, ZHPV18E7 = HPV18E7-specific affibody molecule, Linker = (G4S)3 peptide linker. b The predicted 3D structure of the dimeric affibody molecules. c SDS-PAGE and Western blot analysis of the purified affibody molecules. Lane 1, Bispecific affibody molecule ZHPV16E7-ZHPV18E7; lane 2, ZWT; lane 3, Western blot analysis of ZHPV16E7-ZHPV18E7; lane 4, ZHPV16E7-ZHPV16E7; lane 5, ZHPV16E7 ; lane 6, Western blot analysis of ZHPV16E7-ZHPV16E7; lane 7, ZHPV18E7-ZHPV18E7; lane 8, ZHPV18E7; lane 9, Western blot analysis of ZHPV18E7-ZHPV18E7; M, protein ladder. The mouse anti-His mAb served as the primary antibody in Western blot assay.

d In vivo uptake of affibody by subcutaneous tumor xenografts. Mice bearing SiHa and HeLa cells xenografts (circles) were intravenously injected with Dylight755-labeled heterodimers, homodimers or monomers affibody molecules followed by dynamic scanning with in vivo NIR System.   

This breakthrough research has led by Dr. Lifang Zhang and her team in Institute of Molecular Virology and Immunology, Department of Microbiology and Immunology, School of Basic Medical Sciences, Wenzhou Medical University, Wenzhou, 325035, Zhejiang, People’s Republic of China.

Research Support:

This research has been supported by:

81172463/National Nature Science Foundation of China/

81502242/National Nature Science Foundation of China/

LGF18C010004/Public Welfare Foundation of Zhejiang Province

Citations:

  1. Andersson KG, Oroujeni M, Garousi J, Mitran B, Ståhl S, Orlova A, Löfblom J, Tolmachev V (2016) Feasibility of imaging of epidermal growth factor receptor expression with ZEGFR:2377 affibody molecule labeled with99mTc using a peptide-based cysteine-containing chelator. Int J Oncol 49(6):2285–2293.  https://doi.org/10.3892/ijo.2016.3721CrossRefPubMedPubMedCentralGoogle Scholar
  2. Badaracco G, Venuti A, Sedati A, Marcante ML (2002) HPV16 and HPV18 in genital tumors: significantly different levels of viral integration and correlation to tumor invasiveness. J Med Virol 67(4):574–582CrossRefPubMedGoogle Scholar
  3. Byrne H, Conroy PJ, Whisstock JC, O’Kennedy RJ (2013) A tale of two specificities: bispecific antibodies for therapeutic and diagnostic applications. Trends Biotechnol 31(11):621–632.  https://doi.org/10.1016/j.tibtech.2013.08.007CrossRefPubMedGoogle Scholar
  4. Cheng Q, Wållberg H, Grafström J, Lu L, Thorell JO, Hägg O, Linder S, Johansson K, Tegnebratt T, Arnér ES, Stone-Elander S, Ahlzén HS, Ståhl S, Sel-tag imaging project (2016) Preclinical PET imaging of EGFR levels: pairing a targeting with a non-targeting Sel-tagged Affibody-based tracer to estimate the specific uptake. EJNMMI Res 6(1):58.  https://doi.org/10.1186/s13550-016-0213-8CrossRefPubMedPubMedCentralGoogle Scholar
  5. Choi JW, Hong ST, Kang DE, Paik KC, Han MS, Lim CS, Cho BR (2016) A two-photon tracer for human epidermal growth factor receptor-2: detection of breast cancer in a live tissue. Anal Chem 88(19):9412–9418CrossRefPubMedGoogle Scholar
  6. Dall’Acqua W, Simon AL, Mulkerrin MG, Carter P (1998) Contribution of domain interface residues to the stability of antibody CH3 domain homodimers. Biochemistry 37(26):9266–9273CrossRefPubMedGoogle Scholar
  7. de Sanjose S, Quint WG, Alemany L, Geraets DT, Klaustermeier JE, Lloveras B, Tous S, Felix A, Bravo LE, Shin HR, Vallejos CS, de Ruiz PA, Lima MA, Guimera N, Clavero O, Alejo M, Llombart-Bosch A, Cheng-Yang C, Tatti SA, Kasamatsu E, Iljazovic E, Odida M, Prado R, Seoud M, Grce M, Usubutun A, Jain A, Suarez GA, Lombardi LE, Banjo A, Menéndez C, Domingo EJ, Velasco J, Nessa A, Chichareon SC, Qiao YL, Lerma E, Garland SM, Sasagawa T, Ferrera A, Hammouda D, Mariani L, Pelayo A, Steiner I, Oliva E, Meijer CJ, Al-Jassar WF, Cruz E, Wright TC, Puras A, Llave CL, Tzardi M, Agorastos T, Garcia-Barriola V, Clavel C, Ordi J, Andújar M, Castellsagué X, Sánchez GI, Nowakowski AM, Bornstein J, Muñoz N, Bosch FX, Retrospective International Survey and HPV Time Trends Study Group (2010) Human papillomavirus genotype attribution in invasive cervical cancer: a retrospective cross-sectional worldwide study. Lancet Oncol 11(11):1048–1056.  https://doi.org/10.1016/S1470-2045(10)70230-8CrossRefPubMedGoogle Scholar
  8. Ekerljung L, Wållberg H, Sohrabian A, Andersson K, Friedman M, Frejd FY, Ståhl S, Gedda L (2012) Generation and evaluation of bispecific affibody molecules for simultaneous targeting of EGFR and HER2. Bioconjug Chem 23(9):1802–1811CrossRefPubMedGoogle Scholar
  9. Fedorova A, Zobel K, Gill HS, Ogasawara A, Flores JE, Tinianow JN, Vanderbilt AN, Wu P, Meng YG, Williams SP, Wiesmann C, Murray J, Marik J, Deshayes K (2011) The development of peptide-based tools for the analysis of angiogenesis. Chem Biol 18(7):839–845.  https://doi.org/10.1016/j.chembiol.2011.05.011CrossRefPubMedGoogle Scholar
  10. 10. Frejd FY, Kim K (2017) Affibody molecules as engineered protein drugs. Exp Mol Med 49(3):e306.  https://doi.org/10.1038/emm.2017.35CrossRefPubMedPubMedCentralGoogle Scholar
  11. Friedman M, Lindström S, Ekerljung L, Andersson-Svahn H, Carlsson J, Brismar H, Gedda L, Frejd FY, Ståhl S (2009) Engineering and characterization of a bispecific HER2 x EGFR-binding affibody molecule. Biotechnol Appl Biochem 54(2):121–131.  https://doi.org/10.1042/BA20090096CrossRefPubMedGoogle Scholar
  12. Garousi J, Andersson KG, Mitran B, Pichl M, Ståhl S, Orlova A, Löfblom J, Tolmachev V (2016) PET imaging of epidermal growth factor receptor expression in tumours using 89Zr-labelled ZEGFR:2377 affibody molecules. Int J Oncol 48(4):1325–1332.  https://doi.org/10.3892/ijo.2016.3369CrossRefPubMedPubMedCentralGoogle Scholar
  13. Hopman AH, Smedts F, Dignef W, Ummelen M, Sonke G, Mravunac M, Vooijs GP, Speel EJ, Ramaekers FC (2004) Transition of high-grade cervical intraepithelial neoplasia to micro-invasive carcinoma is characterized by integration of HPV 16/18 and numerical chromosome abnormalities. J Pathol 202(1):23–33CrossRefPubMedGoogle Scholar
  14. Kontermann RE (2012) Dual targeting strategies with bispecific antibodies. Mabs 4(2):182–197.  https://doi.org/10.4161/mabs.4.2.19000CrossRefPubMedPubMedCentralGoogle Scholar
  15. Kronqvist N, Malm M, Göstring L, Gunneriusson E, Nilsson M, Höidén Guthenberg I, Gedda L, Frejd FY, Ståhl S, Löfblom J (2011) Combining phage and staphylococcal surface display for generation of ErbB3-specific affibody molecules. Protein Eng Des Sel 24(4):385–396.  https://doi.org/10.1093/protein/gzq118CrossRefPubMedGoogle Scholar
  16. Löfblom J, Feldwisch J, Tolmachev V, Carlsson J, Ståhl S, Frejd FY (2010) Affibody molecules: engineered proteins for therapeutic, diagnostic and biotechnological applications. FEBS Lett 584(12):2670–2680.  https://doi.org/10.1016/j.febslet.2010.04.014CrossRefPubMedGoogle Scholar
  17. Löfblom J, Frejd FY, Ståhl S (2011) Non-immunoglobulin based protein scaffolds. Curr Opin Biotechnol 22(6):843–848.  https://doi.org/10.1016/j.copbio.2011.06.002CrossRefPubMedGoogle Scholar
  18. Lu P, Feng MG (2008) Bifunctional enhancement of a β-glucanase-xylanase fusion enzyme by optimization of peptide linkers. Appl Microbiol Biotechnol 79(4):579–587.  https://doi.org/10.1007/s00253-008-1468-4CrossRefPubMedGoogle Scholar
  19. Malm M, Bass T, Gudmundsdotter L, Lord M, Frejd FY, Ståhl S, Löfblom J (2014) Engineering of a bispecific affibody molecule towards HER2 and HER3 by addition of an albumin-binding domain allows for affinity purification and in vivo half-life extension. Biotechnol J 9(9):1215–1222.  https://doi.org/10.1002/biot.201400009CrossRefPubMedGoogle Scholar
  20. McBride WJ, Zanzonico P, Sharkey RM, Norén C, Karacay H, Rossi EA, Losman MJ, Brard PY, Chang CH, Larson SM, Goldenberg DM (2006) Bispecific antibody pretargeting PET (immunoPET) with an 124I-labeled hapten-peptide. J Nucl Med 47(10):1678–1688PubMedGoogle Scholar
  21. Mirabello L, Yeager M, Yu K, Clifford GM, Xiao Y, Zhu B, Cullen M, Boland JF, Wentzensen N, Nelson CW, Raine-Bennett T, Chen Z, Bass S, Song L, Yang Q, Steinberg M, Burdett L, Dean M, Roberson D, Mitchell J, Lorey T, Franceschi S, Castle PE, Walker J, Zuna R, Kreimer AR, Beachler DC, Hildesheim A, Gonzalez P, Porras C, Burk RD, Schiffman M (2017) HPV16 E7 genetic conservation is critical to carcinogenesis. Cell 170(6):1164–1174.e6.  https://doi.org/10.1016/j.cell.2017.08.001CrossRefPubMedPubMedCentralGoogle Scholar
  22. Muñoz N, Bosch FX, de Sanjosé S, Herrero R, Castellsagué X, Shah KV, Snijders PJ, Meijer CJ, International Agency for Research on Cancer Multicenter Cervical Cancer Study Group (2003) Epidemiologic classification of human papillomavirus types associated with cervical cancer. N Engl J Med 348(6):518–527CrossRefPubMedGoogle Scholar
  23. Ogilvie GS, Krajden M, Van Niekerk D, Smith LW, Cook D, Ceballos K, Lee M, Gentile L, Gondara L, Elwood-Martin R, Peacock S, Stuart G, Franco EL, Coldman AJ (2017) HPV for cervical cancer screening (HPV FOCAL): complete round 1 results of a randomized trial comparing HPV-based primary screening to liquid-based cytology for cervical cancer. Int J Cancer 140(2):440–448.  https://doi.org/10.1002/ijc.30454CrossRefPubMedGoogle Scholar
  24. Orlova A, Magnusson M, Eriksson TL, Nilsson M, Larsson B, Höidén-Guthenberg I, Widström C, Carlsson J, Tolmachev V, Ståhl S, Nilsson FY (2006) Tumor imaging using a picomolar affinity HER2 binding affibody molecule. Cancer Res 66(8):4339–4348CrossRefPubMedGoogle Scholar
  25. Roman A, Munger K (2013) The papillomavirus E7 proteins. Virology 445(1–2):138–168.  https://doi.org/10.1016/j.virol.2013.04.013CrossRefPubMedPubMedCentralGoogle Scholar
  26. Romanczuk H, Howley PM (1992) Disruption of either the E1 or the E2 regulatory gene of human papillomavirus type 16 increases viral immortalization capacity. Proc Natl Acad Sci U S A 89(7):3159–3163CrossRefPubMedPubMedCentralGoogle Scholar
  27. Ronco G, Dillner J, Elfström KM, Tunesi S, Snijders PJ, Arbyn M, Kitchener H, Segnan N, Gilham C, Giorgi-Rossi P, Berkhof J, Peto J, Meijer CJ, International HPV Screening Working Group (2014) Efficacy of HPV-based screening for prevention of invasive cervical cancer: follow-up of four European randomised controlled trials. Lancet 383(9916):524–532.  https://doi.org/10.1016/S0140-6736(13)62218-7CrossRefPubMedGoogle Scholar
  28. Rosestedt M, Andersson KG, Mitran B, Tolmachev V, Löfblom J, Orlova A, Ståhl S (2015) Affibody-mediated PET imaging of HER3 expression in malignant tumours. Sci Rep 5:15226.  https://doi.org/10.1038/srep15226CrossRefPubMedPubMedCentralGoogle Scholar
  29. Sandström M, Lindskog K, Velikyan I, Wennbor A, Feldwisch J, Sandberg D, Tolmachev V, Orlova A, Sörensen J, Carlsson J, Lindman H, Lubberink M (2016) Biodistribution and radiation dosimetry of the anti-HER2 Affibody molecule 68Ga-ABY-025 in breast cancer patients. J Nucl Med 57(6):867–871.  https://doi.org/10.2967/jnumed.115.169342CrossRefPubMedGoogle Scholar
  30. Saslow D, Solomon D, Lawson HW, Killackey M, Kulasingam SL, Cain J, Garcia FA, Moriarty AT, Waxman AG, Wilbur DC, Wentzensen N, Downs LS Jr, Spitzer M, Moscicki AB, Franco EL, Stoler MH, Schiffman M, Castle PE, Myers ER, ACS-ASCCP-ASCP Cervical Cancer Guideline Committee, American Cancer Society, American Society for Colposcopy and Cervical Pathology, American Society for Clinical Pathology (2012) American Society for Clinical Pathology screening guidelines for the prevention and early detection of cervical cancer. CA Cancer J Clin 62(3):147–172.  https://doi.org/10.3322/caac.21139CrossRefPubMedPubMedCentralGoogle Scholar
  31. Scheffner M, Werness BA, Huibregtse JM, Levine AJ, Howley PM (1990) The E6 oncoprotein encoded by human papillomavirus types 16 and 18 promotes the degradation of p53. Cell 63(6):1129–1136CrossRefPubMedGoogle Scholar
  32. Schoffelen R, Sharkey RM, Goldenberg DM, Franssen G, Mcbride WJ, Rossi EAChang CH, Laverman P, Disselhorst JA, Eek A, van der Graaf WT, Oyen WJ, Boerman OC (2010) Pretargeted immuno-positron emission tomography imaging of carcinoembryonic antigen-expressing tumors with a bispecific antibody and a 68Ga- and 18F-labeled hapten peptide in mice with human tumor xenografts. Mol Cancer Ther 9(4):1019–1027.  https://doi.org/10.1158/1535-7163.MCT-09-0862CrossRefPubMedPubMedCentralGoogle Scholar
  33. Schwarz E, Freese UK, Gissmann L, Mayer W, Roggenbuck B, Stremlau A, zur Hausen H (1985) Structure and transcription of human papillomavirus sequences in cervical carcinoma cells. Nature 314(6006):111–114
  34. Sörensen J, Sandberg D, SandströM M, Wennborg A, Feldwisch J, Tolmachev V, Åström G, Lubberink M, Garske-Román U, Carlsson J, Lindman H (2014) First-in- human molecular imaging of HER2 expression in breast cancer metastases using the 111In-ABY-025 affibody molecule. J Nucl Med 55(5):730–735.  https://doi.org/10.2967/jnumed.113.131243CrossRefPubMedGoogle Scholar
  35. Sörensen J, Velikyan I, Sandberg D, Wennborg A, Feldwisch J, Tolmachev V, Orlova A, Sandström M, Lubberink M, Olofsson H, Carlsson J, Lindman H (2016) Measuring HER2-receptor expression in metastatic breast cancer using [68Ga]ABY-025 Affibody PET/CT. Theranostics 6(2):262–271.  https://doi.org/10.7150/thno.13502CrossRefPubMedPubMedCentralGoogle Scholar
  36. Ståhl S, Gräslund T, Eriksson KA, Frejd FY, Nygren PÅ, Löfblom J (2017) Affibody molecules in biotechnological and medical applications. Trends Biotechnol 35(8):691–712.  https://doi.org/10.1016/j.tibtech.2017.04.007CrossRefPubMedGoogle Scholar

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