Cloning and expression of the V-domain of the CD166 in prokaryotic host cell

Hassan Dana, Vahid Marmari, Ali Mazraeh, Ali Ghamari, Mohammad Mahdi Forghanifard


Purpose: CD166/ALCAM (Activated leukocyte cell adhesion molecule) as an immunoglobulin is implicated in cell migration. It is also involved in tumorigenesis of CRC (colorectal cancer) and known as a cancer stem cell marker. CD166, as a membrane protein, potentially represents either diagnostic or therapeutic capacities for CRC.

Methods: In this study, the sequence of V domain was optimized for expression in prokaryotic host using online tools and cloned into pET-28a plasmid. The recombinant pET28a was transformed into the E. coli BL21DE3 using heat shock method and expression of recombinant V domain was examined using SDS-PAGE (sodium dodecyl sulfate polyacrylamide gel electrophoresis).

Results: The results confirmed protein expression of recombinant 22.77 kDa V domains in bacterial expression system.

Conclusion: V domain of the CD166 was expressed successfully in E. coli bacteria. This recombinant fragment can be introduced as a suitable diagnostic and therapeutic candidate for screening and cancer-therapy of CRC patients, respectively. 


Cloning and Expression, ALCAM / CD166, V domain, Recombinant Protein, Colorectal Cancer

Full Text:



King JA, Ofori-Acquah SF, Stevens T, et al. Activated leukocyte cell adhesion molecule in breast cancer: prognostic indicator. Breast Cancer Research. 2004;6:478-87.

Masedunskas A, King JA, Tan F, et al. Activated leukocyte cell adhesion molecule is a component of the endothelial junction involved in transendothelial monocyte migration. Federation of European Biochemical Societies. 2006;580(11):2637-45

Wang J, Gu Z, Ni P, et al. NF-kappaB P50/P65 hetero-dimer mediates differential regulation of CD166/ALCAM expression via interaction with micoRNA-9 after serum deprivation, providing evidence for a novel negative auto-regulatory loop. Nucleic Acids Res. 2011;39(15):6440-55.

Klonisch T, Wiechec E, Hombach-Klonisch S, et al. Cancer stem cell markers in common cancers – therapeutic implications. Trends Mol Med. 2008;14(10):450-60.

Dalerba P, Dylla S, Park I, et al. Phenotypic characterization of human colorectal cancer stem. Cell pans. 2007;24:10158-63.

Bowen MA, Pate DD, Li X, et al. Cloning, mapping, and characterization of Activated Leukocyte-Cell Adhesion Molecule (ALCAM), a CD6 Ligand. J Exp Med. 1995;181(6):2213-20.

Bowen MA, Bajorath J, Siadak AW, et al. The Amino-terminal Immunoglobulin-like Domain of Activated Leukocyte Cell Adhesion Molecule binds specifically to the membrane-proximal scavenger receptor cysteine-rich domain of CD6 with a 1:1 stoichiometry. J Biol Chem. 1996;271(29):17390-6.

Weidle UH, Eggle D, Klostermann S, et al. ALCAM/CD166: Cancer-related Issues. Cancer genomics & proteomics. 2010;7:231-44.

Wiiger MT, Gehrken HB, Fodstad Q, et al. A novel human recombinant single-chain antibody targeting CD166/ALCAM inhibits cancer cell invasion in vitro and in vivo tumour growth. Cancer Immunol Immunother. 2010;59:1665–74.

Bowen M.A, Aruffo A. Adhesion molecules, their receptors, and their regulation: Analysis of CD6-Activated Leukocyte Cell Adhesion Molecule (ALCAM/CD166) Interactions. Transplant Proc. 1999;31(1-2):795-6.

Ofori-Acquah SF, King JA. Activated leukocyte cell adhesion molecule: a new paradox in cancer. Translational Research. 2008;151:122–28.

Fanali C, Lucchetti D, Farina M, et al. Cancer stem cells in colorectal cancer from pathogenesis to therapy: Controversies and perspectives. World J Gastroentero. 2014;20(4):923-42.

Bowen MA, Aruffo AA, Bajorath J. Cell surface receptors and their ligands: In Vitro analysis of CD6-CD166 interactions. PROTEINS: structure, function, and genetics. 2000;40:420–28.

Skonier JE, Bowen MA, Aruffo A, et al. CD6 recognizes the neural adhesion molecule BEN. Protein Science. 1997;6:1768-70.

Swart GW. Activated leukocyte cell adhesion molecule (CD166/ALCAM): Developmental and mechanistic aspects of cell clustering and cell migration. European Journal of Cell Biology. 2002;81:313–21.

Tachezy M, Zander H, Gebauer F, et al. Activated leukocyte cell adhesion molecule (CD166)-Its prognostic power for colorectal cancer patients. Journal of Surgical Research. 2012;177:15-20.

Pitule P, Čedíková M, Třeška V, et al. Assessing colorectal cancer heterogeneity: one step closer to tailored medicine. Journal of Applied Biomedicine. 2013;11:115–29.

Park JH, McMillan DC, Horgan PG, et al. The impact of anti-inflammatory agents on the outcome of patients with colorectal cancer. Cancer Treatment Reviews. 2014;40:68–77.

Mejia A, Schulz S, Hyslop T, et al. Molecular staging individualizing cancer management. J Surg Oncol. 2012;105(5):468–74.

Rohani Rasaf M, Ramezani R, Mehrazma M, et al. Inequalities in cancer distribution in Tehran; A disaggregated estimation of 2007 incidence by 22 districts. International Journal of Preventive Medicine. 2012;3:483-92.

Ouakrim DA, Lockett T, Boussioutas A, et al. Screening participation predictors for people at familial risk of colorectal cancer a systematic review. American Journal of Preventive Medicine. 2013;44(5):496–506.

Kim E, Coelho D, Blachier F. Review of the association between meat consumption and risk of colorectal cancer. Nutrition Research. 2013;33:983–94.

Moshfeghi K, Mohammad-Beigi A, Hamedi-Sanani D, et al. Evaluation the role of nutritional and individual factors in colorectal cancer. Zahedan J Res Med Sci. 2011;13(4):12-7.

Darabi P, Galehdari H, Khatami SR , et al. Codon optimization, cloning and expression of the human Leukemia Inhibitory Factor (hLIF) in E. Coli. Iran J Biotech. 2013;11(1):47-53.

Ni C, Zhang Z, Zhu X, et al. Prognostic value of CD166 expression in cancers of the digestive system: a systematic review and meta-analysis. PLoS One. 2013;5:8(8):e70958.

Wang F, Scoville D, He XC, et al. Isolation and Characterization of intestinal stem cells based on surface marker combinations and colony-formation assay. Gastroenterology. 2013;145(2):383–95.

Vaiopoulos A G, Kostakis I D, Koutsilleris M, et al. Concise review: colorectal cancer Stem Cells. Stem Cells. 2012;30:363–71.

Sanders MA, Majumdar APN. Colon cancer stem cells: implications in carcinogenesis. Front Biosci (Landmark Ed). 2014;16:1651–62.

Liu AY, Roudier MP, True LD. Heterogeneity in primary and metastatic prostate cancer as defined by cell surface CD profile. American Journal of Pathology. 2004;165:1543-56.

Dana H, Marmari V, Mahmoodi G, et al. CD166 as a Stem Cell Marker? A potential target for therapy colorectal cancer? J Stem Cell Res Ther. 2016;1(6):00041.

Levin TG, Powell AE, Davies PS, et al. Characterization of the intestinal cancer Stem Cell Marker CD166 in the human and mouse gastrointestinal tract. Gastroenterology. 2010;139:2072–82.

Shiozawa Y, Nie B, Pienta KJ, et al. Cancer stem cells and their role in metastasis. Pharmacology & Therapeutics. 2013;138:285–93.

Botchkina C. Colon cancer stem cells – From basic to clinical application. Cancer Letters. 2013;338:127–40.

Dana H, Mahmoodi G, Marmari V, et al. An overview of cancer Stem Cell. J Stem Cell Res Ther. 2016;1(4):00029.

Noori Daloii MR, Fazilaty H, Tabrizi M. Cancer metastasis, genetic and micro environmental factors of distant tissue: a review article. Tehran Univ Med J(TUMJ). 2013; 70(11):671-83.

Noh KH, Lee Y-H, Jeon J-H, et al. Cancer vaccination drives Nanog-dependent evolution of tumor cells towards an immune-resistant and stem-like phenotype. Cancer Research. 2012;72(7):1717-27.

Jager E, Jager D, Knuth A. Antigen-specific immunotherapy and cancer vaccines. Int J Cancer. 2003;106:817–20.


Creative Commons License
This work is licensed under a Creative Commons Attribution 3.0 License.


International Journal of Cancer Therapy and Oncology (ISSN 2330-4049)

© International Journal of Cancer Therapy and Oncology (IJCTO)

To make sure that you can receive messages from us, please add the '' domain to your e-mail 'safe list'. If you do not receive e-mail in your 'inbox', check your 'bulk mail' or 'junk mail' folders.


Number of visits since October, 2013