Association of a characteristic membrane pattern of annexin A2 with high invasiveness and nodal status in colon adenocarcinoma ELENA TRISTANTE, CARLOS M. MART  INEZ, SOF  IA JIM  ENEZ, LUCIA MORA, FERNANDO CARBALLO, ISABEL MART  INEZ-LACACI, and CARLOS DE TORRE-MINGUELA MURCIA AND ELCHE (ALICANTE), SPAIN The identification of tumor cells in lymph nodes is essential for the correct classifica- tion of patients with colorectal cancer who may benefit from adjuvant treatments. Proper classification of tumor stage becomes entangled by variables such as an insufficient number of lymph nodes examined, which can result in erroneous or missed diagnosis. The determination of pathologic factors in the primary tumor asso- ciated with positive lymph nodes is an area of research that has attempted to provide variables to solve this problem. In the present study, we observed that the localization of annexin A2 (AnxA2) in a cell membrane is the characteristic that distinguishes tumor cells with high invasiveness. Localization of AnxA2 expression was also studied in tissue specimens from 58 patients with invasive colorectal carcinoma (T3–T4), who had undergone colectomy with radical lymph node dissection. Interestingly, the membrane pattern observed in tumor cell lines was also identified in patient’s tissue samples and allowed us to distinguish among different cell populations with the tumor. Univariate analysis showed that tumor deposits in pericolic fat, extramural vascular invasion, and amount of cells with AnxA2 membrane pattern in the tumor invasive edge had a significant influence on lymph node metastasis. On the con- trary, multivariate analysis revealed that the number of cells with AnxA2 membrane pattern (P , 0.05) and tumor deposits (P , 0.05) was significantly associated with lymph node metastasis. Furthermore, AnxA2 cellular localization was observed in cell clusters that define tumor budding, and a significant association between both variables was detected. (Translational Research 2015;166:196–206) Abbreviations: CHAPS ¼ 3-[(3-cholamidopropyl)dimethylammonio]-1-propanesulfonate; DIGE ¼ Difference gel electrophoresis; DTT ¼ Dithiothreitol; ECL ¼ Enhanced chemiluminescence; HSP90 ¼ Heat shock protein 90; MS/MS ¼ Tandem mass spectrometry; PAGE ¼ Polyacrylamide gel electrophoresis; PMSF ¼ Phenylmethylsulfonyl fluoride; PVDF ¼ Polyvinylidene difluoride; SDS ¼ Sodium dodecyl sulfate From the Unidad AECC de Investigaci on Traslacional en C ancer, Hospital Cl  ınico Universitario Virgen de la Arrixaca (HCUVA), Murcia, Spain; Grupo Cirug  ıa Experimental, CIBERehd, Hospital Cl  ınico Universitario Virgen de la Arrixaca (HCUVA), IMIB-Arrixaca, Murcia, Spain; Unidad de Prote omica, Hospital Cl  ınico Universitario Virgen de la Arrixaca (HCUVA), IMIB-Arrixaca, Murcia, Spain; Servicio de Gastroenterolog  ıa, Hospital Cl  ınico Universitario Virgen de la Arrixaca (HCUVA), IMIB-Arrixaca, Murcia, Spain; Instituto de Biolog  ıa Molecular y Celular, Universidad Miguel Hern andez, Elche (Alicante), Spain. Elena Tristante and Carlos M. Mart  ınez contributed equally to this article. Submitted for publication October 2, 2014; revision submitted February 12, 2015; accepted for publication February 26, 2015. Reprint requests: Carlos de Torre-Minguela, Unidad de Prote omica, Hospital Cl  ınico Universitario Virgen de la Arrixaca (HCUVA), IMIB-Arrixaca, Ctra de Madrid-Cartagena s/n, 30120 Murcia, Spain; e-mail: carlos.de3@um.es. 1931-5244/$ - see front matter Ó 2015 Elsevier Inc. All rights reserved. http://dx.doi.org/10.1016/j.trsl.2015.02.006 196