Generation and characterization of two human alpha/beta T cell clones. Recognizing autologous breast tumor cells through an HLA- and TCR/CD3-independent pathway.

Abstract

Cell-mediated immune response to breast tumor has only been marginally investigated. To gain insight into this issue we have developed two clones of distinct phenotype, CD3 + a/fl, CD4+, CD8-, CD16-, and CD3+ a/l, CD4-, CD8+, CD16-, respectively, from peripheral blood lymphocytes (PBL) of a breast cancer patient. These effectors, selected on the basis of their cytolytic activity against autologous tumor cells and lack of lysis on NK-sensitive cell lines, preferentially recognize autologous tumor cells. The two clones' cytotoxic activity, while inhibited by anti-LFA-1 mAb, could not be abolished by mAbs to CD3, to class I and class II MHC molecules, and by mAbs to molecules involved in T cell function (i.e., CD4, CD8, CD2).The molecular structure of the a and P T cell receptor chains of the two effector cells, confirmed their clonality and showed that, despite an overlapping killing pattern, they possess distinct TCR a and ,f chains. These findings demonstrate that breast tumor-specific CTL clones can be generated through current technology and that a a/p effector cell population operating through a HLA-unrestricted and TCR/CD3-independent pathway may be involved in the identification and killing of this tumor. (J.

The Cooperation between hMena Overexpression and HER2 Signalling in Breast Cancer

“…The DAL cell line was obtained from the ascitic fluid of a breast cancer patient [32]. MCF7-HER2 and MCF7-pcDNA3 stable transfectants were obtained by selecting MCF7 cells transfected with HER2-pcDNA3.1 (kindly provided by Dr. Oreste Segatto), and with the empty vector respectively, using G418 500 µg/ml (Invitrogen) in complete culture medium.…”

Molecular Cloning of hMena (ENAH) and Its Splice Variant hMena+11a: Epidermal Growth Factor Increases Their Expression and Stimulates hMena+11a Phosphorylation in Breast Cancer Cell Lines

“…The following cell lines were purchased from the American Type Culture Collection (Rockville, MD): MDAMB361, T47D, SKBr3, MCF7, BT474 (breast cancer), A427, Calu3 (lung cancer), SiHa, CaSki (cervical carcinoma), T98G, U87MG, and U373 (glioma). Other cell lines used were glioblastoma U251 (23); SBT and DAL, developed in our laboratory from the ascitic fluid of two breast cancer patients (24); normal human keratinocytes (NHK) and ADF astrocytoma, kindly provided by Drs. A. Venuti and G. Zupi (Regina Elena Cancer Institute, Rome, Italy), respectively; melanoma ME10538, provided by Dr. A. Anichini (Human Tumor Immunobiology Unit, Istituto Nazional per lo Studio e la Cura dei Tumori, Milan, Italy), and MAS melanoma, developed in our laboratory from a primary melanoma lesion.…”

Human mena protein, a serex‐defined antigen overexpressed in breast cancer eliciting both humoral and CD8+ T‐cell immune response

“…The MDAMB231cl2 was established by cloning the MDAMB231 cell line in soft agar, as previously described 15. SAB and DAL cell lines were developed from the ascitic fluid of 2 breast cancer patients16 and MAS from a primary melanoma lesion. Normal human epidermal melanocytes were purchased from Promo Cell (Heidelberg, Germany).…”

“…After 24 hr, IL‐2 (Roche Diagnostics, Mannheim, Germany) and IL‐7 (PeproTech, Rocky Hill, NJ; 2.5 ng/ml and 10 ng/ml, respectively) were added to the culture wells as previously reported 18. Cells were restimulated after 7 days with 1 μg/ml of peptide, then weekly with T2 cells pulsed with 40 μg/ml of peptide, and tested in cytotoxicity assay as described 16…”

“…The cytotoxic activity of the hMena‐502 D.P.N. cell line was determined using a conventional 4‐hr 51 Cr release assay as previously described 16. For peptide recognition, T2 cells were incubated overnight with 40 μg/ml peptide, washed, then labelled.…”