Therapy of human tumors in NOD/SCID mice with patient-derived reactivated memory T cells from bone marrow.

In an analysis of 84 primary-operated breast cancer patients and 11 healthy donors, we found that the bone marrow of most patients contained memory T cells with specificity for tumor-associated antigens. Patients' bone marrow and peripheral blood contained CD8+ T cells that specifically bound HLA/peptide tetramers. In short-term culture with autologous dendritic cells pre-pulsed with tumor lysates, patients' memory T cells from bone marrow (but not peripheral blood) could be specifically reactivated to interferon-gamma-producing and cytotoxic effector cells. A single transfer of restimulated bone-marrow T cells into NOD/SCID mice caused regression of autologous tumor xenotransplants associated with infiltration by human T cells and tumor-cell apoptosis and necrosis. T cells from peripheral blood showed much lower anti-tumor reactivity. Our findings reveal an innate, specific recognition of breast cancer antigens and point to a possible novel cancer therapy using patients' bone-marrow-derived memory T cells.

Generation of dendritic cells from human bone marrow mononuclear cells: advantages for clinical application in comparison to peripheral blood monocyte derived cells.

Dendritic cells (DCs) currently used for vaccination in clinical studies to induce immunity against malignant cells are normally generated from peripheral blood-derived monocytes. Here we studied conditions for the generation of DCs from unseparated human bone marrow (BM) mononuclear cells and compared them functionally with DCs from blood. The two types of DCs, from bone marrow (BM-DC) and peripheral blood (BL-DC), were generated in parallel from the same normal healthy donors by culturing in serum-free X-VIVO 20 medium containing GM-CSF and IL-4, and then the phenotypes and functions were compared. BM-DC generation occurred in 14 days and involved proliferative expansion from CD34 stem cells and differentiation while BL-DC generation occurred in 7 days from CD14 monocytes and involved only differentiation. A 7- to 25-fold higher number of DCs could be obtained from BM than from blood. BM-DC had similar phenotypes as BL-DC. The capacity to stimulate MLR reactivity in allogeneic T lymphocytes was higher with BM-DC than that with BL-DC. Also, the capacity to stimulate autologous memory T cell responses to tetanus toxoid (TT) or tuberculin (PPD) was higher with BM-DC than with BL-DC. These results suggest that BM-DC as produced here may be a very economic and useful source of professional antigen-presenting cells for anti-tumor immunotherapeutic protocols.

Enrichment of memory T cells and other profound immunological changes in the bone marrow from untreated breast cancer patients.

Previous studies with animal tumors showed that bone marrow (BM) is a privileged site where potentially lethal tumor cells are controlled in a dormant state by the immune system. Here, we investigated BM of breast cancer patients with respect to tumor cell content, immune activation status and memory T-cell content. BM-derived cells from primary operated breast cancer patients (n = 90) were compared with those from healthy donors (n = 10) and also with cells from respective blood samples. Cytokeratin 19-positive tumor cells were detected by nested polymerase chain reaction. Three-color flow cytometry was used to identify numbers and activation state of T cells, natural killer (NK) cells, monocytes/macrophages and subsets by a panel of monoclonal antibodies (mAbs). The proportion of memory T cells among the CD4 and CD8 T cells was much higher in BM of cancer patients than in healthy donors (p < 0.001). The extent of memory T-cell increase was related to the size of the primary tumor. Patient-derived BM memory CD8 T cells could be shown to contain specific HLA-A2/Her-2/neu(369-377) tetramer binding cells. Patients with disseminated tumor cells in their BM had more memory CD4 T cells and more CD56(+) CD8(+) cells than patients with tumor cell-negative BM. Only some of the immunological changes seen in BM samples of cancer patients were also detectable in peripheral blood samples. Our hypothesis that BM is a special compartment for immunological memory and tumor dormancy is supported by the above findings. The overall results reveal that BM is a valuable additional compartment for immune diagnosis in pathological conditions and possibly for follow-up treatment strategies.