Tami Katz

Vaccination with Dendritic Cell/Tumor Fusions following Autologous Stem Cell Transplant Induces Immunologic and Clinical Responses in Multiple Myeloma Patients

Purpose: A multiple myeloma vaccine has been developed whereby patient-derived tumor cells are fused with autologous dendritic cells, creating a hybridoma that stimulates a broad antitumor response. We report on the results of a phase II trial in which patients underwent vaccination following autologous stem cell transplantation (ASCT) to target minimal residual disease. Experimental Design: Twenty-four patients received serial vaccinations with dendritic cell/myeloma fusion cells following posttransplant hematopoietic recovery. A second cohort of 12 patients received a pretransplant vaccine followed by posttransplant vaccinations. Dendritic cells generated from adherent mononuclear cells cultured with granulocyte macrophage colony-stimulating factor, interleukin-4, and TNF-α were fused with autologous bone marrow–derived myeloma fusion cells using polyethylene glycol. Fusion cells were quantified by determining the percentage of cells that coexpress dendritic cell and myeloma fusion antigens. Results: The posttransplant period was associated with reduction in general measures of cellular immunity; however, an increase in CD4 and CD8+ myeloma-specific T cells was observed after ASCT that was significantly expanded following posttransplant vaccination. Seventy-eight percent of patients achieved a best response of complete response (CR)+very good partial response (VGPR) and 47% achieved a CR/near CR (nCR). Remarkably, 24% of patients who achieved a partial response following transplant were converted to CR/nCR after vaccination and at more than 3 months posttransplant, consistent with a vaccine-mediated effect on residual disease. Conclusions: The posttransplant period for patients with multiple myeloma provides a unique platform for cellular immunotherapy in which vaccination with dendritic cell/myeloma fusion fusions resulted in the marked expansion of myeloma-specific T cells and cytoreduction of minimal residual disease. Clin Cancer Res; 19(13); 3640–8. ©2013 AACR.

Soluble plasma HLA peptidome as a potential source for cancer biomarkers

The HLA molecules are membrane-bound transporters that carry peptides from the cytoplasm to the cell surface for surveillance by circulating T lymphocytes. Although low levels of soluble HLA molecules (sHLA) are normally released into the blood, many types of tumor cells release larger amounts of these sHLA molecules, presumably to counter immune surveillance by T cells. Here we demonstrate that these sHLA molecules are still bound with their authentic peptide repertoires, similar to those of the membranal HLA molecules (mHLA). Therefore, a single immunoaffinity purification of the plasma sHLA molecules, starting with a few milliliters of patients’ blood, allows for identification of very large sHLA peptidomes by mass spectrometry, forming a foundation for development of a simple and universal blood-based cancer diagnosis. The new methodology was validated using plasma and tumor cells of multiple-myeloma and leukemia patients, plasma of healthy controls, and with cultured cancer cells. The analyses identified thousands of sHLA peptides, including some cancer-related peptides, present among the sHLA peptidomes of the cancer patients. Furthermore, because the HLA peptides are the degradation products of the cellular proteins, this sHLA peptidomics approach opens the way for investigation of the patterns of protein synthesis and degradation within the tumor cells.

Granulocyte colony-stimulating factor administration upregulates telomerase activity in CD34+ haematopoietic cells and may prevent telomere attrition after chemotherapy

Summary. Hematopoietic reconstitution could be associated with premature ageing of the transplanted cells and a high frequency of myelodysplastic syndrome and secondary leukaemia. Telomere length decreases with cell divisions and age, and at a crucial length it is associated with chromosomal instability and cell senescence. Telomerase is a reverse transcriptase enzyme that adds nucleotides to chromosomal ends. Most somatic cells lack telomerase activity yet haematopoietic stem cells retain low levels of telomerase. Some studies have found that chemotherapy and stem cell transplantation lead to the accelerated shortening of telomere length. As granulocyte colony‐stimulating factor (G‐CSF) is routinely used in the mobilization of stem cells for transplantation, we evaluated its effects on telomerase activity and regulation, and on telomere dynamics, in normal donors and selected lymphoma patients. Administration of G‐CSF increased telomerase activity in CD34+ haematopoietic cells compared with controls. In marrow‐derived CD34+ cells, telomerase activity increased sevenfold, compared with a 14‐fold increase in peripheral‐blood‐mobilized CD34+ cells. A parallel increase in the expression of human telomerase enzyme reverse transcriptase RNA and protein kinase C α occurred. In addition, G‐CSF administration to five lymphoma patients after consecutive courses of CHOP (cyclophosphamide, doxorubicin, vincristine, prednisone) chemotherapy, resulted in telomere length preservation or elongation, as opposed to marked attrition in patients who did not receive growth factors. We conclude that the in vivo administration of G‐CSF prevents or attenuates telomere attrition associated with chemotherapy administration. This attenuation may contribute to the preservation of telomere integrity inG‐CSF‐primed transplanted stem cells.

Successful haploidentical bone marrow transplantation in Fanconi anemia

A 10-year-old girl with Fanconi anemia and severe aplastic anemia underwent a haploidentical BMT from her mother due to lack of a matched family donor. T cell depletion was done by positive selection of CD34 cells with immunomagnetic beads. Due to graft rejection a second haploidentical BMT from the father was successfully undertaken. No immunosuppression was given after the transplant. Immunological reconstitution took approximately 6 months, with no GVHD or severe infections. Such a transplant, containing a large purified CD34 cell fraction with a minimal number of added T cells, should be considered as the treatment of choice for patients with Fanconi anemia if no HLA matched donor is available. Bone Marrow Transplantation (2000) 26, 1221–1223.

Contamination of peripheral hematopoeitic stem cell products with a Mycobacterium mucogenicum-related pathogen.

A gram-positive rod with a restriction pattern closely related to the published nucleotide sequence of Mycobacterium mucogenicum was isolated from 6 of 45 units of peripheral blood stem cell products. The source of the contamination was traced to ice cubes used in processing the peripheral blood stem cell products. Substituting reusable ice trays for ice from an ice machine terminated the outbreak.

Chronic lymphocytic leukemia cells acquire regulatory B-cell properties in response to TLR9 and CD40 activation.

Circulating chronic lymphocytic leukemia (CLL) cells share phenotypic features with certain subsets of regulatory B-cells (Bregs). The latter cells have been reported to negatively regulate immune cell responses, mostly by provision of IL-10. The purpose of the current study was to identify and delineate Breg properties of CLL cells. B-cells and T-cells were obtained from the peripheral blood of untreated CLL patients diagnosed according to the 2008 Guidelines of the International Workshop on Chronic Lymphocytic Leukemia. Co-culture assays were used to examine the ability of CLL cells to suppress autologous T-cell immune responses. IL-10 potency of CLL cells was assessed following stimulation with activators of the toll-like receptor 9 (TLR9) or CD40 and was correlated with the inhibitory activity of the cells. TLR9-activated CLL cells were found to increase the frequency of CD4+CD25hiFOXp3+ regulatory T-cells (Tregs) and to inhibit autologous CD4+ T-cell proliferation. This signaling cascade proved to control IL-10 generation in CLL cells, which in turn promoted the inhibition of T-cell proliferation by CLL cells. However, CD40 activation of CLL cells, while exhibiting a similar ability to augment Treg frequency, did not either affect IL-10 generation or T-cell proliferation. In conclusion, CLL cells demonstrate a unique clonal quality of adopting Breg properties which promote modulation of T-cell characteristics. TLR9 appears to be a potent activator of regulatory abilities in CLL cells, possibly contributing to preferential immune escape of TLR9-responsive cells.