Patients with head and neck squamous cell carcinoma (HNSCC) have profound immune deficiencies. In 65% of these patients, there is an increased intra‐tumoral presence of immune‐suppressive CD34+ progenitor cells. The goal of the present study was to determine whether CD34+ cell levels were also increased in the peripheral blood of HNSCC patients and if these immune‐suppressive cells could be differentiated into dendritic cells. Our results showed that CD34+ cell levels are increased in the peripheral blood of HNSCC patients. To assess if these CD34+ cells could differentiate into dendritic cells, they were isolated from the blood of HNSCC patients and cultured for 12 days with various cytokine combinations. Culturing CD34+ cells with stem cell factor (c‐kit ligand) plus granulocyte‐macrophage colony‐stimulating factor resulted in the appearance of a significant proportion of cells expressing phenotypic markers characteristic of dendritic cells. Also, including tumor necrosis factor‐α yielded a significant proportion of cells resembling the bi‐potential precursor cells for dendritic cells and monocytes (CD14+CD1a+), in addition to the dendritic‐like cells. When the differentiation inducer 1α,25‐dihydroxyvitamin D3 [1,25(OH)2D3] was added along with the cytokine combinations, the yield of cells having characteristics of dendritic cells was further increased. Cells that were derived from CD34+ cell cultures containing 1,25(OH)2D3 had a more potent capacity to present the recall antigen tetanus toxoid to autologous peripheral blood leukocytes and to stimulate a mixed leukocyte response compared to cultures to which 1,25(OH)2D3 had not been added. Our results show that CD34+ cells, whose frequency is increased in HNSCC patients, can be differentiated into cells that phenotypically and functionally resemble dendritic cells and that 1,25(OH)2D3 accentuates this differentiation. Int. J. Cancer 73:663–669, 1997. © 1997 Wiley‐Liss, Inc.

Patients with head and neck squamous cell carcinoma (HNSCC) have profound defects in their immune defenses. Using immunofluorescent staining and flow cytometric analysis, we found that most patients with HNSCC have increased levels of CD34+ cells within their peripheral blood. These circulating CD34+ cells contribute to the depressed functional competence of the peripheral blood T-lymphocytes. This was demonstrated by the increased level of proliferative responsiveness to interleukin-2 by the patients' peripheral blood T-cells after depletion of CD34+ cells. These results show the importance of CD34+ cells in contributing to the depression of T-lymphocyte function in patients with HNSCC and suggest that strategies designed to reduce the levels of circulating CD34+ cells may enhance the immune reactivity of the patients' circulating T-lymphocytes against the HNSCC.

Within human head and neck squamous cell carcinomas (HNSCC) that produce granulocyte-macrophage CSF are CD34+ cells that exhibit natural suppressive (NS) activity. The present study aimed to identify how these NS cells mediate suppression and how to diminish their presence. CD34+ cells that were immunomagnetically isolated from fresh surgical HNSCC specimens produced a soluble product that blocked normal T cell stimulation through the TCR/CD3 complex. This inhibitory activity could be neutralized with Abs to TGF-beta1. Since prior studies showed that the CD34+ NS cells within HNSCC cancers are myelomonocytic progenitor cells, the feasibility of using cytokines that can induce myeloid cell differentiation to diminish the presence of CD34+ NS cells was tested. Adding low doses of 100 U/ml IFN-gamma plus 10 U/ml TNF-alpha to bulk cultures of dissociated HNSCC cancers diminished the frequency of CD34+ cells. Studies with CD34+ cells that were isolated from the HNSCC cancers showed that this cytokine treatment induced differentiation of the CD34+ cells predominantly into monocytic cells. The consequence of treating CD34+ NS cells with the myeloid differentiation treatment was the loss of suppressive activity, a decline in TGF-beta production, and the production of TNF-alpha by the resulting monocytic cells. In HNSCC bulk cultures containing high levels of CD34+ NS activity, IFN-gamma/TNF-alpha not only reduced CD34+ cell levels, but also increased the capacity of the intratumoral T cells to express the p55 IL-2R. These studies show that IFN-gamma/TNF-alpha can induce differentiation of TGF-beta-secreting CD34+ NS cells into nonsuppressive monocytic cells that secrete TNF-alpha.

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