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Progressive growth of metastatic Lewis lung carcinoma (LLC-LN7) tumors is associated with increased levels of bone-marrow-derived CD34+ cells having natural suppressor (NS) activity toward T cells. The present studies determined whether tumor-derived products are responsible for this induction of NS activity. Culturing normal bone marrow cells with LLC-LN7-conditioned medium (LLC-CM) or with recombinant granulocyte/macrophage-colony-stimulating factor (GM-CSF) resulted in the appearance of NS activity. The development of NS activity coincided with a prominent increase in the levels of CD34+ cells. That the CD34+ cells were responsible for the NS activity of the bone marrow cultures containing LLC-CM was shown by the loss of NS activity when CD34+ cells were depleted. The stimulation of CD34+ NS cells by LLC-CM was attributed to tumor production of GM-CSF, since neutralization of GM-CSF within the LLC-CM reduced its capacity to increase CD34+ cell levels. Studies also showed that the induction of CD34+ NS cells by LLC-CM and GM-CSF could be overcome by including in the cultures an inducer of myeloid differentiation, 1alpha,25-dihydroxyvitamin D3 [1,25(OH)2D3]. These results demonstrate that the mechanism by which the LLC-LN7 tumors stimulate increased levels of CD34+ NS cells from normal bone marrow is by their production of GM-CSF and that this can be blocked with the myeloid differentiation inducer 1,25(OH)2D3.

The sterol 1alpha,25-dihydroxyvitamin D3 [1,25(OH)2D3] can inhibit T cell activation as well as restore the functional competence of suppressed T cells, The present studies determined whether 1,25(OH)2D3 had a differential effect on the activation of normal T cells or of suppressed T cells from mice bearing Lewis lung carcinoma tumors. Normal spleen cell proliferation in response to immobilized anti-CD3 was unaffected by the lower doses of 0.1-10 nM 1,25(OH)2D3, and was inhibited by the higher dose of 100 nM 1,25(OH)2D3. In contrast, 1,25(OH)2D3 increased proliferation and interferon gamma secretion by T cells of tumor bearers in response to stimulation through T cell receptor/CD3. Assessment of mechanisms associated with the 1,25(OH)2D3 stimulation of tumor-bearer T cells implicated protein phosphatase 2A (PP-2A). First, PP-2A activity of spleen cells from tumor bearers was reduced compared to that of normal spleen cells but was increased by 1,25(OH)2D3. Second, 1,25(OH)2D3 stimulation of tumor-bearer T cell proliferation was dependent on this PP-2A activity as it was blocked by doses of okadaic acid that selectively inhibit PP-2A. These results suggest that 1,25(OH)2D3 preferentially enhances the responsiveness of immunosuppressed T cells from tumor bearers to TCR/CD3 stimulation by restoring PP-2A activity.