Stephen G. Emerson

Interferon-alpha restores the deficient expression of the cytoadhesion molecule lymphocyte function antigen-3 by chronic myelogenous leukemia progenitor cells

Hematopoietic cells from the malignant clone in chronic myelogenous leukemia (CML) maintain and expand a proliferative advantage over normal hematopoietic cells within the bone marrow. This advantage is often ameliorated or reversed in vivo by IFN alpha. Based upon earlier studies suggesting decreased adhesiveness of CML progenitor cells, we asked whether CML progenitor cells are deficient in their expression of the cytoadhesion molecule lymphocyte function antigen-3 (LFA-3, CD58) which is normally expressed on hematopoietic progenitors. Progenitor cells from untreated CML patients showed greatly reduced or absent LFA-3 expression, whereas progenitors from patients treated with IFN alpha in vivo or in vitro expressed surface LFA-3 at more normal levels. LFA-3-deficient CML progenitor cells were unable to stimulate normal regulatory proliferative responses in autologous T cells. We hypothesize that IFN alpha-sensitive LFA-3 deficiency reflects a cell surface cytoadhesion defect which may help explain adhesive abnormalities of CML progenitor cells in vitro and clonal proliferation in vivo.

Synergistic regulation of human megakaryocyte development.

Little information exists concerning differing levels of regulation occurring during human megakaryocyte development. We hypothesize that megakaryocytic proliferation and maturation is controlled by two, synergistic regulatory factors. One, megakaryocyte colony-stimulating activity, is an obligate requirement for colony formation and drives the development of relatively immature cells. Megakaryocyte colony-stimulating activity is a functional component of the human recombinant proteins, interleukin 3 or GM-CSF. Human recombinant growth factors, interleukin 1, interleukin 6, or crythropoietin, do not effect megakaryocyte development either alone or in combination with interleukin 3. Full maturation requires a second synergistic activity which increases megakaryocyte number, size, and cytoplasmic and antigenic content. In culture, this synergistic regulator augments maturation by increasing the number of colonies, colony cellularity, and size. In suspension cultures, this cofactor increases megakaryocyte cytoplasmic and antigenic content, and shifts the morphological distribution from immature to mature megakaryocytes. Finally, this activity also increases the number of antigen positive megakaryocytes, either by stimulating proliferation or conversion of antigen-negative to antigen-positive cells. Comparative studies of megakaryocytic regulation suggests that this in vitro regulator mimicks some of the known effects of thrombopoietin in vivo.

Recombinant human granulocyte-colony-stimulating factor in the treatment of patients with neutropenia

The results of an open-label, randomized, Phase III trial of r-methionyl human granulocyte-colony-stimulating factor (r-metHuG-CSF) in 41 patients with severe chronic neutropenia (SCN) are reported. Patients with diagnoses of congenital, cyclic, and idiopathic neutropenia, with histories of recurrent infections, were evaluated. The primary objective of the trial was to evaluate the ability of r-metHuG-CSF to increase the ANC to greater than 1500/mm3. A secondary objective was to evaluate variables associated with infection-related morbidity in SCN. r-metHuG-CSF treatment consisted of 1 month of dose titration followed by 4 months of treatment at an optimal dose. Patients were randomized to either immediate treatment with r-metHuG-CSF (Group A) or four months of observation followed by r-metHuG-CSF treatment (Group B). r-metHuG-CSF was administered by daily, subcutaneous injection with initial doses of 3 to 10 micrograms/kg/day. Forty of 41 patients who received r-metHuG-CSF had a complete response (median ANC greater than 1500/mm3 during 4 months of r-metHuG-CSF treatment). All cases of gingivitis and severe mouth ulcers resolved upon treatment with r-metHuG-CSF. Serious infections were also eliminated. Only one patient failed to show clinical improvement in response to r-metHuG-CSF treatment. Adverse reactions during the first 5 months of treatment were mild. Splenomegaly (mild) was noted in some patients. The administration of r-metHuG-CSF in patients with SCN significantly increased the ANC (P less than 0.001) and was accompanied by a marked reduction in infectious complications.

Acute lymphoblastic leukemia with eosinophilia

Acute lymphoblastic leukemia with eosinophilia is a rare but distinctive clinical entity. The eosinophilia in these patients can present before, concomitantly, or after the diagnosis of leukemia. Patients with this syndrome often suffer from the cardiovascular complications of severe eosinophilia, suffering excess morbidity and mortality as a result of their eosinophilia. Treatment of the eosinophilia in this syndrome consists of administration of induction chemotherapy, followed by prednisone and hydroxyurea if required for persistent eosinophilia. Eosinophilia often resolves with remission of leukemia, only to return at the time of relapse in a high percentage of cases. Patients with this syndrome characteristically have cytogenetic abnormalities involving the long arms of chromosomes 5 and 14. These cytogenetic abnormalities are not commonly seen in acute lymphoblastic leukemia and suggest that this syndrome may have a distinct pathophysiology and etiology. The affected region on chromosome 5 contains genes that control hematopoiesis, including eosinophilopoiesis. Further investigations into these cytogenetic abnormalities may provide insight into the etiology of the leukemia and eosinophilia characteristic of this syndrome.

Regulation of cytotoxic T lymphocyte-mediated graft rejection following bone marrow transplantation.

Cytotoxic T lymphocytes have been implicated as the effector cell mediating graft rejection following human allogeneic bone marrow transplantation. We have studied a BMT patient who rejected haploidentical T cell-depleted marrow. In vitro studies demonstrated that the circulating lymphocytes were CD3+ and CD8+, of recipient origin, and exhibited selective cytotoxicity against donor-specific class I major histocompatibility complex antigens. Cytotoxicity was inhibited by monoclonal antibodies directed against CD3, CD8, CD2, and lymphocyte function-associated antigen-1 on the T cell, and against MHC class I proteins on the target cell. Furthermore, these circulating cells inhibited the in vitro growth and differentiation of enriched donor bone marrow progenitor cells, an inhibition that was partially reversed by anti-CD3 MAb. Donor-specific recipient-derived CTL may mediate resistance to engraftment, and CTL activity may be inhibited by a number of MAb. The implications of these findings for host preparation and treatment are discussed.

The influence of extra-cellular matrix and stroma remodeling on the productivity of long-term human bone marrow cultures

The stromal cell layer is believed to play an important role in long-term human bone marrow cultures (LTHBMCs). At present, neither the role that the stromal cell extra-cellular matrix (ECM) plays in influencing stroma behavior is well understood nor are the effects of stroma aging. Rapid medium exchanged LTHBMCs were established on surfaces precoated with human natural fibronectin and type 1 rat tail collagen. Although initial adhesion of hematopoietic cells was improved by the presence of both ECMs, the overall progenitor and nonadherent cell productivity was not improved nor did the stroma grow to confluency faster. Thus, the ECMs used did not significantly influence the cell productivity of LTHBMCs. To examine the influence of stromal cell layer aging, conditioned medium was obtained from the first two weeks of LTHBMCs that was subsequently concentrated and used as a medium supplement in a second set of slowly exchanged LTHBMCs. The presence of the concentrated conditioned medium (conCM) enhanced the production of nonadherent cells three-fold compared with control over an eight week culture period. Control cultures that were exposed to conCM after 4 weeks in culture significantly improved their cell productivity during the latter 4 weeks of culture compared with control. The productivity of cultures exposed to conCM for 4 weeks dropped significantly when unsupplemented medium was used for the latter 4 weeks of culture. Interestingly, phytohemagglutin-stimulated leukocyte-conditioned medium stimulated LTHMBCs in a similar fashion, as did conditioned medium from early LTHBMCs. Taken together, these results strongly suggest that the stromal cell layer does produce important factors for active hematopoiesis during its growth to confluence.