Assessment of the therapeutic potential of cytokines, cytotoxic drugs and effector cell populations for the treatment of multiple myeloma using the 5T33 murine myeloma model.

The therapeutic potential of six cytokines, eight cytotoxic drugs and two effector cell populations for the treatment of multiple myeloma was assessed in vitro using the 5T33 murine myeloma model. The efficacy of combination IFN-alpha and melphalan therapy was also evaluated in vitro and in vivo. Of the cytokines tested in vitro using the MTT assay, only IFN-alpha demonstrated significant inhibition of myeloma cell growth at non-toxic concentrations (ED50 = 1508.3 +/- 181.3 U/mL and 2617.9 +/- 334.0 U/mL for murine IFN-alpha [mIFN-alpha] and human IFN-alpha hybrid B/D [hIFN-alpha B/D], respectively). The ED50 for the eight cytotoxic drugs tested ranged from 2.3 x 10(-9) to 4.3 x 10(-13) mol/L and all were within the therapeutic range for humans. Combination hIFN-alpha B/D and melphalan were found to be additive in their inhibitory effects on myeloma cell growth in vitro and this finding was confirmed in vivo in C57BL/KaLwRij mice bearing disseminated 5T33 myeloma. Control animals demonstrated a median survival duration of 25.3 days whereas hIFN-alpha B/D or melphalan treatment alone increased survival to 30.5 and 33.3 days, respectively (P < 0.001). Combination IFN-alpha/melphalan therapy increased median survival duration to 38.5 days (P < 0.001) which was also significantly greater than that obtained with single agent therapy (P < 0.01). The murine myeloma cells were found to be resistant to NK cell lysis but susceptible to lysis by LAK cells (49.3 +/- 6.3% lysis at an effector to target ratio of 100:1).(ABSTRACT TRUNCATED AT 250 WORDS)

Effects of interferon alpha on autocrine growth factor loops in B lymphoproliferative disorders.

The B lymphoproliferative disorders B chronic lymphocytic leukemia (B-CLL) and hairy cell leukemia (HCL) produce a number of autocrine growth factors, including tumor necrosis factor (TNF), interleukin 6 (IL-6), and IL-1, all of which may induce positive feedback growth loops. If such malignancies depend on these autocrine growth loops for survival, their interruption may be therapeutically valuable. Interferon alpha (IFN-alpha) abrogates TNF- or IL-6-induced proliferation of HCL and B-CLL cells in vitro and has therapeutic activity in these diseases. We have investigated the possibility that IFN-alpha may act by interrupting autocrine growth factor loops. If purified B-CLL or HCL cells are cultured in the presence of TNF, there is induction of mRNA for TNF, IL-1 alpha, IL-1 beta, and IL-6. However, culture in the presence of IFN-alpha in addition to TNF reduced the level of mRNA for all these cytokines, compared with cells cultured in TNF alone. While cytokine mRNA levels were diminished, levels of mRNA for the ribonuclease activator 2-5A synthetase were increased. Analysis of the kinetics of cytokine mRNA production showed that levels fall shortly after the rise of 2-5A synthetase mRNA. IFN-alpha may produce these effects by shortening the half-life of cytokine mRNA, since TNF mRNA half-life in B-CLL and HCL cells is substantially reduced when the cells are cultured with IFN-alpha. These data suggest that IFN-alpha may mediate its therapeutic effects in these malignancies by blocking autocrine growth factor loops.

Cytokine-induced enhancement of the susceptibility of hairy cell leukaemia lymphocytes to natural killer cell lysis.

In seven patients with HCL we have investigated how the cytokines TNF, alpha and gamma IFN and IL2 modify the interactions between HCL lymphocytes and MHC unrestricted cytotoxic effector cells [natural killer (NK) and lymphokine activated killer (LAK), cells]. We find that IL2 and alpha IFN increase patient NK activity against K562, though killing remains subnormal. IL2 and alpha IFN also induce normal levels of LAK activity, measured against an NK resistant B lymphoblastoid cell line. In contrast, gamma IFN has no effect on patient effector function. However, promotion of cytotoxic effector activity is unlikely to produce therapeutic benefit since HCL cells themselves are entirely resistant to NK/LAK killing. Susceptibility of HCL cells can be induced by culturing the cells in the presence of both gamma IFN and TNF, but not by culture with either cytokine alone. This synergy is not mediated by a gamma IFN induced increase in TNF receptors on HCL lymphocytes, and therefore occurs at a post-receptor level.

In vitro analysis of the interactions of recombinant IL-2 with regenerating lymphoid and myeloid cells after allogeneic marrow transplantation.

Although administration of rIL-2 post-T depleted allogeneic bone marrow transplantation (TD-BMT) offers the prospect of augmenting immune reconstitution and thereby reducing the risks of infection and relapse, it has been unclear what direct or indirect effects this agent would have on the regenerating myeloid system. We find that addition of 200 IU or rIL-2 to patient lymphocytes obtained within 6 wk of TD-BMT results in a substantial (2 to 3 log) increase in INF-gamma secretion and the production of TNF. Cytokines present in supernatants obtained from IL-2-stimulated patient lymphocytes have two contrasting effects on myeloid cells from normal donors and from marrow recipients. They prime granulocytes for enhanced oxidative metabolism as measured by ability to generate chemiluminescence in response to FMLP, whereas IL-2 added directly to neutrophils has no effect. However, these IL-2-induced cytokines also act to inhibit myeloid progenitor growth and reduce granulocyte macrophage (GM) colony formation by a mean of 53%. Preincubation of supernatants with anti-IFN-gamma antibody partially abrogates both enhancement of granulocyte chemiluminescence and suppression of marrow CFU-GM. Addition of IL-2 directly to recipient marrow also produces inhibition, leading to a 25% reduction of GM-colony growth. This effect is not due to direct interaction between myeloid progenitor cells and IL-2, because it is completely abrogated by removal of CD8 and Leu-7+ lymphocytes from the marrow. Although the suppressive effects on marrow growth in vitro are of particular concern after BMT, the potential of IL-2 to promote granulocyte function, immune reconstitution, and anti-leukemic activity after TD-BMT justify further consideration of IL-2 therapy in this setting.

Tumour necrosis factor as an autocrine tumour growth factor for chronic B-cell malignancies.

Recombinant tumour necrosis factor (TNF) promotes survival and induces proliferation in the tumour cells from two malignancies of B lymphocytes--hairy-cell leukaemia and B-chronic lymphocytic leukaemia. Culture with TNF also induces TNF mRNA and protein, so the cytokine may act as an autocrine tumour growth factor. These growth promoting effects are antagonised by alpha but not by gamma interferon.

Effects of tumour necrosis factor and alpha interferon on chronic B cell malignancies.

Tumour necrosis factor (TNF) induces the lysis of many malignant cells in vitro and regression of some tumours in vivo. However, TNF is also a growth factor for normal fibroblasts, T cells and B cells and we have recently shown that TNF can also act as a growth factor for chronic B cell neoplasms, including hairy cell leukaemia and B-CLL. In these cells it promotes proto-oncogene expression, RNA and DNA synthesis and increases overall cell survival. Stimulation appears to be autocrine in nature since exposure of the neoplastic cells to recombinant TNF protein induces the corresponding messenger RNA and synthesis of the protein itself. TNF induced proto-oncogene expression and DNA synthesis occur over a substantially longer time period than when the cells are stimulated with agents such as TPA and Calcium ionophore (2), but we have no evidence that the delay represents the time taken to generate TNF dependent secondary cytokines such as IL-1 and IL6. Alpha interferon opposes TNF mediated activation and our recent data indicate that this effect is independent of alpha interferon down regulation of TNF receptors. It appears to be related instead to a decreased accumulation of TNF mRNA which occurs contemporaneously with an alpha interferon induced rise in 2-5 A synthetase. If TNF dependent growth is important for the survival of B-CLL cells, then agents which mimic alpha interferon or which block TNF induced autocrine growth would be predicted to be of therapeutic benefit.

Giardia infection causes vitamin B12 deficiency.

A patient presented with hematological evidence of vitamin B12 deficiency. The Schilling test performed suggested intestinal malabsorption and further investigation revealed heavy infestation with Giardia lamblia. Specific treatment of the giardiasis with tinidazole resulted in correction of the abnormalities in vitamin B12 absorption. These findings, together with the absence of other causes of vitamin B12 deficiency, suggest that giardiasis should be considered as a cause of vitamin B12 deficiency.

Reversible renal failure in malignant hypertension.

Acute renal failure requiring dialysis to sustain life may be due to malignant hypertension. If the blood pressure is controlled during a period of dialysis, then it is possible that the renal lesions may heal, with some recovery of renal function. This report describes eight patients with acute renal failure due to malignant hypertension who required temporary dialysis. In all eight cases adequate control of blood pressure was achieved and all recovered renal function such that dialysis could be discontinued. The longest period of follow-up was five years, and one patient achieved a creatinine clearance of 23 ml/min. Renal histology, available in five cases, showed changes of malignant hypertension only, with no evidence of other renal lesions. A review of the literature is presented and the potentially reversible nature of acute oliguric renal failure due to malignant hypertension is emphasized.