Heather Edwards MB BCh
Jonathan Kell MB BChir MD FRCPath
University Hospital of Wales,
Acute myeloid leukaemia (AML) is the most common form of acute leukaemia in adults, with a median age at presentation of 65 years, an overall incidence of 3.7 per 100,000 and an age-related mortality of 2.7–18 per 100,000. Whereas most patients will enter a complete remission with conventional chemotherapy, most of these patients will relapse in the first two years following treatment. In acute lymphoblastic leukaemia, this high risk of relapse post treatment is mitigated by maintenance therapy for up to three years following intensive induction and consolidation therapy. There are few studies examining the value of maintenance chemotherapy in AML. The UK NCRN AML16 study in older patients examined the use of the hypomethylating agent azacitidine in maintenance in AML and the results are awaited.
Interleukin-2 (IL-2) is a pro-inflammatory cytokine targeting activated T cells and natural killer (NK) cells and has been used as a biological adjunct in the treatment of various haematological malignancies. These studies have shown modest benefit but marked toxicity at conventional doses. More recently, combination of IL-2 with histamine dihydrochloride (HDC) has been shown to be a safe and potentially effective combination that may prevent relapse in patients with AML when given as maintenance therapy.
Biology of IL-2 and HDC
IL-2 is produced by T cells and stimulates CD25-activated T cells and NK cells to proliferate. NK cells stimulated by IL-2 recognise and kill malignant cells and virally infected cells in vivo and are capable of killing AML blasts in vitro.(1–3) However, this lysis of AML blasts is usually seen in pure cultures of NK cells with leukaemic blasts. This has not translated into clinical benefit, raising the possibility of an inhibitor factor.
Monocytes produce reactive oxygen radicals, such as superoxide (O2–), hydroxyl ions (OH–) and halous ions such as hypochlorous acid (HOCl–), by the respiratory burst using NADPH oxidase. It has been shown that using scavengers of reactive oxygen metabolites in the presence of monocytes hinders the suppression of NK cells. This was shown in a study where both catalase, which reduces hydrogen peroxide to water, and histamine were individually added to NK cells alone and NK cells in the presence of monocytes.(3)
Catalase had no effect on NK cell function in the absence of monocytes but abrogated the monocyte-induced inhibition of NK cells in culture.(3) Histamine also reversed monocytic inhibition of NK cells at 10-7M when added within the first hour of co-incubation of the monocytes and NK cells. However, histamine was ineffective in the presence of catalase. This was true both for monocytes that were resting and for those that had been induced to generate reactive oxygen metabolites. Furthermore, the addition of H2O2 in small concentrations will reverse this inhibition of monocytes inhibiting NK cells. It is concluded that monocyte function on NK cells is dependent on the generation of oxygen radicals.
Hellstrand et al have further shown that culture with monocytes will also abrogate IL-2-mediated NK cell lysis. Both histamine and catalase restored the function of IL-2 in these experiments. Histamine inhibits the respiratory burst in monocytes by acting through the histamine-2 receptor (H2R). Other H2R agonists also inhibit the respiratory burst and the effect can be overcome with H2 antagonists such as ranitidine. Interestingly, monocytes derived from patients with chronic granulomatous disease, which have a defective respiratory burst, fail to inhibit NK function in these experiments.(3)
IL-2 and histamine synergistically enhance NK cell-mediated cytotoxicity in murine models of human cancer. Treatment of mice with histamine of other H2 agonists has resulted in increased clearance of YAC-1 lymphoma cells and inhibited the metastasis of B16/F1 melanoma cells in mice.(4,5) Furthermore, in a murine ALL model, tissue depletion of histamine dihydrochloride resulted in a decreased surviv
al, whereas repletion of HDC improved survival.(6)
Brune(7) confirmed the effects of IL-2 and histamine on fresh leukaemic blasts and treated seven patients in remission from AML with the combination of IL-2 and histamine. Although very small numbers of patients are involved (too few to make robust deductions), this is an important proof of principle in that the combination was relatively non-toxic, with only short-lasting flushes, modest arterial hypotension and headache being reported. The crucial finding is that the dose of IL-2 used in this study (6.3–9 x105 iu bd) is a fraction of the historical dose used in the initial studies of IL-2 (12–24Mu/m2/day), which caused significant toxicity.(8)
By inhibiting monocytes, IL-2 can stimulate T cells and NK cells to lyse peripheral blasts, thus producing similar effects in vivo as in vitro. Hellstrand et al suggest that histamine inhibits the formation of H2O2 by monocytes by acting on the H2R. Hellstrand et al(4) gave ranitidine to patients donating the blood used in the trial, which blocked the H2R and therefore prevented histamine’s suppressive effect on monocytes.
Current chemotherapy regimes are successful in inducing remission in many patients with AML, but most patients will experience a relapse with generally poor outcome after re-induction. Various means of preventing relapse have been tried with intensification of consolidation therapy(9), maintenance with azacitidine (as in the AML16 study) and allogeneic bone marrow transplantation. Of these, allogeneic transplantation carries the best prospect of cure, relying on the donor immune system to eradicate minimal residual disease in the recipient.
However, there are significant toxicities associated with bone marrow transplantation and it is available to only younger patients with a suitably matched donor. Nonetheless, the utilisation of immune cells to control malignant disease is an attractive thesis. NK cells are thought to be fundamental in the prevention of relapse of AML following bone marrow transplants and patients who do not relapse after remission have been found to have good functioning NK cells. T-cell depletion of donor grafts results in a higher relapse rate. Several studies have examined using IL-2 to stimulate innate T-cell responses in patients with AML in remission.(8)
The amalgamated results of these studies were reviewed in a recent meta-analysis.(10) They examined five randomised controlled trials, using individual patient data, which recruited a total of 905 patients, of whom 449 were treated with IL-2 with 456 controls. A sixth study with grouped results was also included. A range of doses and dosing regimens was used. None of the individual trials produced a statistically significant reduction in relapse rates in the treatment groups when compared with the control groups and the meta-analysis showed hazard ratios for leukaemia-free survival of 0.97 (p=0.74) and 1.08 for overall survival. It was concluded that there was no increase in survival without relapse or overall survival time when monotherapy of IL-2 was used.
Combination of IL-2 and histamine
A phase III trial by Brune et al(11) compared the use of IL-2 and HDC with no treatment in 320 patients with AML in first or subsequent complete remission; 160 were randomly assigned to either arm. Patients in the treatment group received twice-daily subcutaneous injections of both HDC and IL-2, which they administered at home for 10 cycles of three weeks over an 18-month period.
Histamine was given at 0.5mg bd and IL-2 at 16.4kU/kg bd. The IL-2 dose is 1.6Mu for a 100kg individual and is, therefore, a tenth of the dose used in single agent studies. The leukaemic-free and overall survivals three years after the last enrolment were measured as an endpoint.
Toxicities were limited and the side effect profile was largely related to injection site reactions, low-grade fevers, myalgia, grade 1 and 3 headache and systemic effects of histamine, such as facial flushing, arterial hypotension and palpitations. There were no grade 3 or 4 hypotensive reactions.
Leukaemia-free survival (LFS) was improved at one year following remission (48% vs 42%), at two years (41% vs 29%) and at three years (34% vs 27%). In patients in first clinical remission (CR), the LFS at three years is 40% vs 26% (p=0.1). There was no effect on overall survival at any of these time points. Also, the advantage of maintenance with IL-2/histamine was limited to the patients in first CR, with no effect seen in patients in second or subsequent remissions.
There is good scientific rationale for the use of the combination of IL-2 and histamine as maintenance therapy in first CR in AML. Histamine is able to overcome the monocyte-mediated inhibition of IL-2 stimulated NK cells in vitro. Interleukin-2 alone does not appear to affect long-term outcome in AML as a single agent, but Brune and colleagues(11) have published a randomised study showing a significant difference in three-year leukaemia-free survival for patients in first CR receiving the IL-2/histamine combination with tolerable toxicity.
Given this 50% increase in LFS, the question arises as to why this therapy is not standard practice, as the results were published in 2006. The study has been criticised primarily for the apparent poor outcome in the control arm. At 26%, this is significantly worse than that generally reported in trials of around 40–45%. The study reports a median follow-up of 46 months. Consequently, the study was recruiting patients in the years 2000–2002 and results were published in 2006. It is well recognised that the supportive care offered to patients with AML has been improving and this may go some way to explaining the discrepancy in the published results compared with other historical studies. In addition, it must be remembered that the Brune study was a prospective randomised trial and, therefore, both trial and control subjects were recruited at the same time and following similar induction therapies and so we must presume that the difference reported represents a genuine result, at least based on these data.
There remains reluctance among many opinion leaders, certainly in the UK, to bring IL-2/HDC maintenance therapy to the front-line treatment of AML, based on the results of this single randomised study. It would certainly be of great interest to have a second study to confirm these results and there is now a study recruiting in Europe to examine the effect of the combination on minimal residual disease and two-year leukaemia-free survival in first CR from AML (ClinicalTrials.gov identifier NCT01347996). Another potentially exciting area for development is combining HDC with azacitidine in high-risk myelodysplasia, and the Groupe Francophone des Myelodysplasies (GFM) is exploring this question (ClinicalTrials.gov NCT01324960).
Maintenance therapy for AML remains in its infancy, with failed attempts at using IL-2 as a single agent to stimulate auto-immune leukaemia cell killing. As our knowledge of the biology of NK cells has increased, we can now modulate that immune response further by using histamine to abrogate monocyte-induced inhibition of NK cells. The combination of IL-2 with HDC is potent in preclinical models and has now entered clinical trials. The first of these, a randomised controlled study in AML in remission, has shown a tantalising glimpse of a prolonged leukaemia-free survival that now needs to be confirmed in a second study which is currently recruiting in Europe. A further exciting area is in high-risk myelodysplasia and we await the results of these trials with great interest.
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- Schaich M et al. Cytarabine dose of 36 g/m² compared with 12 g/m² within first consolidation in acute myeloid leukemia: results of patients enrolled onto the prospective randomized AML96 study. J Clin Oncol 2011;29:2696–702.
- Buyse M et al. Individual patient data meta-analysis of randomized trials evaluating IL-2 monotherapy as remission maintenance therapy in acute myeloid leukemia. Blood 2011;11:7007–13.
- Brune M et al. Improved leukemia-free survival after postconsolidation immunotherapy with histamine dihydrochloride and interleukin-2 in acute myeloid leukemia: results of a randomized phase 3 trial. Blood 2006;108:88–96.