The current study took advantage of widespread recruitment of patients with steroid-responsive HES for participation in a clinical trial, in order to estimate the proportion of patients with L-HES and/or increased serum TARC values in a large cohort, and to explore responses of patients with T-cell driven disease to highly targeted eosinophil-specific therapy. Diagnosis of L-HES was based on detection of abnormal T-cell phenotypes associated with evidence of clonal T-cell expansion. Demonstration of T-cell clonality alone was not considered sufficient evidence for L-HES, based on recent studies showing clonal TCR rearrangements in high proportions of HES patients, the majority of whom have normal T-cell phenotypes and no evidence of increased IL-5 production [13
]. The proportion of patients with L-HES included in this clinical trial (13/63) was approximately 20%, which is similar to that reported by Simon et al (16/60 [25%]) in a cohort of HES patients referred mainly to dermatologists [7
]. A recent, large retrospective multicenter study, designed to minimize the referral bias of single-center studies, reported 17% patients with L-HES, but the actual proportion with well-documented T-cell phenotype abnormalities was only 8.7% [15
]. The relatively high proportion of phenotypically abnormal T-cell subsets observed in our study can be explained by exclusion of patients with FIP1L1/PDGFRA-associated HES from the clinical trial, and by selection of patients who responded to CS-monotherapy, which likely enriched the cohort for L-HES patients compared with more aggressive, presumably myeloproliferative, disease forms.
Results of T-cell analyses on this patient cohort illustrate the heterogeneity within L-HES and how challenging formal diagnosis of this variant has become. In addition to patients with previously well-characterized phenotypically aberrant T-cell subsets, (oligo)clonal TCR rearrangement patterns were observed in some patients with phenotype abnormalities which have not yet been investigated in the setting of HES (e.g. double positive T-cells). Although demonstration of increased IL-5 production at the single-cell level should clarify the possible pathogenic role of these T-cell subsets, this is not within the bounds of routine explorations, and biomarkers of T-cell driven hypereosinophilia, which override phenotypic and functional differences, are desperately needed. Previous studies show that TARC, which is produced by various cell-types in response to IL-4 [17
], may be increased in various conditions associated with in vivo
Th2-cell activation, including L-HES [9
]. We therefore measured serum TARC to identify additional patients with presumed Th2-driven disease in this study, beyond those with clear-cut phenotype abnormalities. Forty percent (33/81) of HES patients in this study had a peak serum TARC level above 1000 pg/ml (), including all 13 patients with phenotypically proven L-HES although they were under CS, and even when the proportion of aberrant cells was very low (less than 2.5% lymphocytes in 3 cases).
Mepolizumab was an effective CS-sparing agent in patients with L-HES and/or increased serum TARC, with response rates similar to patients with a normal T-cell profile for the primary and more stringent PDN end-points (). Remarkably, approximately half of the patients were able to taper completely off CS by the end of the study, regardless of their T-cell profile. In contrast, a lower proportion of patients with L-HES and/or serum TARC above 1000 pg/ml durably maintained eosinophil levels below 600/μl during mepolizumab therapy ( and ). Viewed differently, the only 4 patients in the trial whose eosinophil levels increased above 600/μl despite mepolizumab had increased serum TARC and clear-cut evidence for Th2-mediated disease. Indeed, 2 of them had CD3-
L-HES (patients 5 and 6, ), 1 had a clonal TCR rearrangement pattern and extremely elevated serum TARC (patient 17, ), and the fourth patient had increased serum TARC and IgE levels, as well as the highest serum IL-4 level observed in this study (125 pg/ml, not shown) (patient T27, ). For patients 6 and 17, the rise in eosinophilia was easily overcome with low-dose PDN maintenance therapy (<10 mg daily); in these conditions, HES-related clinical manifestations were well-controlled, and eosinophils never exceeded 1000/μl (Table S2
). In contrast, patients 5 and T27 were truly resistant to mepolizumab, with uncontrolled eosinophilia and recurrent HES-related symptoms as PDN-tapering was attempted, resulting in permanent withdrawal from the study in one case, and HES-related death in the other. Mepolizumab’s failure to deplete eosinophils in these cases likely reflects incomplete neutralization of IL-5, due to marked IL-5 over-production in vivo and/or antibody-inaccessible local production of IL-5; and possibly uninhibited effects of other eosinophilopoïetic factors produced by dysregulated T-cells. Although T-cell production of IL-5 was not assessed in the current study, several publications demonstrate that CD3-
, and CD3+
T-cells from patients with L-HES [4
], and PBMC from patients with increased serum TARC [9
], produce more IL-5 in vitro than CD4 T-cells and PBMC from control subjects and other HES patients. Further evidence that increased TARC levels are indicative of a stronger eosinophilopoïetic drive is provided in the placebo-treatment arm, wherein a higher proportion of patients with serum TARC above 2000 pg/ml experienced recurrence of hypereosinophilia within 8 weeks during the initial CS-tapering period (90% vs 46%, p=0.025, data not shown). The ongoing open-label extension study should clarify the long-term practical implications of these findings, namely the impact on intervals between mepolizumab infusions, as well as the requirement for CS maintenance therapy.
Overall, these results indicate that mepolizumab 1) represents an effective CS-sparing agent for the majority of patients with T-cell driven HES (i.e. L-HES and/or increased serum TARC), similar to other HES patients, but 2) may provide only incomplete disease control as monotherapy in a subset of these patients, whose eosinophil levels remain above 600/μl. Given its excellent tolerance and safety profile [11
], mepolizumab represents a very tempting alternative to IFNα, the currently preferred but poorly tolerated CS-sparing agent for this HES variant. This study indicates that once mepolizumab is initiated, blood eosinophil levels and clinical status should, nevertheless, be monitored closely as CS doses are reduced for early detection of resistance or requirement of a PDN maintenance dose. Maintenance of low-dose CS may actually be preferable for all patients with L-HES under mepolizumab, because, in contrast to CS and IFNα [10
], mepolizumab is not expected to exert inhibitory effects on the pathogenic T-cells involved in L-HES. Indeed, the abnormal T-cells remain detectable in peripheral blood throughout the study (Table S1
) and there is reason to believe that both mepolizumab itself [22
] and the CS-tapering it enables may increase their production of Th2 cytokines.
Finally, although the 2 patients with T-cell lymphoma were excluded from our analyses, we underscore that neither reached the primary end-point under mepolizumab. The patient with mycosis fungoïdes presented persistent marked erythroderma requiring continued CS despite effective depletion of blood and skin-infiltrating eosinophils with mepolizumab, whereas the patient with AITL responded neither in terms of disease manifestations nor eosinophil levels. Both received appropriate therapy for their lymphoma; the first is in remission, and the second died a week later of cardiorespiratory failure, due to heart infiltration by the malignant T-cells. The fact that 2 patients with high serum TARC levels initially diagnosed with HES were ultimately diagnosed with lymphoma suggests that increased serum TARC should prompt careful assessment for underlying lymphoma, both at diagnosis and at regular intervals. Furthermore, it is conceivable that targeted anti-eosinophil strategies may unmask and even accelerate progression of such diseases by enabling CS tapering in some patients, and thereby alleviating some of the pressure on abnormal T-cells. This possibility is of utmost relevance for patients with L-HES, namely CD3-CD4+ associated disease, as these T-cells may be pre-malignant, and are sensitive to the pro-apoptotic effects of CS. Prolonged follow-up of patients with L-HES on mepolizumab therapy in the open-label extension study should provide some insight into the effects of treatment on numbers and activation status of pathogenic T-cell subsets.
In conclusion, this study shows that HES patients with phenotypically abnormal T cell subsets and/or increased serum TARC levels respond to mepolizumab and are able to taper CS at a rate that is comparable to that of the overall HES population. However, in contrast to patients with normal T-cell profiles, mepolizumab fails to deplete eosinophils in a subset of patients with T-cell driven disease, suggesting that overproduced IL-5 and/or other eosinophilopoïetic factors are not (fully) neutralized. Furthermore, CS-tapering on mepolizumab may unmask CS-suppressed T cell disease. Pursuing low-dose CS may therefore be prudent in L-HES patients, in order to target the potentially pre-malignant T-cells, in parallel with eosinophil-directed therapy.