Effects Of Ruxolitinib Therapy On Megakaryocyte Morphology and Inflammatory Bone Marrow Reaction In Patients With Myelofibrosis

Bone marrow (BM) microenvironment plays an important role in the initiation and progression of myelofibrosis (MF). The dysregulation of proinflammatory cytokine production by both hematopoietic stem cells via JAK-activating mutations and surrounding stromal cells generates a microenvironment that is functionally linked to disease-associated increase in BM fibrosis, constitutional symptoms, splenomegaly, and extramedullary hematopoiesis. It has been shown that proliferation of atypical megakaryocytes (MK) and their pathologic interaction with the marrow stroma plays a central role in MF. Recent data indicate that ruxolitinib (RUX) treatment results in a reduction in the level of cytokines and other inflammatory markers. However, it is still not clear whether these effects are reflected by corresponding BM changes, in particular with regard to macrophages (MAK), plasma cells (PL) and morphology of MK.

A total of 46 patients (pts) with MF presenting at baseline with various degrees of BM fibrosis (grade 1: n=8; grade 2: n=24; grade 3: n=14) were selected from a MD Anderson Cancer Center phase 1/2 study (NCT00509899). All cases had a sequential BM biopsy taken at 24 months (mo) following RUX therapy. Analysis included immunohistochemical and morphometric assessment of MK (overall frequency, degree of clustering and dysplastic changes), amount of erythropoiesis (ERY), frequency of macrophages (MAK) and plasma cells (PL). Specific antibodies were applied to identify these subpopulations (CD61 – MK; Glycophorin C [Ret40f] – ERY; CD68 [PGM-1] – MAK; MUM1 – PL). Individual changes were calculated for each parameter and correlated with clinical features.

Following therapy the majority of pts revealed a stabilization or improvement of BM fibrosis (improvement: 13%, stabilization: 57%, worsening: 30%). In line with previously published data an increased number of MAK was found at baseline independent of BM fibrosis grade. In 56% of pts therapy induced a reduction of MAK, 37% pts revealed no effect, and 3 cases showed an increase (7%). As indicator of an underlying inflammatory stroma reaction, about half of the pts displayed increased numbers of PL at baseline. In the majority of pts amount of PL remained stable (64%), 24% had a reduction, and 12% showed a further increase. Initially 80% of cases presented with a marked increase in MK, however, frequency was reduced over therapy in about 50%. Dense clustering of MK was slightly decreased (baseline 63% vs 24 mo 39%), but no significant treatment effect was observed concerning evolution of myelodysplastic features. In most cases a significant reduction of hematopoietic cellularity was encountered (median change -36%). Therapy had no significant effect on ERY (median change -5%).

Pts with increased numbers of MAK at 24 mo had a greater spleen size reduction compared to those with reduced MAK during therapy (median change -13.5 vs -6.5 cm; Mann-Whitney U test: p=0.006). Therapy induced increase in MAK was not associated with hemoglobin levels at 24 mo (median change -0.2 vs -0.9 g/dl; p=0.217), furthermore no correlation with platelets or symptoms was found.

In contrast, reduction of PL at 24 mo was associated with pronounced spleen size reduction (median change -16.0 vs -5.5 cm; p=0.062) and higher levels of hemoglobin (median change +0.6 vs -0.8 g/dl; p=0.077). No effects were observed on platelet counts and reduction of MF related symptoms (p=0.265).

Improvement in BM fibrosis at 24 mo was associated in 67% with a corresponding lowering of overall MK frequency, contrasting cases with progression that revealed in 80% an increase in dysplastic features. Overall, changes in MK frequency and morphology were not linked with spleen size reduction, hemoglobin or symptoms. However, decrease in MK quantity resulted in lower platelet counts at 24 mo (median change -299 vs -162 x10⁹/l; p=0.165).

Our results highlight recent data on JAK inhibitor therapy in pts with MF with regard to improvement in BM fibrosis and an overall reduction of an inflammatory condition. RUX therapy induces a modulation of the BM microenvironment that is linked with spleen size reduction and normalization of MK morphology. Ongoing research is further exploring the significance of our current findings, including a correlation with other proinflammatory cytokines and extracellular matrix proteins like LOX.

Kvasnicka:Novartis: Consultancy; Incyte Corporation: Consultancy; Novartis: Research Funding; Incyte Corporation: Research Funding; Shire: Research Funding; AOP Orphan Pharmaceuticals: Research Funding; Novartis: Honoraria; Shire: Honoraria; Incyte Corporation: Honoraria. Thiele:AOP Orphan Pharmaceuticals: Consultancy; Incyte Corporation: Consultancy; Novartis: Consultancy; Shire: Consultancy; Sanofi: Consultancy; Novartis: Research Funding; Shire: Research Funding; AOP Orphan Pharmaceuticals: Honoraria; Incyte Corporation: Honoraria; Novartis: Honoraria; Shire: Honoraria; Sanofi: Honoraria. Cortes:Ambit: Research Funding. Kantarjian:Ariad: Research Funding; Novartis: Research Funding; BMS: Research Funding; Pfizer: Research Funding. Verstovsek:Incyte Corporation: Research Funding.