Treatment of primary plasma cell leukaemia with carfilzomib and lenalidomide-based therapy (EMN12/HOVON-129): final analysis of a non-randomised, multicentre, phase 2 study.

BACKGROUND: Primary plasma cell leukaemia is a rare and aggressive plasma cell disorder with a poor prognosis. The aim of the EMN12/HOVON-129 study was to improve the outcomes of patients with primary plasma cell leukaemia by incorporating carfilzomib and lenalidomide in induction, consolidation, and maintenance therapy. METHODS: The EMN12/HOVON-129 study is a non-randomised, phase 2, multicentre study conducted at 19 academic centres and hospitals in seven European countries (Belgium, Czech Republic, Denmark, Italy, Norway, The Netherlands, and the UK) for previously untreated patients with primary plasma cell leukaemia aged 18 years or older. Inclusion criteria were newly diagnosed primary plasma cell leukaemia (defined as >2 ×109 cells per L circulating monoclonal plasma cells or plasmacytosis >20% of the differential white cell count) and WHO performance status 0-3. Patients aged 18-65 years (younger patients) and 66 years or older (older patients) were treated in age-specific cohorts and were analysed separately. Younger patients were treated with four 28-day cycles of carfilzomib (36 mg/m2 intravenously on days 1, 2, 8, 9, 15, and 16), lenalidomide (25 mg orally on days 1-21), and dexamethasone (20 mg orally on days 1, 2, 8, 9, 15, 16, 22, and 23). Carfilzomib-lenalidomide-dexamethasone (KRd) induction was followed by double autologous haematopoietic stem-cell transplantation (HSCT), four cycles of KRd consolidation, and then maintenance with carfilzomib (27 mg/m2 intravenously on days 1, 2, 15, and 16 for the first 12 28-day cycles, and then 56 mg/m2 on days 1 and 15 in all subsequent cycles) and lenalidomide (10 mg orally on days 1-21) until progression. Patients who were eligible for allogeneic HSCT, could also receive a single autologous HSCT followed by reduced-intensity conditioning allogeneic HSCT and then carfilzomib-lenalidomide maintenance. Older patients received eight cycles of KRd induction followed by maintenance therapy with carfilzomib and lenalidomide until progression. The primary endpoint was progression-free survival. The primary analysis population was the intention-to-treat population, irrespective of the actual treatment received. Data from all participants who received any study drug were included in the safety analyses. The trial was registered at www.trialregister.nl (until June 2022) and https://trialsearch.who.int/ as NTR5350; recruitment is complete and this is the final analysis. FINDINGS: Between Oct 23, 2015, and Aug 5, 2021, 61 patients were enrolled and received KRd induction treatment (36 patients aged 18-65 years [20 (56%) were male and 16 (44%) female], and 25 aged ≥66 years [12 (48%) were male and 13 (52%) female]). With a median follow-up of 43·5 months (IQR 27·7-67·8), the median progression-free survival was 15·5 months (95% CI 9·4-38·4) for younger patients. For older patients, median follow-up was 32·0 months (IQR 24·7-34·6), and median progression-free survival was 13·8 months (95% CI 9·2-35·5). Adverse events were most frequently observed directly after treatment initiation, with infections (two of 36 (6%) younger patients and eight of 25 (32%) older patients) and respiratory events (two of 36 [6%] younger patients and four of 25 [16%] older patients) being the most common grade 3 or greater events during the first four KRd cycles. Treatment-related serious adverse events were reported in 26 (72%) of 36 younger patients and in 19 (76%) of 25 older patients, with infections being the most common. Treatment-related deaths were reported in none of the younger patients and three (12%) of the older patients (two infections and one unknown cause of death). INTERPRETATION: Carfilzomib and lenalidomide-based therapy provides improved progression-free survival compared with previously published data. However, results remain inferior in primary plasma cell leukaemia compared with multiple myeloma, highlighting the need for new studies incorporating novel immunotherapies. FUNDING: Dutch Cancer Society, Celgene (a BMS company), and AMGEN.

CD4+ CCR6+ T cells, but not γδ T cells, are important for the IL-23R-dependent progression of antigen-induced inflammatory arthritis in mice.

IL-23 plays an important role in the development of arthritis and the IL-23 receptor (IL-23R) is expressed on different types of T cells. However, it is not fully clear which IL-23R+ T cells are critical in driving T cell-mediated synovitis. We demonstrate, using knock-in IL-23R-GFP reporter (IL-23RGFP/+ ) mice, that CD4+ CCR6+ T cells and γδ T cells, but not CD8+ T cells, express the IL-23R(GFP). During early arthritis, IL-23R(GFP)+ CD4+ CCR6+ T cells, but not IL-23R(GFP)+ γδ T cells, were present in the inflamed joints. IL-23RGFP/+ mice were bred as homozygotes to obtain IL-23RGFP/GFP (IL-23R deficient/IL-23R-/- ) mice, which express GFP under the IL-23R promotor. Arthritis progression and joint damage were significantly milder in IL-23R-/- mice, which revealed less IL-17A+ cells in their lymphoid tissues. Surprisingly, IL-23R-/- mice had increased numbers of IL-23R(GFP)+ CD4+ CCR6+ and CCR7+ CD4+ CCR6+ T cells in their spleen compared to WT, and IL-23 suppressed CCR7 expression in vitro. However, IL-23R(GFP)+ CD4+ CCR6+ T cells were present in the synovium of IL-23R-/- mice at day 4. Finally, adoptive transfer experiments revealed that CD4+ CCR6+ T cells and not γδ T cells drive arthritis progression. These data suggest that IL-23R-dependent T cell-mediated synovitis is dependent on CD4+ CCR6+ T cells and not on γδ T cells.

The role of IL-23 receptor signaling in inflammation-mediated erosive autoimmune arthritis and bone remodeling.

The IL-23/Th17 axis has been implicated in the development of autoimmune diseases, such as rheumatoid arthritis (RA) and psoriatic arthritis (PsA). RA and PsA are heterogeneous diseases with substantial burden on patients. Increasing evidence suggests that the IL-23 signaling pathway may be involved in the development of autoimmunity and erosive joint damage. IL-23 can act either directly or indirectly on bone forming osteoblasts as well as on bone resorbing osteoclasts. As IL-23 regulates the activity of cells of the bone, it is conceivable that in addition to inflammation-mediated joint erosion, IL-23 may play a role in physiological bone remodeling. In this review, we focus on the role of IL-23 in autoimmune arthritis in patients and murine models, and provide an overview of IL-23 producing and responding cells in autoimmune arthritic joints. In addition, we discuss the role of IL-23 on bone forming osteoblasts and bone resorbing osteoclasts regarding inflammation-mediated joint damage and bone remodeling. At last, we briefly discuss the clinical implications of targeting this pathway for joint damage and systemic bone loss in autoimmune arthritis.

Inhibition of MicroRNA-494 Reduces Carotid Artery Atherosclerotic Lesion Development and Increases Plaque Stability.

OBJECTIVES: Unstable atherosclerotic lesions in carotid arteries require surgical endarterectomy to reduce the risk of ischemic stroke. We aimed to identify microRNAs that exert a broad effect on atherosclerotic plaque formation and stability in the carotid artery. BACKGROUND: We made a selection of 164 genes involved in atherosclerosis. Using www.targetscan.org, we determined which microRNAs potentially regulate expression of these genes. We identified multiple microRNAs from the 14q32 microRNA cluster, which is highly involved in vascular remodeling. In human plaques, collected during carotid endarterectomy surgery, we found that 14q32 microRNA (miR-494) was abundantly expressed in unstable lesions. METHODS: We induced atherosclerotic plaque formation in hypercholesterolemic ApoE mice by placing semiconstrictive collars around both carotid arteries. We injected "Gene Silencing Oligonucleotides" against miR-494 (GSO-494) or negative control (GSO-control). Using fluorescently labeled GSOs, we confirmed uptake of GSOs in affected areas of the carotids, but not elsewhere in the vasculature. RESULTS: After injection of GSO-494, we observed significant downregulation of miR-494 expression in the carotid arteries, although miR-494 target genes were upregulated. Further analyses revealed a 65% decrease in plaque size after GSO-494 treatment. Plaque stability was increased in GSO-494-treated mice, determined by an 80% decrease in necrotic core size and a 50% increase in plaque collagen content. Inhibition of miR-494 also resulted in decreased cholesterol levels and decreased very low-density lipoprotein (VLDL) fractions. CONCLUSIONS: Treatment with GSO-494 results in smaller atherosclerotic lesions with increased plaque stability. Inhibition of miR-494 may decrease the risk of surgical complications or even avert endarterectomy surgery in some cases.

Characterizing memory T helper cells in patients with psoriasis, subclinical, or early psoriatic arthritis using a machine learning algorithm.

BACKGROUND: Psoriasis patients developing psoriatic arthritis (PsA) are thought to go through different phases. Understanding the underlying events in these phases is crucial to diagnose PsA early. Here, we have characterized the circulating memory T helper (Th) cells in psoriasis patients with or without arthralgia, psoriasis patients who developed PsA during follow-up (subclinical PsA), early PsA patients and healthy controls to elucidate their role in PsA development. METHODS: We used peripheral blood mononuclear cells of sex and age-matched psoriasis patients included in Rotterdam Joint Skin study (n=22), early PsA patients included in Dutch South West Early Psoriatic Arthritis Cohort (DEPAR) (n=23) and healthy controls (HC; n=17). We profiled memory Th cell subsets with flow cytometry and used the machine learning algorithm FlowSOM to interpret the data. RESULTS: Three of the 22 psoriasis patients developed PsA during 2-year follow-up. FlowSOM identified 12 clusters of memory Th cells, including Th1, Th2, Th17/22, and Th17.1 cells. All psoriasis and PsA patients had higher numbers of Th17/22 than healthy controls. Psoriasis patients without arthralgia had lower numbers of CCR6-CCR4+CXCR3+ memory Th cells and higher numbers of CCR6+CCR4-CXCR3-memory Th cells compared to HC. PsA patients had higher numbers of Th2 cells and CCR6+CCR4+CXCR3- cells, but lower numbers of CCR6+CCR4+CXCR3+ memory Th cells compared to HC. The number of CCR6+ Th17.1 cells negatively correlated with tender joint counts and the number of CCR6+ Th17 cells positively correlated with skin disease severity. CONCLUSIONS: Unsupervised clustering analysis revealed differences in circulating memory Th cells between psoriasis and PsA patients compared to HC; however, no specific subset was identified characterizing subclinical PsA patients.

IL-23 receptor deficiency results in lower bone mass via indirect regulation of bone formation.

The IL-23 receptor (IL-23R) signaling pathway has pleiotropic effects on the differentiation of osteoclasts and osteoblasts, since it can inhibit or stimulate these processes via different pathways. However, the potential role of this pathway in the regulation of bone homeostasis remains elusive. Therefore, we studied the role of IL-23R signaling in physiological bone remodeling using IL-23R deficient mice. Using µCT, we demonstrate that 7-week-old IL-23R-/- mice have similar bone mass as age matched littermate control mice. In contrast, 12-week-old IL-23R-/- mice have significantly lower trabecular and cortical bone mass, shorter femurs and more fragile bones. At the age of 26 weeks, there were no differences in trabecular bone mass and femur length, but most of cortical bone mass parameters remain significantly lower in IL-23R-/- mice. In vitro osteoclast differentiation and resorption capacity of 7- and 12-week-old IL-23R-/- mice are similar to WT. However, serum levels of the bone formation marker, PINP, are significantly lower in 12-week-old IL-23R-/- mice, but similar to WT at 7 and 26 weeks. Interestingly, Il23r gene expression was not detected in in vitro cultured osteoblasts, suggesting an indirect effect of IL-23R. In conclusion, IL-23R deficiency results in temporal and long-term changes in bone growth via regulation of bone formation.

Dendritic Cell-Specific Deletion of ß-Catenin Results in Fewer Regulatory T-Cells without Exacerbating Autoimmune Collagen-Induced Arthritis.

Dendritic cells (DCs) are professional antigen presenting cells that have the dual ability to stimulate immunity and maintain tolerance. However, the signalling pathways mediating tolerogenic DC function in vivo remain largely unknown. The ß-catenin pathway has been suggested to promote a regulatory DC phenotype. The aim of this study was to unravel the role of ß-catenin signalling to control DC function in the autoimmune collagen-induced arthritis model (CIA). Deletion of ß-catenin specifically in DCs was achieved by crossing conditional knockout mice with a CD11c-Cre transgenic mouse line. Bone marrow-derived DCs (BMDCs) were generated and used to study the maturation profile of these cells in response to a TLR2 or TLR4 ligand stimulation. CIA was induced by intra-dermal immunization with 100 µg chicken type II collagen in complete Freund's adjuvant on days 0 and 21. CIA incidence and severity was monitored macroscopically and by histology. The T cell profile as well as their cytokine production were analysed by flow cytometry. Lack of ß-catenin specifically in DCs did not affect the spontaneous, TLR2- or TLR4-induced maturation and activation of BMDCs or their cytokine production. Moreover, no effect on the incidence and severity of CIA was observed in mice lacking ß-catenin in CD11c+ cells. A decreased frequency of splenic CD3+CD8+ T cells and of regulatory T cells (Tregs) (CD4+CD25highFoxP3+), but no changes in the frequency of splenic Th17 (CCR6+CXCR3-CCR4+), Th2 (CCR6-CXCR3-CCR4+) and Th1 (CCR6-CXCR3+CCR4-) cells were observed in these mice under CIA condition. Furthermore, the expression of IL-17A, IL-17F, IL-22, IL-4 or IFNγ was also not affected. Our data indicate that ablation of ß-catenin expression in DCs did not alter the course and severity of CIA. We conclude that although deletion of ß-catenin resulted in a lower frequency of Tregs, this decrease was not sufficient to aggravate the onset and severity of CIA.