ALBERT

Description: Idiopathic multicentric Castleman disease (iMCD) is a rare and poorly understood hematologic disorder characterized by lymphadenopathy, systemic inflammation, cytopenias, and life-threatening multiorgan dysfunction. Interleukin-6 (IL-6) inhibition effectively treats approximately one-third of patients. Limited options exist for nonresponders, because the etiology, dysregulated cell types, and signaling pathways are unknown. We previously reported 3 anti-IL-6 nonresponders with increased mTOR activation who responded to mTOR inhibition with sirolimus. We investigated mTOR signaling in tissue and serum proteomes from iMCD patients and controls. mTOR activation was increased in the interfollicular space of iMCD lymph nodes (N = 26) compared with control lymph nodes by immunohistochemistry (IHC) for pS6, p4EBP1, and p70S6K, known effectors and readouts of mTORC1 activation. IHC for pS6 also revealed increased mTOR activation in iMCD compared with Hodgkin lymphoma, systemic lupus erythematosus, and reactive lymph nodes, suggesting that the mTOR activation in iMCD is not just a product of lymphoproliferation/inflammatory lymphadenopathy. Further, the degree of mTOR activation in iMCD was comparable to autoimmune lymphoproliferative syndrome, a disease driven by mTOR hyperactivation that responds to sirolimus treatment. Gene set enrichment analysis of serum proteomic data from iMCD patients (n = 88) and controls (n = 42) showed significantly enriched mTORC1 signaling. Finally, functional studies revealed increased baseline mTOR pathway activation in peripheral monocytes and T cells from iMCD remission samples compared with healthy controls. IL-6 stimulation augmented mTOR activation in iMCD patients, which was abrogated with JAK1/2 inhibition. These findings support mTOR activation as a novel therapeutic target for iMCD, which is being investigated through a trial of sirolimus (NCT03933904).

Description: Idiopathic multicentric Castleman disease (iMCD) is a rare hematologic illness involving episodic disease flares with polyclonal cytokine-induced lymphoproliferation, systemic inflammation, and life-threatening multi-organ dysfunction. iMCD is further classified by clinical features and the most severe cases of iMCD often fall into the thrombocytopenia, anasarca, fever/elevated C-reactive protein, reticulin myelofibrosis, renal dysfunction, and organomegaly (TAFRO) clinical subtype. A cytokine storm involving interleukin(IL)-6 drives disease pathogenesis in a subset of patients, however only 34% of patients were found to respond to anti-IL-6 therapy with siltuximab in its registrational clinical trial. The identification of next generation therapeutics for iMCD-TAFRO patients has been challenging as the etiology, pathological cell types, and signaling pathways involved in iMCD-TAFRO are largely unknown. In this study, we aimed to identify cellular drivers and pathophysiological mechanisms of iMCD-TAFRO through the use of an unbiased multi-omics approach. We obtained paired bulk peripheral blood mononuclear cells (PBMCs) from a cohort of ten iMCD-TAFRO patients isolated during disease flare and clinical remission. These paired PBMC samples were utilized for flow cytometry to assess immune cell frequency and phenotype between iMCD-TAFRO flare and remission as well as between flare and age/sex matched healthy donors (n = 10). Three paired iMCD-TAFRO samples were also selected for transcriptional profiling using single-cell RNA sequencing (scRNAseq). We observed phenotypic differences across the T cell, monocyte, and NK cell compartments. We observed a significant increase in the frequency of CD8 T cells within the T cell compartment as well as an increased frequency of granzyme B and perforin expressing CD8 T cells in iMCD-TAFRO flare compared to healthy donors. We also observed a significant increase in the frequency of CD56+ NK cells within the NK cell compartment and a significant increase in the frequency of CD14+ monocytes within the monocyte compartment during iMCD-TAFRO flare compared to healthy donors. Together, these data suggest activation and involvement of CD8 T cell, NK cell, and monocyte subsets during iMCD-TAFRO flare. We next utilized Gene Set Enrichment Analysis (GSEA) of our single-cell transcriptomics dataset to ask whether circulating immune cell frequencies display enrichment of the 50 Hallmark gene sets during flare compared to remission across three iMCD-TAFRO patients. We found significant enrichment (FDR 〈 0.01) of genes within the Interferon Alpha Response gene set in circulating non-naïve CD8 T cells, classical monocytes, nonclassical monocytes, NK cells, and dendritic cells. These data suggest that a number of circulating immune cell populations may be responding to Type I interferon (IFN-I) during iMCD-TAFRO flare. In addition, our group has recently reported mTORC1 activation in iMCD and has characterized in three iMCD-TAFRO patients a clinical response following mTOR inhibition with sirolimus. Within our scRNAseq dataset, we identified mTORC1 signaling to be enriched only in circulating classical and nonclassical monocytes during flare. Having observed enrichment of both the Interferon Alpha Response Gene set and the mTORC1 signaling gene set in circulating monocytes, we then asked whether the relative expression of IFN-I response genes and mTORC1 signaling genes are correlated within circulating classical and nonclassical monocytes. Indeed, we observed a significant positive correlation between the average relative expression of mTORC1 signaling genes and IFN-I Response genes across classical, but not nonclassical, monocytes from all three iMCD-TAFRO patients (all R2 ≥0.6, p

Description: Human herpesvirus-8(HHV-8)-negative/idiopathic multicentric Castleman disease (iMCD) is a rare and poorly understood disorder diagnosed in ~1,000 individuals in the USA each year. It involves polyclonal lymphoproliferation, constitutional symptoms, systemic inflammation and an uncontrollable cytokine storm resulting in life-threatening multi-organ failure. Diagnosis and treatment can be difficult due to limited etiological understanding and heterogeneous presentation - clinical, laboratory, and histopathological abnormalities overlap with infectious, autoimmune and oncological diseases. iMCD symptoms and disease progression are largely believed to be driven by interleukin-6 (IL-6). However, approximately 66% of iMCD patients did not respond to anti-IL-6 therapy, siltuximab, the only FDA-approved iMCD therapy, in its phase II study (NCT01024036). Few treatment options exist for anti-IL-6 refractory patients because alternative driver cytokines and signaling pathways are not known. Herein we report the largest study to-date of iMCD serum proteomes with correlative anti-IL6 response data from 92 iMCD patients in disease flare (n=75 of which were collected as part of NCT01024036), in order to: (1) molecularly define iMCD, (2) identify predictors of response to anti-IL6 therapy, and (3) gain insights into the pathogenesis of iMCD. Proteomes of HHV8-positive MCD (n=20), Hodgkin lymphoma (n=20), rheumatoid arthritis (n=20) and healthy individuals (n=44) were also analyzed. Of the ~1,300 analytes measured using SomaLogic SOMAscan, 1,178 passed QC and were included in analyses. Each analyte was log2 transformed and capped at the 2.5th and 97.5th percentiles. Clinical and laboratory data collected at the time of sample draw were used to calculate disease activity following a modified CHAP scale: C-reactive protein, hemoglobin and albumin; missing performance status. Response to siltuximab was determined in NCT01024036. Data analysis was performed using the Medidata Rave Omics machine learning platform and R v3.4.4. Clustering of baseline proteomic data for iMCD patients identified six clusters that ranged in size from seven to 27 subjects. No associations with race, site, sex, age, or batch were found. Analytes identified among the strongest differentiators include cytokines, chemokines and inflammatory molecules. Interestingly, the largest cluster was associated with response to siltuximab (p

Description: Castleman disease (CD) describes a group of heterogeneous hematologic disorders with characteristic histopathological features. CD can present with unicentric or multicentric (MCD) regions of lymph node enlargement. Some cases of MCD are caused by human herpesvirus-8 (HHV-8), whereas others are HHV-8–negative/idiopathic (iMCD). Treatment of iMCD is challenging, and outcomes can be poor because no uniform treatment guidelines exist, few systematic studies have been conducted, and no agreed upon response criteria have been described. The purpose of this paper is to establish consensus, evidence-based treatment guidelines based on the severity of iMCD to improve outcomes. An international Working Group of 42 experts from 10 countries was convened by the Castleman Disease Collaborative Network to establish consensus guidelines for the management of iMCD based on published literature, review of treatment effectiveness for 344 cases, and expert opinion. The anti–interleukin-6 monoclonal antibody siltuximab (or tocilizumab, if siltuximab is not available) with or without corticosteroids is the preferred first-line therapy for iMCD. In the most severe cases, adjuvant combination chemotherapy is recommended. Additional agents are recommended, tailored by disease severity, as second- and third-line therapies for treatment failures. Response criteria were formulated to facilitate the evaluation of treatment failure or success. These guidelines should help treating physicians to stratify patients based on disease severity in order to select the best available therapeutic option. An international registry for patients with CD (ACCELERATE, #NCT02817997) was established in October 2016 to collect patient outcomes to increase the evidence base for selection of therapies in the future.

Description: Background Human herpes virus-8 (HHV8) negative/idiopathic multicentric Castleman disease (iMCD) is a rare hematological disorder with one FDA-approved treatment and no early indicators of patient response after treatment is commenced. Clinical onset can be sudden and severe, involving polyclonal lymphoproliferation with characteristic dysmorphic germinal centers, constitutional symptoms, systemic inflammation and life-threatening cytokine storm-driven multi-organ failure. CXCL13, a key regulator of lymph node germinal center development, was recently found to be the most elevated cytokine in iMCD flare, but the clinical significance of this finding is not yet clear. Interleukin-6 (IL-6) is the known driver of pathogenesis in a portion of patients and the target of the only FDA-approved treatment, siltuximab. In the Phase II study of siltuximab (NCT01024036), one-third of patients met response criteria, which were assessed after a minimum of 48 weeks. Early indicators of response to siltuximab are urgently needed to inform clinicians about the likelihood of patient response to therapy, adjust treatments if needed, and identify novel therapeutic targets for siltuximab non-responders. Methods Clinical data and serum samples were collected as part of NCT01024036. We measured serum protein analytes in the 52 subjects who were treated with anti-IL6 therapy, as well as the 26 patients in the control arm, at day 1, day 8, and day 64 of therapy (infusions administered every 21 days). Serum samples from 44 healthy donors were also analyzed. Of the 1,305 analytes measured using SomaLogic SOMAscan, 1,178 passed QC and were included in analyses. Each analyte was log2 transformed and capped at the 2.5th and 97.5th percentiles. Response to anti-IL6 therapy was determined by independent review in NCT01024036. Data processing was performed using the Medidata Rave Omics machine learning platform and R v3.4.4. Linear mixed effects models were used to detect whether kinetic changes in protein expression were associated with anti-IL6 response. Upon running the full model with the selected covariates, the p-values of the interaction between time point and response were used to test for differences between responders and non-responders. A separate model was fitted using each protein, and False Discovery Rates (FDR) were estimated by the Benjamini-Hochberg method with alpha 〈 0.05. Results Seven days after siltuximab was first administered (day 8), 9 proteins were significantly different between responders and non-responders: IgA, BCMA, NPS-PLA2, ART, IL-18 BPa, CD5L, b2-Microglobulin, CXCL13, and NRP1. All 9 of these proteins were significantly decreased in responders compared to non-responders. At day 64, the number of significantly different proteins increased to 121, including 8 of the 9 proteins from day 8; NPS-PLA2 did not achieve significance at day 64. This result indicates that there may be early indicators of response in serum as early as day 8. Given that CXCL13 was recently discovered as a key cytokine in iMCD, the early and significant decline of CXCL13 in responders versus non-responders was highly notable (Day 8: FDR = 0.02, Day 64: FDR = 0.005). Prior to treatment, CXCL13 was significantly higher in this cohort of iMCD patients than in a group of age-matched healthy donors (p = 8.19e-09). By day 64, CXCL13 levels in siltuximab responders decreased to levels approaching the healthy donor range but remained elevated in non-responders and placebo patients. Conclusions This analysis represents the first use of high-quality serum proteomics data to study early indicators of response to treatment in a rare hyperinflammatory, lymphoproliferative disorder. The decline in CXCL13 levels in responders and continued elevation in non-responders suggests that CXCL13 is downstream of IL-6 in responders and independent of IL-6 signaling in non-responders. CXCL13, along with several other proteins that demonstrated significant decline by day 8 including IgA and beta2-microglobulin, can be routinely measured and could serve as a panel that indicates the likelihood of response soon after commencing therapy, if validated in a separate cohort. These proteins could also provide a more continuous scale of response than traditional outcome measures. Given that iMCD may have a sudden and severe onset, early indicators of response to anti-IL6 therapy are critical for timely treatment administration. Figure Disclosures Katz: Medidata Solutions: Employment. Nabel:Thermo Fisher: Patents & Royalties: royalty income. Diamond:Brigham and Women's Hospital and Harvard Medical School's Department of Medicine, Division of Genetics, Genomes2People Translational Genomics Research Program: Consultancy. Karvir:Medidata Solutions: Employment. Zhang:Medidata Solutions: Employment. Reddy:Janssen Pharmaceuticals: Employment, Equity Ownership. Guilfoyle:Janssen Pharmaceuticals: Employment, Equity Ownership. Tendler:Janssen Pharmaceuticals: Employment, Equity Ownership. van Rhee:Takeda: Consultancy; Sanofi Genzyme: Consultancy; Castleman Disease Collaborative Network: Consultancy; EUSA: Consultancy; Adicet Bio: Consultancy; Kite Pharma: Consultancy; Karyopharm Therapeutics: Consultancy. Beineke:Medidata Solutions: Employment. Oromendia:Medidata Solutions: Employment, Equity Ownership. Fajgenbaum:Janssen Pharmaceuticals: Research Funding.

Description: Background Human herpes-virus 8-negative/idiopathic multicentric Castleman disease (iMCD) is a rare inflammatory disorder involving multicentric lymphadenopathy with characteristic histopathology. Clinical presentation is heterogenous and includes cytokine-driven constitutional symptoms, cytopenias, systemic inflammation, and multi-organ dysfunction. International consensus treatment guidelines are based on a large cohort of case studies and a few clinical trials, but the available evidence is limited. Siltuximab, an anti-IL-6 therapy, is the only FDA-approved treatment for iMCD; 34% of patients achieved durable symptomatic and tumor response in the phase II trial. Tocilizumab, an anti-IL-6 receptor therapy, is frequently used off label and demonstrated promising results in an open-label study in Japan. The treatment guidelines recommend siltuximab ± corticosteroids (CS) as first-line therapy for all iMCD patients and tocilizumab as a substitute when siltuximab is not available. Rituximab, a CD20 antibody, is recommended as an alternate first-line therapy in patients who are non-severe and do not exhibit marked cytokine-driven symptoms. In all other patients, rituximab is recommended second-line; however, it has never been systematically evaluated in iMCD. Chemotherapies, immunosuppressants, and immunomodulators are recommended second- or third-line, but again, effectiveness is not well described. Better understanding of treatment effectiveness is urgently needed. Herein, we describe treatment and response in a real-world cohort of iMCD patients. Methods Data were collected and abstracted for 68 patients enrolled in an on-going IRB-approved natural history study of Castleman disease. Diagnosis is graded by an expert panel of clinicians and pathologists on an on-going basis; patients unlikely to have iMCD were excluded from analysis (N=12). Of the 56 patients included, 37 (66%) are expert panel-confirmed and 19 (34%) are awaiting confirmation. Durable response is defined as achieving ≥50% improvement in the proportion of abnormal iMCD minor clinical and laboratory diagnostic criteria sustained for ≥1 year. Small sample size prevented statistical comparisons. Results Median age at diagnosis is 33 years (range: 1-65 years). The cohort is 52.9% female, 63% white, and 4 (7%) patients died. Thirty-three unique drugs, including anti-IL-6 therapies, CS, chemotherapies, immunosuppressants, and others have been administered across the 56 patients. Rituximab is the most frequently used drug, administered to 39 (70%) patients. Siltuximab (29 patients, 52%) and tocilizumab (19 patients, 34%) are the next two most frequently used targeted therapies. There was a 52% response (15/29) to regimens inclusive of siltuximab, 50% response (9/18) to those inclusive of tocilizumab, and 25% response (9/26) to those inclusive of rituximab. Siltuximab±CS induced response in 15/24 (63%) patients, tocilizumab±CS in 4/7 (57%), and rituximab±CS in 2/13 (15%). Among the 37 expert-confirmed iMCD patients, we found a 58% response (11/19) to regimens inclusive of siltuximab, 47% (8/17) to those inclusive of tocilizumab, and 27% (7/26) to those inclusive of rituximab. Further, in these patients, siltuximab±CS induced response in 11/16 (69%), tocilizumab±CS in 3/6 (50%), and rituximab±CS in 1/6 (17%) patients. Of note, 3 of 4 deceased patients received both anti-IL-6 therapy and rituximab and all 4 received chemotherapies and immunosuppressants but did not respond to any drug. Discussion These data reveal that despite there being one FDA-approved treatment, iMCD is treated with a variety of agents. Among the full cohort, siltuximab±CS demonstrated a 63% durable response, which was higher than the response reported in the clinical trial (not statistically compared). This may reflect differences in response criteria and/or disease activity of patients in clinical trials versus real world settings. Siltuximab and tocilizumab have never been systematically compared; in this cohort they demonstrated similar response. Considering the morbidity and mortality of iMCD, these data suggest that current therapies demonstrate important activity. However, additional agents are needed for refractory patients, who have few options and are at risk of death due to disease progression. Further data are needed to compare groups and identify optimal treatment protocols. Disclosures Liu: BridgeBio Pharma: Employment, Equity Ownership. Gibson:EUSA Pharma: Employment. Kanhai:EUSA Pharma: Employment. Martin:EUSA Pharma: Employment. Srkalovic:Takeda: Speakers Bureau; Janssen: Speakers Bureau; Foundation Medicine: Speakers Bureau; EUSA Pharma: Speakers Bureau. Uldrick:Patent: Patents & Royalties: co-inventor on US Patent 10,001,483 entitled ; Celgene: Other: research support from Celgene through a CRADA at the NCI; Roche: Other: commercial research support through a CTA with Fred Hutchinson Cancer Research Center; Merck: Other: drug for a clinical trial from Merck through a CRADA with the NCI. van Rhee:Takeda: Consultancy; Sanofi Genzyme: Consultancy; Karyopharm Therapeutics: Consultancy; EUSA: Consultancy; Adicet Bio: Consultancy; Kite Pharma: Consultancy; Castleman Disease Collaborative Network: Consultancy. Fajgenbaum:Janssen Pharmaceuticals: Research Funding. OffLabel Disclosure: Tocilizumab, a monoclonal antibody directed against IL-6-receptor, is approved for use in rheumatoid arthritis in the US. It is frequently used off-label in idiopathic multicentric Castleman disease (iMCD) and is recommended as a substitute first-line therapy in the International Consensus iMCD treatment guidelines. Rituximab, a monoclonal antibody directed against CD20, is used in rheumatoid arthritis and other autoimmune and cancerous disorders. It is frequently used off-label in iMCD and is recommended as an alternate first-line or a second-line therapy in the International Consensus iMCD treatment guidelines. Corticosteroids are used broadly in iMCD and are recommended as needed as useful adjunctive therapy in the International Consensus iMCD treatment guidelines.