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ImportanceTreatment of newly diagnosed multiple myeloma (NDMM) with a quadruplet regimen consisting of a monoclonal antibody, proteasome inhibitor, immunomodulatory imide, and corticosteroid has been associated with improved progression-free survival (PFS) compared with triplet regimens. The optimal quadruplet combination, and whether this obviates the need for frontline autologous stem cell transplant (ASCT), remains unknown. We evaluated elotuzumab and weekly carfilzomib, lenalidomide, and dexamethasone (Elo-KRd) without ASCT in NDMM.ObjectiveTo investigate the efficacy of Elo-KRd using a measurable residual disease (MRD)-adapted design in NDMM regardless of ASCT eligibility.Design, Setting, and ParticipantsThis multicenter, single-arm, phase 2 study enrolled patients between July 2017 and February 2021. Median follow-up was 29 months.InterventionsTwelve to 24 cycles of Elo-KRd; consecutive MRD-negative results at 10−6 by next-generation sequencing (NGS) after cycles 8 (C8) and 12 determined the duration of Elo-KRd. This was followed by Elo-Rd (no carfilzomib) maintenance therapy until disease progression.Main Outcomes and MeasuresThe primary end point was the rate of stringent complete response (sCR) and/or MRD-negativity (10−5) after C8 Elo-KRd. Secondary end points included safety, rate of response, MRD status, PFS, and overall survival (OS). As an exploratory analysis, MRD was assessed using liquid chromatography mass spectrometry (MS) on peripheral blood samples.ResultsForty-six patients were enrolled (median age 62 years, 11 [24%] aged >70 years). Overall, 32 (70%) were White, 6 (13%) were Black, 3 (6%) were more than 1 race, and 5 (11%) were of unknown race. Thirty-three (72%) were men and 13 (28%) were women. High-risk cytogenetic abnormalities were present in 22 (48%) patients. The rate of sCR and/or MRD-negativity after C8 was 26 of 45 (58%), meeting the predefined statistical threshold for efficacy. Responses deepened over time, with the MRD-negativity (10−5) rate increasing to 70% and MS-negativity rate increasing to 65%; concordance between MRD by NGS and MS increased over time. The most common (>10%) grade 3 or 4 adverse events were lung and nonpulmonary infections (13% and 11%, respectively). There was 1 grade 5 myocardial infarction. The estimated 3-year PFS was 72% overall and 92% for patients with MRD-negativity (10−5) at C8.Conclusions and RelevanceAn MRD-adapted design using elotuzumab and weekly KRd without ASCT showed a high rate of sCR and/or MRD-negativity and durable responses. This approach provides support for further evaluation of MRD-guided deescalation of therapy to decrease treatment exposure while sustaining deep responses.Trial RegistrationClinicalTrials.gov Identifier: NCT02969837

We conducted a phase 1/2 study of carfilzomib, pomalidomide, and dexamethasone (KPd) followed by another cohort of KPd with daratumumab (Dara-KPd) in relapsed/refractory multiple myeloma. The primary endpoints were identification of a maximum tolerated dose (MTD) of KPd for phase 1, and rates of overall response (ORR) and near complete response (nCR) after 4 cycles of KPd and Dara-KPd, respectively, for phase 2. The MTD for KPd was carfilzomib 20/27 mg/m2 on days 1, 2, 8, 9, 15, 16 (cycles 1-8) and days 1, 2, 15, 16 for cycles 9+; oral pomalidomide 4 mg on days 1-21; and oral dexamethasone 40 mg weekly in 28-day cycles. Sixty-six patients received KPd, including 34 at the MTD. The ORR after 4 cycles of KPd at the MTD was 27/34 (79%, 95% CI 62%-91%), meeting the statistical threshold for efficacy. At a median follow-up of 44 months, the median PFS and OS were 13 and 44 months, respectively. Twenty-eight patients received Dara-KPd. The rate of nCR or better after 4 cycles was 11/28 (39%, 95% CI 22%-59%), meeting the statistical threshold for efficacy. As best response to Dara-KPd, the ORR was 25/28 (89%) and the rate of measurable residual disease negativity by flow cytometry (10-5) was 17/26 (65%). At a median 26 months follow-up, the median PFS and OS for Dara-KPd were not reached. Dara-KPd induced deeper and more durable responses than KPd without compromising safety in a predominantly high risk, lenalidomide-refractory population, warranting further evaluation of this quadruplet for this patient population. This trial is registered at www.clinicaltrials.gov as NCT01665794.

In this phase 2 multicenter study, we evaluated the incorporation of autologous stem cell transplantation (ASCT) into a carfilzomib-lenalidomide-dexamethasone (KRd) regimen for patients with newly diagnosed multiple myeloma (NDMM). Transplant-eligible patients with NDMM received 4 cycles of KRd induction, ASCT, 4 cycles of KRd consolidation, and 10 cycles of KRd maintenance. The primary end point was rate of stringent complete response (sCR) after 8 cycles of KRd with a predefined threshold of ≥50% to support further study. Seventy-six patients were enrolled with a median age of 59 years (range, 40-76 years), and 35.5% had high-risk cytogenetics. The primary end point was met, with an sCR rate of 60% after 8 cycles. Depth of response improved over time. On intent-to-treat (ITT), the sCR rate reached 76%. The rate of minimal residual disease (MRD) negativity using modified ITT was 70% according to next-generation sequencing (

Multiple myeloma (MM) continues to be considered incurable, necessitating new drug discovery. The mitotic kinase T‐LAK cell‐originated protein kinase/PDZ‐binding kinase (TOPK/PBK) is associated with proliferation of tumor cells, maintenance of cancer stem cells, and poor patient prognosis in many cancers. In this report, we demonstrate potent anti‐myeloma effects of the TOPK inhibitor OTS514 for the first time. OTS514 induces cell cycle arrest and apoptosis at nanomolar concentrations in a series of human myeloma cell lines (HMCL) and prevents outgrowth of a putative CD138+ stem cell population from MM patient‐derived peripheral blood mononuclear cells. In bone marrow cells from MM patients, OTS514 treatment exhibited preferential killing of the malignant CD138+ plasma cells compared with the CD138− compartment. In an aggressive mouse xenograft model, OTS964 given orally at 100 mg/kg 5 days per week was well tolerated and reduced tumor size by 48%‐81% compared to control depending on the initial graft size. FOXO3 and its transcriptional targets CDKN1A (p21) and CDKN1B (p27) were elevated and apoptosis was induced with OTS514 treatment of HMCLs. TOPK inhibition also induced loss of FOXM1 and disrupted AKT, p38 MAPK, and NF‐κB signaling. The effects of OTS514 were independent of p53 mutation or deletion status. Combination treatment of HMCLs with OTS514 and lenalidomide produced synergistic effects, providing a rationale for the evaluation of TOPK inhibition in existing myeloma treatment regimens., Multiple myeloma (MM) is currently considered to be incurable. For the first time, we show that an inhibitor of T‐LAK cell‐originated protein kinase suppressed several MM‐supportive pathways and induced potent MM‐selective killing that synergized with a common component of current treatment regimens, making OTS514 a suitable candidate for targeting in a clinical setting.

Gut anaerobes protect against pathogen invasion Intestinal infections are a common problem for young animals. One explanation is that the protective gut microbiota is not fully established in infants. How the microbiota might protect against pathogens is unclear. Kim et al. found that members of the group of strictly anaerobic, spore-forming bacteria known as clostridia protect neonatal mice against diarrhea-causing pathogens. The protective effect is enhanced by giving mice the metabolite succinate in drinking water. Succinate favors colonization of the neonatal gut by cluster IV and XIVa clostridia and concomitantly excludes Salmonella typhimurium. Science , this issue p. 315

Introduction : tRNA-derived fragments (tRFs) have been postulated to play a significant role in human disease and specifically cancer. tRFs can function as small non-coding RNAs, expressed more highly in conditions of oxidative stress and tissue damage, and are postulated to have a regulatory function. Previous work has identified overexpression of tRNAs in multiple myeloma (MM), but the expression of tRFs in MM has not yet been characterized. In this descriptive study, we set out to apply modern next-generation sequencing tools to describe tRNA and tRF expression in MM. Methods: Bone marrow aspirate samples from 24 patients, 8 with monoclonal gammopathy of undetermined significance (MGUS) or smoldering MM (SMM), 8 with newly diagnosed MM (NDMM), and 8 with relapsed/refractory MM (RRMM) were selected. Total RNA fraction from mononuclear cell fractions was extracted and used to prepare tRNA sequencing libraries which generated data of different small RNA families such as tRNA, tRF, and 5S rRNA in one library. Results: We found a trend toward increased total expression of tRFs in NDMM and RRMM patient samples. Analysis of specific classes of tRFs revealed differential expression along the course of disease progression, with specific tRFs increasing in expression from MGUS/SMM to RRMM. Specifically, tRFs from tRNAHis showed a significant increase in expression from MGUS/SMM to NDMM (p Conclusions : These preliminary results indicate that tRNA fragment expression has the potential to serve as an objective, quantitative measure of clinical MM progression. While the function of these small RNA species remains unclear, specific cleavage patterns suggest that tRF expression in MM may be regulated during disease progression. Future work should explore potential for miRNA-like function of tRFs in MM, and ongoing work to be presented at the meeting will further explore tRF abundance as well as post-transcriptional modifications of tRNAs which may provide a mechanistic understanding of patterns of tRNA fragmentation. Citation Format: Alexandra E. Rojek, Christopher D. Katanski, Andrew Stefka, Benjamin A. Derman, Andrzej Jakubowiak, Tao Pan. tRNA expression and tRFs in multiple myeloma: Progression from monoclonal gammopathies to relapsed/refractory disease [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 2394.

8011 Background: The addition of a monoclonal antibody to triplet induction regimens in patients (pts) with MM with intent for autologous stem cell transplant (ASCT) has resulted in higher overall and deep response rates. In this study we are investigating the impact of the addition of Elo to KRd on complete response (CR) and/or MRD-negative rates in newly diagnosed MM regardless of transplant eligibility. Methods: Pts were enrolled from four MM Research Consortium sites into this phase 2 study. All patients receive 12 cycles of Elo-KRd in 28-day cycles: Elo per standard dosing, K 20/56/70 mg/m2 days 1, 8 and 15, R 25 mg days 1-21, and dexamethasone 40 mg days 1, 8, 15, 22. ASCT eligible candidates can undergo stem cell collection after cycle 4 and then resume treatment; pts who elect to proceed to ASCT are censored for response at that time. Pts MRD(-) (-5) by NGS after cycles 8 (C8) and 12 (C12) proceed to Elo-Rd until progression. Patients who convert from MRD(+) to MRD(-) between C8 and C12 receive an additional 6 cycles of Elo-KRd (total 18 cycles) followed by Elo-Rd, and pts MRD(+) after C12 receive an additional 12 cycles of Elo-KRd (total 24) followed by Elo-Rd. The primary endpoint of the study is sCR and/or MRD(-) rate after C8 E-KRd. MRD status was determined by ClonoSEQ next generation sequencing (NGS, -5) [Adaptive Biotechnologies]. An improvement in the sCR and/or MRD(-) rate by NGS from a historical 30% to 50% at the end of C8 will be considered promising. Results: 44 pts are enrolled, 39 of whom are evaluable for response (cutoff Jan 10 2021). Median age is 62 years (range 43-81, 23% age >70) and 23 (52%) have high-risk cytogenetic abnormalities (HRCA) including 13 (30%) with >2 high-risk abnormalities (6 pts unknown cytogenetics). 34/39 (87%) have MRD trackable by clonoSEQ. The rate of sCR and/or MRD(-) by NGS at the end of C8 is 19/33 (58%), meeting the statistical threshold for establishing efficacy (2 pts censored for elective ASCT before C8 and 4 pts receiving therapy but have not reached C8). With a median follow-up of 24 months, estimated 2-year progression free survival is 87% (100% for standard risk, 79% for HRCA) and estimated 2-year overall survival is 89% (82% for HRCA). No pt who was MRD(-) by NGS after C8 has progressed, including 6 pts with HRCA. Serious adverse events occurred in 30 pts (68%). 89% experienced treatment emergent AEs, the most common (>10%) of which was pneumonia (14%). One pt had grade 5 myocardial infarction. Conclusions: Elo-KRd demonstrates tolerability consistent with known toxicities of these agents and met the primary endpoint of sCR and/or MRD(-) of >50% after 8 cycles. With longer follow-up, the study results may validate that an MRD-adaptive design for de-escalation of therapy in MM can generate deep responses while reducing treatment exposure. Clinical trial information: NCT02969837.

Introduction: Lenalidomide (LEN) is a cornerstone in the treatment of newly diagnosed MM both as part of induction and maintenance therapy. As a result, most patients at first or second relapse are LEN exposed/refractory creating a need for effective 2nd line and salvage therapies. While no standard of care regimen has been established in early relapse, several therapies have been evaluated combining second generation immunomodulatory agents, proteasome inhibitors, and daratumumab. We have previously reported the results from the phase I/II trial of KPd demonstrating excellent efficacy in a LEN-refractory patient population. Here, we present the first efficacy and safety results from the cohort in which DARA was added to KPd (D-KPd). Methods: This is a multi-center, open-label, Phase 1b/2 study in subjects with relapsed MM with two sequential treatment cohorts: KPd and D-KPd. Eligibility criteria and KPd doses and schedules were identical for both cohorts. Subjects with measurable disease that progressed after at least 1 prior therapy (LEN refractory disease was required for 2nd-line therapy and LEN refractory/exposed for ≥ 3rd line) were eligible. The KPd cohort has completed enrollment (n=67) and results have been previously reported. The D-KPd cohort received treatment on the following 28-day schedule: Pomalidomide 4 mg daily on days 1-21 for cycles 1-8+, Carfilzomib 20/27 mg/m2 on days 1,2,8,9,15,16 for C1-8, then 1,2,15,16 for C9+ (maintenance), dexamethasone 20 mg days 1,2,8,9,15,16,22,23 for C1-2, then 40 mg days 1,8,15,22 for C3+. DARA was administered as per standard schedule, weekly for the first 2 cycles, then every 2 weeks for cycles 3-6, and monthly thereafter. A Minimax two-stage design was employed for enrollment of subjects on this cohort. Twenty-one patients were required to accrue to the first stage, with at least 4 responders of ≥nCR at 4 cycles necessary to accrue to second stage for a total of 34 pts. Primary endpoint was rate of nCR/CR as per IMWG criteria. Minimal residual disease (MRD) was evaluated by 10-color flow cytometry with limit of detection (LOD) 10-4-10-5 and will also be assessed by next-generation sequencing (LOD 10-5-10-6). Secondary endpoints include overall response rate, depth of response, progression-free survival (PFS), and overall survival. Per study design, a nCR/CR rate of >35% (over the historical 20% rate) would support further study of the D-KPd regimen. Results: As of July 29, 2020, all 22 subjects who were enrolled into the D-KPd cohort were evaluable for safety and the primary endpoint. Median age was 62 (range 37-74) with a median of 1 (range 1-3) prior lines of therapy. 81% of patients were LEN refractory, and high-risk cytogenetics per IMWG criteria were present in 12/19 (68%) evaluable patients. Subjects completed a median of 12.5 cycles (range 2-33) of therapy and 21 (95%) subjects completed at least 4 cycles; 1 subject progressed after cycle 2. In the ITT population (n=22), after 4 cycles, 86% achieved ≥PR and 46% ≥nCR, warranting further enrollment to a second stage. At best response, the ≥PR was 86% with 55% ≥nCR/CR, 45% ≥sCR, and 55% MRD negativity by flow cytometry (n=22). The most common grade 3-4 hematologic adverse events included neutropenia (64%), lymphopenia (36%), and febrile neutropenia (18%). The most common grade 3-4 non-hematologic adverse events included fatigue (27%), respiratory infections (23%), diarrhea (14%), and insomnia (14%). There was one thrombotic event (4.5%) which was grade 2. In comparison to the KPd cohort (67 patients with similar baseline characteristics), there was an improvement in efficacy as demonstrated by an increase in rate of ≥nCR at the end of 4 cycles (from 7% to 46%), as well as the best response (from 20% to 55%). High risk cytogenetics did not significantly affect response (≥nCR 46% at best response [all sCR]). With 20 months of follow-up, median PFS was not reached in the D-KPd cohort and 12-month PFS is 84% vs 63% for KPd. Rates of grade 3/4 cytopenias were higher in the D-KPd cohort. There was no treatment related mortality and 19 of 22 pts are alive. Conclusion: D-KPd demonstrates high efficacy in a population of patients with relapsed/refractory multiple myeloma enriched for high risk cytogenetics. MRD negativity by flow cytometry was achieved in 55% of subjects. The ≥nCR rate of 55% with D-KPd compares favorably to the 20% rate with KPd alone; based on the study design, this warrants further evaluation of D-KPd. Disclosures Zonder: BMS, Celgene: Research Funding; Intellia, Amgen, Takeda, Janssen, Regeneron, Alnylam, Caelum, Oncopeptides: Consultancy. Reece:Janssen, Bristol-Myers Squibb, Amgen, Takeda: Consultancy, Honoraria; Janssen, Bristol-Myers Squibb: Membership on an entity's Board of Directors or advisory committees; Merck: Honoraria, Research Funding; Otsuka: Research Funding. Berdeja:Kesios: Research Funding; Karyopharm: Consultancy; Acetylon: Research Funding; Vivolux: Research Funding; Abbvie: Research Funding; Amgen: Consultancy, Research Funding; Bioclinica: Consultancy; BMS: Consultancy, Research Funding; Celgene: Consultancy, Research Funding; Cellularity: Research Funding; Constellation: Research Funding; CURIS: Research Funding; EMD Sorono: Research Funding; Genentech, Inc.: Research Funding; Glenmark: Research Funding; Janssen: Consultancy, Research Funding; Kite Pharma: Consultancy; Legend: Consultancy; Lilly: Research Funding; Novartis: Research Funding; Poseida: Research Funding; Takeda: Consultancy, Research Funding; Teva: Research Funding; Prothena: Consultancy; Servier: Consultancy; Bluebird: Research Funding; CRISPR Therapeutics: Consultancy, Research Funding. Jakubowiak:Adaptive, Juno: Consultancy, Honoraria; AbbVie, Amgen, BMS/Celgene, GSK, Janssen, Karyopharm: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees.

Environmentally induced alterations in the commensal microbiota have been implicated in the increasing prevalence of food allergy. We show here that sensitization to a food allergen is increased in mice that have been treated with antibiotics or are devoid of a commensal microbiota. By selectively colonizing gnotobiotic mice, we demonstrate that the allergy-protective capacity is conferred by a Clostridia-containing microbiota. Microarray analysis of intestinal epithelial cells from gnotobiotic mice revealed a previously unidentified mechanism by which Clostridia regulate innate lymphoid cell function and intestinal epithelial permeability to protect against allergen sensitization. Our findings will inform the development of novel approaches to prevent or treat food allergy based on modulating the composition of the intestinal microbiota.

Tregs play a crucial role in the maintenance of intestinal immune homeostasis. However, significant numbers of Foxp3+ Tregs accumulate in the inflamed lesions in experimental colitis and in IBD patients. Treg production of the proinflammatory cytokines IFN-γ and/or IL-17 may arguably explain their ineffectiveness in suppressing intestinal inflammation. However, it remains unknown whether iTreg and tTreg produce proinflammatory cytokines and how TLR signaling regulates this process. Here, we found that Foxp3+Tregs were increased in the intestines of B6.TLR4−/− and B6.IL-10−/− mice when compared with WT B6 mice. TLR4−/− and IL-10−/− resulted in more Tregs within inflamed intestines. The majority of Foxp3+ Tregs in the spleen was Helios+Nrp1+, whereas most Foxp3+ Tregs in the intestinal LP were Helios−Nrp1−. More Helios+Nrp1+ Tregs expressed IFN-γ and/or IL-17 than did Helios−Nrp1− Tregs in the spleen and intestine, which was increased with TLR4−/−. TLR4 signaling in T cells and APCs inhibited Foxp3+ induction via MyD88-dependent, TRIF-independent pathways, which was negatively regulated by SOCS3. Collectively, these data demonstrate Helios+Nrp1+ tTregs and Helios−Nrp1− iTregs produce proinflammatory cytokines in the intestines during inflammation, which was regulated by TLR4 signaling.

Dietary intervention with extensively hydrolyzed casein formula supplemented with Lactobacillus rhamnosus GG (EHCF+LGG) accelerates tolerance acquisition in infants with cow's milk allergy (CMA). We examined whether this effect is attributable, at least in part, to an influence on the gut microbiota. Fecal samples from healthy controls (n=20) and from CMA infants (n=19) before and after treatment with EHCF with (n=12) and without (n=7) supplementation with LGG were compared by 16S rRNA-based operational taxonomic unit clustering and oligotyping. Differential feature selection and generalized linear model fitting revealed that the CMA infants have a diverse gut microbial community structure dominated by Lachnospiraceae (20.5±9.7%) and Ruminococcaceae (16.2±9.1%). Blautia, Roseburia and Coprococcus were significantly enriched following treatment with EHCF and LGG, but only one genus, Oscillospira, was significantly different between infants that became tolerant and those that remained allergic. However, most tolerant infants showed a significant increase in fecal butyrate levels, and those taxa that were significantly enriched in these samples, Blautia and Roseburia, exhibited specific strain-level demarcations between tolerant and allergic infants. Our data suggest that EHCF+LGG promotes tolerance in infants with CMA, in part, by influencing the strain-level bacterial community structure of the infant gut.

Transplantation is the only cure for end-stage organ failure, but without immunosuppression, T cells rapidly reject allografts. While genetic disparities between donor and recipient are major determinants of the kinetics of transplant rejection, little is known about the contribution of environmental factors. Because colonized organs have worse transplant outcome than sterile organs, we tested the influence of host and donor microbiota on skin transplant rejection. Compared with untreated conventional mice, pretreatment of donors and recipients with broad-spectrum antibiotics (Abx) or use of germ-free (GF) donors and recipients resulted in prolonged survival of minor antigen-mismatched skin grafts. Increased graft survival correlated with reduced type I IFN signaling in antigen-presenting cells (APCs) and decreased priming of alloreactive T cells. Colonization of GF mice with fecal material from untreated conventional mice, but not from Abx-pretreated mice, enhanced the ability of APCs to prime alloreactive T cells and accelerated graft rejection, suggesting that alloimmunity is modulated by the composition of microbiota rather than the quantity of bacteria. Abx pretreatment of conventional mice also delayed rejection of major antigen-mismatched skin and MHC class II-mismatched cardiac allografts. This study demonstrates that Abx pretreatment prolongs graft survival, suggesting that targeting microbial constituents is a potential therapeutic strategy for enhancing graft acceptance.

SummaryBackground The role of the fungal microbiota in digestive diseases is poorly defined, but is becoming better understood due to advances in metagenomics. Aim To review the gastrointestinal fungal microbiota and its relationship with digestive diseases. Methods Search of the literature using PubMed and MEDLINE databases. Subject headings including ‘fungal-bacterial interactions’, ‘mycotoxins’, ‘immunity to fungi’, ‘fungal infection’, ‘fungal microbiota’, ‘mycobiome’ and ‘digestive diseases’ were used. Results The fungal microbiota is an integral part of the gastrointestinal microecosystem with up to 106 microorganisms per gram of faeces. Next-generation sequencing of the fungal 18S rRNA gene has allowed better characterisation of the gastrointestinal mycobiome. Numerous interactions between fungi and bacteria and the complex immune response to gastrointestinal commensal or pathogenic fungi all impact on the pathophysiology of inflammatory bowel disease and other gastrointestinal inflammatory entities such as peptic ulcers. Mycotoxins generated as fungal metabolites contribute to disturbances of gastrointestinal barrier and immune functions and are associated with chronic intestinal inflammatory conditions as well as hepatocellular and oesophagogastric cancer. Systemic and gastrointestinal disease can also lead to secondary fungal infections. Fungal genomic databases and methodologies need to be further developed and will allow a much better understanding of the diversity and function of the mycobiome in gastrointestinal inflammation, tumourigenesis, liver cirrhosis and transplantation, and its alteration as a consequence of antibiotic therapy and chemotherapy. Conclusions The fungal microbiota and its metabolites impact gastrointestinal function and contribute to the pathogenesis of digestive diseases. Further metagenomic analyses of the gastrointestinal mycobiome in health and disease is needed.

Introduction In a phase 1/2 study designed to assess KRd w/o ASCT, KRd provided a high rate of stringent complete response (sCR) (55%) in NDMM patients (pts) after a median of 24 cycles and 47.5 months (mo) of follow-up (f/u), with 4-year (yr) progression-free survival (PFS) and overall survival (OS) rates of 64% and 93%, respectively (Jakubowiak et al. Blood. 2012;120:1801-9; Jakubowiak et al, EHA 2016). To further improve response and outcomes, we designed a phase 2 study to assess KRd with ASCT. Methods The study enrolled ASCT-eligible pts with NDMM requiring tx per International Myeloma Working Group (IMWG) criteria with no age limitation. Pts received initial four 28-day cycles of KRd induction with CFZ IV 20/36 mg/m2 on Days (D) 1, 2, 8, 9, 15, 16 (20 mg/m2 given on D1, 2 in Cycle [C] 1 only); LEN PO D1-21 at 25 mg, DEX PO 40 mg/week followed by stem cell collection using G-CSF and plerixafor, melphalan 200 mg/m2 and ASCT, and KRd consolidations (C 5-8) using the same doses and schedule except LEN 15 mg in C5 with option to escalate to prior dose and DEX reduced to 20 mg weekly. After C8, pts received maintenance KRd for an additional 10 cycles using the same doses as in C8 except CFZ on D 1, 2, 15, 16 only. Single-agent LEN was recommended off-study after C18. The primary endpoint was rate of sCR at the end of C8, with minimal residual disease (MRD) among secondary endpoints. We estimated that an sCR rate of 50% or better for KRd plus ASCT would indicate superior outcome compared with a historical sCR rate of 30% for KRd w/o ASCT at this time point. Responses were assessed using current IMWG criteria. MRD was evaluated by 10-color multiparameter flow cytometry (MFC) with 10-4-10-5 sensitivity and by next generation sequencing (NGS) at landmark time-points: after KRd induction (after C4), ASCT, and KRd consolidation (after C8); at the end of KRd tx (after C18); and then yearly. NGS analysis was done using the immunoSEQ® MRDplatform with a threshold at 10-6 for MRD negativity. MRD-negative status required CR, as per current IMWG criteria. Results As of July 1, 2016, enrollment was completed (76 pts); 72 pts completed KRd induction, 71 ASCT, 66 KRd consolidation, and 44 KRd maintenance with 25 pts remaining on protocol tx. Median age was 59 y (range 40-76), ISS stage II/III 57%, high-risk cytogenetics 36% as per IMWG criteria. Efficacy and toxicity data were available for 73 pts. Based on January 1, 2016 cut-off date, response rates at the end of C8 were 96% VGPR, 73% CR, and 69% sCR. The rate of sCR has been improving during the post-transplant phase of the KRd tx, from 20% post-ASCT to 69% after 4 cycles of KRd consolidation (C8), and to 82% after 10 additional cycles of KRd maintenance (C18). MRD rates in pts evaluated to date were 82% by MFC (N=33) and 66% by NGS (N=29) at the end of C8, and 90% (N=20) and 71% (N=16), respectively, at the end of C18. MRD rates in a subset of high-risk pts evaluated for MRD were 90% by MFC (10 of 11 pts with high-risk) and 63% by NGS (6 of 8 pts) at the end of C8, and 100% (6 of 6 pts) by MFC and 80% (4 of 5) by NGS at the end of C18. After median f/u of 17.5 mo, 2-yr PFS was 97% and 2-yr OS 99% for all 76 pts. For NGS and/or MFC MRD-negative pts at the end of C8, 2-yr PFS/OS was 100% and for MRD-positive/unknown PFS was 93% and OS 98%. For high-risk disease pts (N=27), 2-yr PFS was 96%. KRd-related adverse events (AEs) were generally Grade (G) 1/2, and included (any G) for hematologic AEs thrombocytopenia (57%), lymphopenia (39%), anemia (39%), and neutropenia (28%) and for non-hematologic AEs fatigue (53%), peripheral neuropathy (39%), diarrhea (3%), and infection (34%). Most common G3/4 AEs were lymphopenia (28%), neutropenia (18%), and infections (8%). Two of 71 pts evaluated pre-transplant had asymptomatic decrease of ejection fraction (EF) 45-50%, with no baseline ECHO or MUGA, all remaining pts had normal pre-transplant EF. Updated results, including larger sample of MRD data, will be presented at the meeting, with nearly all patients completing 18 KRd cycles at the next data cut. Conclusions These results show that extended KRd tx with incorporated ASCT results in high rates of sCR and MRD-negative disease in both standard and high risk disease, which correspond to high rates of PFS and OS. These results compare favorably with data from the KRd w/o ASCT study, based on pre-specified improvement of sCR rate at the end of 8 cycles and beyond, and with historical studies in NDMM. These results will require validation in ongoing and planned randomized trials. Disclosures Vij: Janssen: Honoraria; Amgen: Honoraria, Research Funding; Karyopharm: Honoraria; Celgene: Consultancy; Takeda: Honoraria, Research Funding; Novartis: Honoraria; Bristol-Myers Squibb: Honoraria. Reece:Merck: Research Funding; Otsuka: Honoraria, Research Funding; BMS: Honoraria, Research Funding; Celgene: Consultancy, Honoraria, Research Funding; Janssen: Consultancy, Honoraria, Research Funding; Takeda: Consultancy, Honoraria, Research Funding; Novartis: Honoraria, Research Funding; Amgen: Consultancy, Honoraria, Research Funding. Berdeja:Abbvie, Acetylon, Amgen, Bluebird, BMS, Calithera, Celgene, Constellation, Curis, Epizyme, Janssen, Karyopharm, Kesios, Novartis, Onyx, Takeda, Tragara: Research Funding. Rosenbaum:Celgene: Speakers Bureau. Richardson:Celgene: Membership on an entity's Board of Directors or advisory committees. Dytfeld:Janssen Poland: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Celgene: Research Funding; Amgen: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Novartis: Membership on an entity's Board of Directors or advisory committees. Jakubowiak:BMS: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees; Karyopharm: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees; SkylineDx: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees; Takeda: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees; Sanofi: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees; Amgen Inc.: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees; Celgene: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees.

Currently considered incurable, multiple myeloma (MM) is characterized by proliferation of malignant plasma cells (PC) predominantly in the bone marrow, which overproduce monoclonal immunoglobulin proteins, and a perturbed tumor microenvironment, which promotes PC survival, inhibits osteoblast activity, increases osteoclast activity, and leads to hallmark osteolytic bone disease, all of which contribute to the clinical manifestations of the disease. Disrupting these sequelae by incorporation of proteasome inhibitors and immunomodulatory drugs into treatment regimens has improved overall survival of MM patients; however, the majority of patients will become refractory to the most effective currently available therapeutic options and relapse.1 It is hypothesized that a drug-resistant population of myeloma stem cells promotes disease relapse, but the characteristics and identity of this cell type(s) remain uncertain.2 Expression of maternal embryonic leucine zipper kinase (MELK) is increased in a number of cancers and is associated with poorer prognosis. MELK activity modulates many cellular and biological processes, including proliferation, apoptosis, hematopoiesis and oncogenesis, and is believed to have a critical role in cancer stem cell maintenance.3

The incidence of food allergy in developed countries is rising at a rate that cannot be attributed to genetic variation alone. In this review, we discuss the environmental factors that may contribute to the increasing prevalence of potentially fatal anaphylactic responses to food. Decreased exposure to enteric infections due to advances in vaccination and sanitation, along with the adoption of high-fat (Western) diets, antibiotic use, Cesarean birth, and formula feeding of infants, have all been implicated in altering the enteric microbiome away from its ancestral state. This collection of resident commensal microbes performs many important physiological functions and plays a central role in the development of the immune system. We hypothesize that alterations in the microbiome interfere with immune system maturation, resulting in impairment of IgA production, reduced abundance of regulatory T cells, and Th2-skewing of baseline immune responses which drive aberrant responses to innocuous (food) antigens.

In an effort to develop a molecular classification scheme for Crohn's disease (CD), mucosal biopsies from 69 CD patients and 28 normal controls were analyzed for expression of the RelA subunit of nuclear factor (NF)-kappaB, A20 (a negative regulator of NF-kappaB), polymeric immunoglobulin receptor (pIgR), tumor necrosis factor (TNF), and interleukin (IL)-8. Principal component analysis was used to classify individuals into three subsets based on patterns of biomarker expression. Set 1 included normal subjects and CD patients with mild disease and good responses to therapy, thus defining "normal" biomarker expression. CD patients in set 2, characterized by low expression of all five biomarkers, had moderate to severe disease and poor responses to immunosuppressive and anti-TNF therapy. Patients in set 3, characterized by low expression of RelA, A20, and pIgR, normal TNF and elevated IL-8, had acute inflammation that responded well to therapy. Classification of CD patients by these biomarkers may predict disease behavior and responses to therapy.

Background & Aims The commensal microbiota is believed to have an important role in regulating immune responsiveness and preventing intestinal inflammation. Intestinal microbes produce signals that regulate inflammation via Toll-like receptor (TLR) signaling, but the mechanisms of this process are poorly understood. We investigated the role of the anti-inflammatory cytokine interleukin (IL)-10 in this signaling pathway using a mouse model of colitis. Methods Clinical, histopathologic, and functional parameters of intestinal inflammation were evaluated in TLR4 −/− , IL-10 −/− , and TLR4 −/− × IL-10 −/− mice that were free of specific pathogens and in TLR4 −/− × IL-10 −/− mice following eradication and reintroduction of Helicobacter hepaticus. Regulatory T-cell (Treg) function was evaluated by crossing each of the lines with transgenic mice that express green fluorescent protein under control of the endogenous regulatory elements of Foxp3 . Apoptotic cells in the colonic lamina propria were detected by a TUNEL assay. Results TLR4-mediated signals have 2 interrelated roles in promoting inflammation in TLR4 −/− × IL-10 −/− mice. In the absence of TLR4-mediated signals, secretion of proinflammatory and immunoregulatory cytokines is dysregulated. Tregs (Foxp3 + ) that secrete interferon-γ and IL-17 accumulate in the colonic lamina propria of TLR4 −/− × IL-10 −/− mice and do not prevent inflammation. Aberrant control of epithelial cell turnover results in the persistence of antigen-presenting cells that contain apoptotic epithelial fragments in the colonic lamina propria of Helicobacter- infected TLR4 −/− mice. Conclusions In mice that lack both IL-10- and TLR4-mediated signals, aberrant regulatory T-cell function and dysregulated control of epithelial homeostasis combine to exacerbate intestinal inflammation.

Secretory IgA serves as the first line of antigen-specific immunity in the intestine, protecting against pathogens while preventing inflammation in response to commensal microbes. Intestinal epithelial cells (IEC) regulate IgA secretion by controlling expression of the polymeric immunoglobulin receptor (pIgR). Many pro- and anti-inflammatory factors, including pIgR, are transcriptional targets of Toll-like receptor (TLR) signaling in response to molecular patterns from resident microbes. MyD88 is an important TLR signaling adaptor that has been implicated in regulation of intestinal inflammation. To assess the role of MyD88 in IEC gene expression, we compared mRNA levels of pIgR, TNF, and two regulatory factors, A20 and MKP-1, in the small intestines and colons of wild-type and MyD88−/− mice. Gene expression was significantly higher in the colon than in the small intestine of all mice, correlating with the higher microbial load. In the colon, expression of all genes was significantly higher in wild-type than in MyD88−/−mice. We conclude that commensal microorganisms set the tone of intestinal gene expression through MyD88-dependent TLR signaling. Supported by Crohn’s & Colitis Foundation of America.