Bruton Tyrosine Kinase Inhibitor Ibrutinib Research Articles

COSMOS: MOR208 plus idelalisib or venetoclax in patients with relapsed or refractory (R/R) chronic lymphocytic leukemia (CLL) or small lymphocytic lymphoma (SLL) previously treated with a Bruton's tyrosine kinase inhibitor (BTKi)—A two-cohort phase II study.

TPS7567 Background: Patients (pts) with R/R CLL who discontinue treatment with the BTKi ibrutinib due to progression have a particularly dismal prognosis. A phase I study showed that the Fc-enhanced, humanized, CD19 antibody MOR208 was well tolerated with encouraging single-agent activity in pts with R/R CLL/SLL. In preclinical models, MOR208 showed synergy with idelalisib (an inhibitor of PI3K delta) and venetoclax (an inhibitor of BCL-2), both approved for the treatment of CLL. Methods: This two-cohort, phase II study will investigate MOR208 combined with idelalisib (cohort A) or venetoclax (cohort B) in pts with R/R CLL or R/R SLL and includes a safety run-in phase for each cohort, to be evaluated by an Independent Data Monitoring Committee. Key inclusion criteria: aged ≥18 years, R/R CLL/SLL while receiving a BTKi therapy or intolerance of such therapy, BTKi administered as a single-agent or in combination for at least 1 month as the most recent prior anticancer therapy, ECOG performance status of 0–2, and adequate organ function. Key exclusion criteria: transformed CLL/SLL or Richter’s syndrome, BTKi treatment within 5 days prior to study drug dosing, prior treatment with a CD19-targeted therapy, a PI3K inhibitor (cohort A) or a BCL-2 inhibitor (cohort B). Pts will be treated for a maximum of 24 (28-day) cycles or until disease progression. Treatment will be MOR208 12 mg/kg IV (weekly for the first 3 months, every second week for the next 3 months, and monthly thereafter) in combination with oral idelalisib 150 mg twice-daily or venetoclax administered on a weekly ramp-up dosing schedule to the recommended daily dose of 400 mg. Primary endpoint: overall response rate based on independent review; secondary and exploratory endpoints include: progression-free and overall survival, duration of response, safety, pharmacokinetics, MOR208 immunogenicity, quality of life and minimal residual disease negativity. 120 pts per cohort are planned. Clinical trial information: 2015-002915-14.

Computational Regression and Prediction Analysis on a Dataset of 52 Pyrrolo[2,3-b]Pyridines and Pyrimidines Rheumatoid Arthritis Inhibitors

Objectives: To evaluate the dependency of physico-chemical properties of a set of BTK inhibitors on activity by linear regression analysis. Methods/Statistical Analysis: A multivariate regression analysis was implemented on 52 Pyrrolo[2,3-b]pyridines and pyrimidines which are reported as Bruton's Tyrosine Kinase (BTK) inhibitors to construct a regression model using complete data set in the relationship between dependent variable and independent variable was estimated using python based regression analysis. Leverages were calculated in order to exclude outlying data from analysis Findings: A linear regression analysis on a complete dataset of BTK inhibitors as dependent variables and few independent variables resulted in F-test: 4.87, r value: 0.737 and r2 value of 0.543, respectively. The dataset investigated for the existence of outliers resulted in 50 BTK inhibitors after excluding 4q and 4zd from the dataset which resulted in improved r2 of 0.701 with better statistics. Further 44 compound training set resulted in improved r and r2 coefficients such as r: 0.816 and r2: 0.666, respectively and applying on a 6 compound validation set which determines regression model reliability and significance. Application/Improvements: Application of regression model to screen novel compounds with decreased H-bond acceptors, logP and KAlpha2 followed by an increase in KAlpha3 and randic index would enhance inhibitory activity against BTK. 1. Introduction Rheumatoid Arthritis (RA) is an autoimmune disease categorized by auto antibody circulation in plasma, inflammation in synovial, destruction of cartilage and bone etc.1,2 Bruton's Tyrosine Kinase (BTK) belongs to Tec kinase member family of enzymes which are necessary for activation of B-cells.3 A designated function for BTK is found in activation of cells mediated by BCR and cell survival.4-6 Compounds which inhibit BTK are known to hinder B cell receptor signaling7, several such inhibitors are currently in clinical trials and the first BTK inhibitor Ibrutinib was approved for treating mantle cell lymphoma. 8 Efforts from medicinal chemistry, synthesis, crystallography has led to the discovery of some non-covalent BTK inhibitory compounds. Several BTK inhibitors such as Pyrrolo[2,3-b]pyridines9, Pyrrolo[2,3-d]pyrimidines10, thieno[3,2-c]pyridin-4-amines11, phenylpyridin-2(1H)- ones12, Imidazo[1,5-a]quinoxalines13, Purine derivatives14, CarbazoleCarboxamides15, PyridazinoneAnalogs16, Diaminopyrimidines17, imidazo[1,5‑a]pyrazines18 are reported. In this paper, linear regression analysis was implemented to study the dependency of physico-chemical properties on

The regulation of tumor‐suppressive microRNA, miR‐126, in chronic lymphocytic leukemia

The introduction of miR profiling of chronic lymphocytic leukemia (CLL) patients with different cytogenetic profiles and responses to therapy has allowed incorporation of important miR‐mRNA interactions into the understanding of disease biology. In this study, we performed miR expression analysis using NanoString nCounter to discover differentially regulated miRs after therapy with the Bruton tyrosine kinase inhibitor ibrutinib. Of the differentially regulated miRs in the discovery set, miR‐29c and miR‐126 were confirmed using real‐time PCR to be upregulated in CLL patient cells with ibrutinib therapy. In the validation set, an inverse correlation was observed between miR‐126 levels and expression of its putative target p85β, an isoform of the phosphoinositide 3‐kinase p85 regulatory subunit. We found that mRNA for the host gene EGFL7, primary unprocessed miR‐126, and mature miR‐126 are all downregulated in CLL cells compared to normal B cells. Patients in later stages of disease have a greater decrease in miR‐126 expression compared to treatment‐naive patients, indicating that lower miR‐126 levels may associate with disease progression. Overexpression of miR‐126 in leukemia cell lines significantly downregulates p85β expression and decreases activation of prosurvival mitogen‐activated protein kinase (MAPK) signaling. These results implicate miR‐126 in the pathology of CLL.

IMYC: Proof-of-concept study of ibrutinib in c-MYC and HER2 amplified oesophagogastric carcinoma.

TPS221 Background: The MYC proto-oncogene is among the most frequently dysregulated genes in human cancer and is amplified in 10-30% of oesophagogastric (OG) cancers. Such genes which code for transcription factors are challenging to target directly with small molecule inhibitors or monoclonal antibodies. High-throughput siRNA screening of OG cell lines has found silencing of Bruton’s tyrosine kinase (BTK) to result in selective lethality in the presence of MYC amplification. Sensitivity to the orally available BTK inhibitor ibrutinib has been confirmed in-vitro in MYC, HER2 and co-amplified OG cancer cell lines (Chong, Ann Onc 2014), suggesting it may be a potentially effective therapeutic strategy in both MYC and HER2 amplified OG cancer. Methods: i MYC is an open label, phase II non-randomised study to assess the efficacy of ibrutinib in advanced pre-treated OG cancer. A novel FISH assay for tumour MYC amplification has been developed and pts are pre-screened for MYC and HER2 amplification. Eligibility includes metastatic or locally advanced inoperable OG cancer (SCC or adeno), MYC (ratio >2.5) and/or HER2 (ratio>2) amplification, progression after at least 1 prior line of chemotherapy +/- trastuzumab for advanced disease, PS 0-2 and no history of significant cardiovascular or bleeding disorders. Of the first 9 pts, at least 4 will demonstrate MYC amplification and remaining 5 will show either MYC or HER2 amplification, or co-amplification of both. Pts will be treated with ibrutinib monotherapy until progression or unacceptable toxicity. A Simon 2 stage design will be used for the primary endpoint of response rate, with interim analysis after 9 pts and maximum recruitment to 17 pts. Secondary endpoints include PFS, OS and exploratory translational and imaging biomarker analysis. Mandatory biopsy will be obtained at baseline and optional biopsies will be undertaken at day 10-14, week 8 and progression. The trial represents the first attempt at targeting MYC amplification via synthetic lethal gene interactions in OG cancer through the novel application of an existing anti-cancer drug. As of September 2016, 61 patients have consented for pre-screening, 19 completed MYC analysis and first patient has started treatment. Clinical trial information: 02884453.

Leukemia cell proliferation and death in chronic lymphocytic leukemia patients on therapy with the BTK inhibitor ibrutinib.

BACKGROUND. Ibrutinib is an effective targeted therapy for patients with chronic lymphocytic leukemia (CLL) that inhibits Bruton's tyrosine kinase (BTK), a kinase involved in B cell receptor signaling. METHODS. We used stable isotopic labeling with deuterated water (2H2O) to measure directly the effects of ibrutinib on leukemia cell proliferation and death in 30 patients with CLL. RESULTS. The measured average CLL cell proliferation ("birth") rate before ibrutinib therapy was 0.39% of the clone per day (range 0.17%-1.04%); this decreased to 0.05% per day (range 0%-0.36%) with treatment. Death rates of blood CLL cells increased from 0.18% per day (average, range 0%-0.7%) prior to treatment to 1.5% per day (range 0%-3.0%) during ibrutinib therapy, and they were even higher in tissue compartments. CONCLUSIONS. This study provides the first direct in vivo measurements to our knowledge of ibrutinib's antileukemia actions, demonstrating profound and immediate inhibition of CLL cell proliferation and promotion of high rates of CLL cell death. TRIAL REGISTRATION. This trial was registered at clinicaltrials.gov (NCT01752426). FUNDING. This study was supported by a Cancer Center Support Grant (National Cancer Institute grant P30 CA016672), an NIH grant (CA081554) from the National Cancer Institute, MD Anderson's Moon Shots Program in CLL, and Pharmacyclics, an AbbVie company.

Simultaneous Inhibition of PI3Kδ and PI3Kα Induces ABC-DLBCL Regression Via Attenuation of BCR-Dependent and Independent Activation of NF-Kb and AKT

Introduction: ABC-DLBCL is a subtype of DLBCL with less favorable clinical outcomes to the standard of care (SoC) therapies. Constitutive activation of NF-κB by various genetic alterations in ABC-DLBCL has been identified as one of the key mechanisms driving chemotherapy resistance. Inhibition of B cell receptor (BCR) signaling with BTK (Bruton’s tyrosine kinase) inhibitor ibrutinib demonstrated encouraging clinical responses in ABC-DLBCL. However, patients with CD79wt/MyD88mut, or CARD11mut did not respond to ibrutinib, indicating the need for new therapies targeting ibrutinib refractory ABC-DLBCL. Recent approval of PI3Kδ selective inhibitor idelalisib for the treatment of indolent NHL (iNHL) as monotherapy highlighted selective inhibition of PI3Kδ as an effective therapeutic strategy. However, idelalisib did not show clinical activity in DLBCL in a Phase I expansion cohort study. Here we report immunohistochemical (IHC) analysis of clinical tumor samples from follicular lymphoma (FL) and DLBCL patients and a series of in vitro and in vivo mechanistic and functional studies to explore the importance of PI3K isoforms in regulating key signaling pathways in ABC-DLBCL.Methods: Expression of PI3K isoforms and PTEN was assessed by IHC and western blot from a panel of 45 FL and 45 DLBCL primary tumors. Effects of PI3K inhibitors (PI3Kδ-selective inhibitor idelalisib, PI3Kα-selective inhibitor BLY-719, PI3Kβ-selective inhibitor TGX-221, predominant PI3Kα/δ inhibitor copanlisib and BTK inhibitor ibrutinib) on nuclear NF-κB activation were determined using stable cell lines expressing NF-κB-luciferase reporter (obtained by lentiviral infection), IHC staining of p65 NF-κB, expression of CCL4, IL-6, and IL-10 by RT-PCR and protein production by ELISA assays. In vitro and in vivo mechanisms of action were addressed by assessing the activities of the key survival signaling pathways. In vitro and in vivo anti-tumor activities were investigated using cell lines and patient derived xenograft ABC-DLBCL models representing the key molecular features of BCR-dependent and independent ABC-DLBCL.Results: Although PI3Kδ was predominantly expressed in both FL and DLBCL, high PI3Kα expression was more prevalent in DLBCL (60% vs 18%), a patient population resistant to PI3Kδ-selective inhibition in the clinic. Simultaneous inhibition of PI3Kα and PI3Kδ with BYL-719 plus idelalisib or copanlisib alone dramatically enhanced anti-tumor profile in ABC-DLBCL models compared to selective inhibition of PI3Kδ, PI3Kα or BTK. The anti-tumor activity was associated with suppression of p-AKT and a novel mechanism of blocking NF-κB activation driven by CD79mut, CARD11mut, A20mut or MyD88mutin vitro and in vivo. Suppression of NF-kB activation by PI3K inhibition is independent from AKT, but involves a novel mechanism of modulating c-IAP expression. Inhibition of PI3Kα/δ resulted in complete tumor regression in an ibrutinib-resistant MyD88mut-LY0257 patient-derived (PDx) ABC-DLBCL model. Furthermore, rebound activation of BTK and AKT was identified as a mechanism limiting CD79mut ABC-DLBCL to show robust response to PI3K and BTK inhibitors, respectively, as single agents in vivo. Combination of ibrutinib with PI3Kα/δ inhibitor copanlisib dosed intermittently iv was well tolerated and produced complete tumor regression in CD79Bmut TMD-8 cell line and Ly2298 PDx ABC-DLBCL models.Conclusions: High expression of PI3Kα in addition to PI3Kδ in ABC-DLBCL is associated with intrinsic resistance to PI3Kδ selective inhibition. Simultaneous inhibition of PI3Kα/δ by copanlisib modulates not only the PI3K/AKT pathway but also BCR-dependent and independent activation of nuclear NF-κB via a novel AKT-independent mechanism, indicating a promising utility for the treatment of clinically relevant ibrutinib-resistant ABC-DLBCL patients with CD79wt/MyD88mut, A20mut, or CARD11mut tumor genotypes. Combination of PI3Kα/δ and BTK inhibitors demonstrated promising potential for ibrutinib-responsive ABC-DLBCL to achieve complete tumor regression by blocking rebound activation of BTK and AKT. Thus, our findings presented here provide additional insights on intrinsic and acquired resistance mechanisms to selective PI3Kδ and BTK inhibitors and provide rationale for clinical development of PI3K inhibitors with specific isoform profile in combination for the treatment of ABC-DLBCL. DisclosuresPaul:Bayer AG: Employment. Sojoun:Bayer AG: Employment. Wengner:Bayer AG: Employment. Zitzmann-Kolbe:Bayer AG: Employment. Sturz:Bayer AG: Employment. Haike:Bayer AG: Employment. Martin:Bayer AG: Employment. Mumberg:Bayer AG: Employment. Ziegelbauer:Bayer AG: Employment. Liu:Bayer AG: Employment.

Efficacy and Safety of the Bruton Tyrosine Kinase Inhibitor Ibrutinib in Patients with Hairy Cell Leukemia: Stage 1 Results of a Phase 2 Study

▪Background: Effective therapy for classical hairy cell leukemia (c-HCL) refractory to purine nucleoside analog (PNA) treatment is limited, and there are no accepted treatment standards for variant hairy cell leukemia (v-HCL). Ibrutinib, an oral small molecule inhibitor of Bruton tyrosine kinase (BTK), has shown single-agent efficacy and acceptable tolerability in patients with various B-cell malignancies, including chronic lymphocytic leukemia, mantle cell lymphoma, and Waldenstrom's. While BTK is expressed in HCL cells, the clinical activity of ibrutinib in this disease has not been previously assessed. We conducted a single-arm, multi-center phase 2 study (NCI #9268) of single-agent ibrutinib to characterize the overall response rate (complete + partial response) and safety of single-agent ibrutinib treatment for HCL.Methods: Patients with c-HCL (unfit for or relapsed after PNA) and v-HCL (relapsed or treatment-naïve) were eligible. Enrolled patients required treatment, had ECOG ≤ 2, no active infection, and preserved end-organ function. Patients received continuous once daily ibrutinib in 28-day cycles. Using a Simon 2-stage design, the first 13 patients were treated at 420 mg daily, and a second cohort of 13 patients was accrued at the 840 mg dose-level. Ultimately, the pre-specified number of objective responses was reached at the 420 mg dose-level, and accrual to the second stage of the design continues at that dose. Response, including bone marrow biopsy with immunohistochemistry for minimal residual disease (MRD), was assessed after 8 and 12 cycles. Patients experiencing clinical benefit may continue ibrutinib until unacceptable toxicity or progressive disease.Results: As of 15 May 2016, 28 patients had been enrolled and were evaluable for response: 420 mg/day (n=15) and 840 mg/day (n=13). Enrolled patients (1 treatment-naïve v-HCL, 10 relapsed v-HCL, 17 relapsed c-HCL) included 22 male and 6 female patients with median age 65 years (range: 43-78). Relapsed patients received median 4 prior therapies (range: 1-11); all had prior PNA, 81% had prior rituximab, and 3 c-HCL patients had prior vemurafenib. Response data are summarized by dose and histology in the table. To date, the ORR is 46% [4 CR (all c-HCL) and 9 PR (6 c-HCL)], with objective responses more commonly observed in patients with c-HCL. Eight additional patients (29%) with stable disease have experienced clinical benefit (resolution of symptoms, improvement in peripheral blood cell counts) not meeting criteria for PR, most commonly attributable to persistent thrombocytopenia, and continue on treatment. At median follow-up of 22 months, 20 patients (71%) remain on treatment, 3 patients (1 v-HCL, 2 c-HCL) have progressed, 2 patients (c-HCL) with severe neutropenia at baseline (c-HCL) had early deaths from pneumonia, 1 (c-HCL) discontinued during cycle 8 for failure to resolve baseline neutropenia, and 2 other patients (v-HCL) have discontinued for adverse events. Estimated 24-month progression-free survival (PFS) was 79% (95% CI: 57-91%) and the median PFS has not been reached. Redistribution lymphocytosis was observed in patients with circulating disease at baseline. Soluble interleukin-2 receptor (sIL2R) level was increased at baseline in all c-HCL patients for whom data were available and correlated with response. In general, toxicity did not differ by dose level or disease histology. The most frequent (>10%) grade ≥3 adverse events (AEs) were lymphopenia (21%), neutropenia (18%), lung infection (18%), thrombocytopenia (14%), hypophosphatemia (11%), and hypertension (11%). Common grade 1/2 non-hematologic toxicities included myalgia (57%), diarrhea (54%), fatigue (50%), nausea/vomiting (46%), bruising (46%), and rash (46%); grade 1/2 atrial fibrillation (no grade ≥3) was observed in 5 patients. Grade ≥3 infection was observed in 25%, but no grade ≥3 bleeding AEs were reported.Conclusions: Ibrutinib can induce remission in both c-HCL and v-HCL patients, including heavily pre-treated patients, but complete remissions have only been observed in c-HCL to date. The drug is generally well-tolerated during long-term administration, and durable clinical benefit is observed in the majority of treated patients even when objective response criteria are not met. [Display omitted] [Display omitted] DisclosuresJones:Pharmacyclics, LLC, an AbbVie Company: Membership on an entity's Board of Directors or advisory committees, Research Funding; AbbVie: Membership on an entity's Board of Directors or advisory committees, Research Funding; Janssen: Membership on an entity's Board of Directors or advisory committees, Research Funding. Lozanski:Stemline Therapeutics Inc.: Research Funding; Beckman Coulter: Research Funding; Genentech: Research Funding; Boehringer Ingelheim: Research Funding.

The Bruton's Tyrosine Kinase (BTK) Inhibitor ARQ 531 Effectively Inhibits Wild Type and C481S Mutant BTK and Is Superior to Ibrutinib in a Mouse Model of Chronic Lymphocytic Leukemia

Introduction: The Bruton's tyrosine kinase inhibitor ibrutinib improves survival in chronic lymphocytic leukemia (CLL) compared to standard chemotherapy or immune therapy. In a subset of patients, somatic mutation (C481S) of its binding site results in acquired resistance to ibrutinib therapy with poor clinical outcome. ARQ 531 is an ATP competitive, orally bioavailable, potent inhibitor of BTK and other relevant kinases including the majority of Src and Tec family kinases. Herein we present preclinical data with ARQ 531 in CLL including C481S mutated BTK and its efficacy versus ibrutinib in the TCL1 mouse model of CLL.Methods: Potency of ARQ 531 and its binding kinetics were measured in enzymatic and Surface Plasmon Resonance (SPR) binding assays. Primary CLL B cells were negatively selected and treated with 1μM ARQ 531. BCR signaling was investigated by immunoblot following a 1 hour drug incubation. CLL cells migrating towards CXCL12 and CXCL13 after 4 hours across a 5.0 micron transwell insert were counted by flow cytometry. Annexin V and propidium iodide flow cytometry was used to measure CLL viability over a range of drug concentrations and time. CpG mediated CLL cell activation was measured by CD40 and CD86 expression by flow cytometry. In vivo investigation utilized B6 mice engrafted with 1E7 CD5+/CD19+ TCL1 lymphocytes via tail vein injection. Mice were randomized to treatment with vehicle, ARQ 531, or ibrutinib following the establishment of a CD5+/CD19+ population >10% in peripheral blood.Results: At 72 hours the viability of CLL cells treated with 0.1µM, 1.0µM, or 10.0µM ARQ 531 was found to be 94%, 67%, and 50% that of untreated samples, respectively (p=0.76, p<0.001, p=0.016) in comparison to the 71% relative viability seen with 1µM ibrutinib (p<0.001). ARQ 531 demonstrates dose dependent inhibition of BTK, AKT, and ERK phosphorylation. Stimulation through TLR9 by CpG ligand increases expression of the activation markers CD40 and CD86; ARQ 531 decreased CpG induced expression of CD40 by 25% and CD86 by 40% (p=0.022, p=0.041). ARQ 531 in the presence of CXCL12 or CXCL13 decreased migration by 51% and 66%, respectively (p=0.015, p=0.025), below baseline migration of untreated CLL.Unlike ibrutinib, ARQ 531 inhibits both wild type and C481S BTK with an IC50 less than 1nM in a biochemical assay, and inhibits phosphorylation of C481S mutated BTK in LV125 transfected HEK293T cells. Cytotoxicity was also observed in CLL cells isolated from patients with C481S BTK mutations, where the viability of cells treated with 1µM ARQ 531 was 72% that of untreated cells at 72 hours (p=0.038). SPR binding assay showed a residence time of 51 minutes in wild type BTK and 56 minutes in C481S mutated BTK.The TCL1 mouse model displays active BCR signaling and responds to treatment with the BTK inhibitors ibrutinib and acalabrutinib (Ponader S, 2012; Herman SEM, 2015). Following establishment of leukemia in the TCL1 transfer model, we randomized mice to treatment with vehicle (n=14), 25 mg/kg ibrutinib (n=6), 50 mg/kg ARQ 531 (n=14) or 75 mg/kg ARQ 531 (n=14) given by daily gavage. ARQ 531 significantly improved survival over both ibrutinib and vehicle (median survival: 36 days vehicle, 53 days ibrutinib, not reached by 74 days 50mg/kg and 75 mg/kg ARQ 531, p<0.0001). Mice receiving ARQ 531 have consistently lower lymphocyte counts on microscopic examination and reduced CD5+/CD19+ populations via flow cytometry compared to vehicle or ibrutinib. Additionally, we have observed increases in absolute neutrophil count over time in mice receiving ARQ 531. Pharmacokinetic studies are pending; however, in a single oral dose study of 10mg/kg in monkeys, the bioavailability was 72.4% with a CMax of 9µM and a half-life greater than 24 hours.Conclusion: The BTK inhibitor ARQ 531 is a potent inhibitor of BTK with promising activity both in vitro and in vivo. Multi-targeted inhibition of cytokine, chemokine, and BCR pathways by ARQ 531 decreases activation, migration, and viability of CLL cells. Unlike ibrutinib, ARQ 531 inhibits activation of C481S mutated BTK variants and maintains cytotoxicity in ibrutinib resistant clones. Additionally, ARQ 531 has demonstrated remarkable efficacy in an in vivo TCL1 adoptive transfer model, improving survival to a greater extent than ibrutinib and potentially restoring granulocyte production. These data justify continued preclinical work and development to facilitate transition to clinical trials. DisclosuresEathiraj:ArQule, Inc: Employment. Abbadessa:ArQule, Inc: Employment. Schwartz:ArQule, Inc: Employment. Woyach:Acerta: Research Funding; Morphosys: Research Funding; Karyopharm: Research Funding.

Ibrutinib Disrupts the Metabolic Program of CLL Cells in-Vivo

Introduction: Unlike their normal resting B cell counterparts, chronic lymphocytic leukemia (CLL) cells proliferate. Approximately 1% of the total CLL cell clone expands daily. To adjust for the increase in energetic demands imposed by continuous proliferation, CLL cells undergo metabolic reprogramming and, as recently shown (Rozovski U, et al. Mol Cancer Res. 2015; 13:944-53), CLL cells utilize fat in a manner similar to that of adipocytes. The recent introduction of the oral Bruton tyrosine kinase inhibitor (BTK) ibrutinib revolutionized the treatment of CLL. Because the proliferation of CLL cells is driven by lipid metabolism and ibrutinib inhibits the B cell receptor-induced proliferation of CLL cells, we sought to determine whether ibrutinib also disrupts the metabolic program that provides CLL cells with their unique energy requirements.Methods: We prospectively studied serial peripheral blood samples from 16 patients with CLL. The patients’ peripheral blood CLL cells were analyzed prior to and during treatment with ibrutinib. All patients received a daily dose of 420 mg ibrutinib. In addition, we performed in vitro studies using CLL cells from 3 ibrutinib-naïve patients. CLL cells were analyzed for free-fatty acids (FFA) consumption and for the rate of cellular apoptosis using propiduim iodide (PI) and annexin V staining analyzed by flow cytometry.Results: To study lipid metabolism of CLL cells we incubated peripheral blood CLL cells from 3 randomly selected ibrutinib-naïve patients in the presence or absence of FFA and measured the concentration of culture media-dissolved O2 (dO2). Like in our previous study (Rozovski U, et al. Mol Cancer Res. 2015; 13:944-53), we found that CLL cells metabolized FFA and, as a result, the levels of dO2 decreased. However when the cells were co-cultured with FFA and ibrutinib, the delta dO2 (dO2 with FFA minus dO2 without FFA) remained unchanged, suggesting that ibrutinib blocked FFA metabolism in CLL cells.Then, to determine whether ibrutinib also inhibited CLL-cell lipid metabolism in patients treated with ibrutinib, we collected 2 to 5 consecutive PB samples (median: 5) from 16 CLL patients prior to and during treatment with ibrutinib. Unlike the 12% reduction in delta dO2 detected in untreated patients’ CLL cells incubated with FFA in vitro, a 6% reduction in delta dO2 was detected in CLL cells of patients treated with ibrutinib 4 days into treatment and after a median of 147 days of ibrutinib treatment a change in delta dO2 was no longer detected. These data suggest that ibrutinib-treated cells lost their capacity to utilize FFA or that the number of FFA consuming circulating CLL cells declined until they were no longer detected. In addition, whereas ibrutinib induced apoptosis of CLL cells in a dose-dependent manner in vitro, ibrutinib did not induce apoptosis at the same time points in vivo, suggesting that interruption of FFA metabolism does not lead to apoptotic cell death and that the metabolic and proapoptotic pathways are not linearly intertwined in CLL cells.In conclusion: Treatment with ibrutinib changes the metabolic profile CLL cells. Even after short exposure to the drug the cells were less capable of utilizing FFA, and after longer exposure, the cells could no longer utilize FFA. Whether ibrutinib induced reduction in FFA metabolism decreases the proliferation capacity of CLL cells remains to be determined. DisclosuresBurger:Pharmacyclics: Research Funding. O'Brien:Janssen: Consultancy, Honoraria; Pharmacyclics, LLC, an AbbVie Company: Consultancy, Honoraria, Research Funding. Jain:Pfizer: Consultancy, Honoraria, Research Funding; Celgene: Research Funding; Abbvie: Research Funding; Novimmune: Consultancy, Honoraria; Servier: Consultancy, Honoraria; Incyte: Research Funding; ADC Therapeutics: Consultancy, Honoraria, Research Funding; Seattle Genetics: Research Funding; Genentech: Research Funding; BMS: Research Funding; Pharmacyclics: Consultancy, Honoraria, Research Funding; Infinity: Research Funding; Novartis: Consultancy, Honoraria. Wierda:Novartis: Research Funding; Abbvie: Research Funding; Acerta: Research Funding; Gilead: Research Funding; Genentech: Research Funding.

Ibrutinib Abrogates TREM-1 Mediated Neutrophil Activation

Triggering receptor expressed on myeloid cells 1 (TREM-1) is an activating receptor on neutrophils (PMN) and important in the innate host defence against microbial pathogens. Here we examined the influence of the Bruton tyrosine kinase (BTK) inhibitor ibrutinib on TREM-1 dependent activation of human PMNs. Firstly, ibrutinib specifically inhibited TREM-1 mediated PMN activation of the oxidative burst and CD62L shedding, whereas TLR mediated activation remained unaffected. Correspondingly, ibrutinib suppressed ERK phosphorylation after TREM-1, but not after TLR ligation. To clarify whether this TREM-1 specific effect of ibrutinib was also relevant in vivo, we treated mice with ibrutinib, and analyzed fungal clearance in vivo upon pulmonary challenge with A. fumigatus and PMN activation ex vivo upon TREM-1 or TLR ligation, again finding that TREM-1, but not TLR induced activation was suppressed. Equivalent results were obtained in PMNs of six lymphoma patients under treatment with ibrutinib. In summary, BTK plays an important role in TREM-1 mediated PMN activation, providing a potential mechanism for the frequently observed infectious complications in patients treated with ibrutinib, and fosters studies using this drug as a novel tool to modulate TREM-1 activity for therapeutic purposes, i. e. in sepsis or autoimmune diseases. DisclosuresHess:Pfizer: Honoraria; Janssen: Honoraria; Celgene: Honoraria; Roche, CTI, Pfizer, Celgene: Research Funding; Roche: Honoraria; Novartis: Honoraria.

Targeting MALT1 with the Small Molecule Inhibitor MI2 Induces a Caspase-Dependent Apoptosis and Inhibits the NF-κb Pathway in Chronic Lymphocytic Leukemia Primary Cells

Targeting the B-cell receptor (BCR) pathway in chronic lymphocytic leukemia (CLL) with the Bruton Tyrosine Kinase (BTK) inhibitor Ibrutinib has transformed the treatment paradigm of this disease. Ibrutinib is currently indicated for treatment of CLL regardless of the line of therapy. However, ibrutinib is not curative, and relapses secondary to C481S mutation in BTK and gain of function mutations in PLCγ2 (R665W and L845F) are being reported.The protease mucosa-associated lymphoid tissue lymphoma translocation 1 (MALT1) is the active component of the CARD11-BCL10-MALT1 (CBM) signaling complex. CBM mediates NF-κB activation downstream of BTK and PLCγ2 within the BCR pathway, which makes of MALT1 an ideal therapeutic target in ibrutinib-resistant clones. Notably, the chemical compound MI2 (C19H17Cl3N4O3) binds to- and suppresses the protease activity of MALT1, and was found to be active against activated B-cell-like diffuse large B-cell lymphoma. However, the role of MALT1 inhibition in CLL has not been investigated.We studied the efficacy of MI2 against a cohort of CLL samples (N=21), and explored its mechanism of action. PBMCs from patients with CLL were tested against serial dilutions of MI2 (0.125-8μM) for 48 h in 96-well plates, and MTS assay was performed to quantify cell viability. We observed a dose-dependent cytotoxicity in all samples, with an inhibitory concentration at 25% (IC25) < 2μM observed in the majority of the samples. MI2 cytotoxicity was independent of the IGHV mutational status, CD38 expression, cytogenetics, previous therapy status, and MALT1 protein level (immunoblot on purified CLL cells). To study the drug apoptosis and drug selectivity, we examined apoptotic induction in both CLL cells and T-cells of the same patients. PBMCs from five patients with CLL were treated with MI2 for 48 h and compared to untreated control. At the end of the incubation, cells were stained with CD5, CD19, and Annexin-V, then analyzed using flow cytometry. We found that MI2 induced a dose-dependent apoptosis in CLL cells (defined as CD5+/CD19+) in all samples, with only mild toxicity to their corresponding T-cells (T-cells represent a fraction of each patient’s PBMCs defined as CD5+/CD19-). This minimal toxicity to T-cells was not statistically significant when compared to untreated control T-cells, and may reflect a T-cell receptor signaling-independence of circulating T-cells, which are mostly composed of terminally differentiated effectors. To determine whether the induced apoptosis is caspase-dependent, we treated PBMCs of CLL patients (N=5) with MI2 (0.5-4 μM) in the presence or absence of 100μM of the pan-caspase inhibitor z-VAD-fmk for 48 h. We found that z-VAD-fmk completely prevented the induction of apoptosis in CD19+/CD5+ CLL cells as determined by Annexin-V staining (flow cytometry), and decreased MI2-induced PARP cleavage (Immunoblot).To determine the effect of the microenvironment on MI2 toxicity, we cultured primary CLL cells with and without MI2 (0.5-4µM) for 48 h, in the presence or absence of Nurse-Like Cells (NLC) (N=5) or anti-IgM (N=4). NLCs and anti-IgM cross-linking of BCR are in-vitromodels for the protective effect of the microenvironment. Co-culture with NLCs or with anti-IgM improved the viability of the untreated primary CLL samples. Similar to ibrutinib, MI2 continues to be effective against CLL cells in the presence of NLCs or anti-IgM, but higher concentrations are required to achieve the desired cytotoxicity against CLL cells. There was no cytotoxic effect on NLCs.To investigate the effect of MI2 on tumor biology, 1 μg of total RNA from purified CLL cells (treated for 8 h with MI2, N=3; and untreated control, N=3) was subjected to RNA sequencing. Out of 56,650 tested genes, there were 438 genes whose expression changed ≥2-fold at P<0.05 (312 down- and 126 up-regulated), several genes are known NF-κB targets. The observer-independent method, Gene Set Enrichment Analysis (GSEA), clearly identified the canonical NF-κB among of the top affected pathways.Further studies aim to verify the RNA sequencing data at the protein level, to determine MI2 efficacy against ibrutinib-resistant clones in vitro, and against CLL growth in mice in vivo.Supported by Hope Foundation SWOG Early Exploration and Development (SEED) Award. We thank the Louisiana Cancer Research Center (LCRC) biospecimen core and our patients for the donation of samples to make this research possible. DisclosuresNo relevant conflicts of interest to declare.

Natural History of Non-Infectious, Ibrutinib-Attributable Adverse Events Leading to Alternative BTK Inhibitor Use in CLL

Introduction:The Bruton tyrosine kinase (BTK) inhibitor ibrutinib (IB) is currently approved for the treatment of patients with chronic lymphocytic leukemia and small lymphocytic lymphoma (CLL/SLL), including patients with del17p disease. Treatment with IB is effective and generally well-tolerated. However, a proportion of patients experience intolerable adverse events (AEs) requiring IB discontinuation. Non-infectious toxicities are generally considered to be due to kinase inhibition and it is uncertain if the AE is directly attributable to inhibition of BTK (on-target effect) or to inhibition of an alternative target of ibrutinib (off-target effect). To indirectly examine this, we reviewed toxicity and tolerability data for patients who discontinued IB due to significant AEs and subsequently received an alternative BTK inhibitor.Methods:Patients with previously treated CLL/SLL who were intolerant to IB due to non-infectious IB-attributable AEs and subsequently received a different BTK inhibitor were included in this analysis. Patients with infectious AEs were excluded. AEs arising during IB therapy that were attributable to IB and resulted in discontinuation based on both patient and investigator preference were detailed and graded according to CTCAE v.4.03. These AEs were assessed for resolution or increasing severity after discontinuing IB both before and during subsequent BTK inhibitor treatment.Results:Twenty-one patients were included in the analysis and the median age was 61-years (range 50-76). Patients had received a median of 4 prior therapies (range 2-11). Fifty-eight percent of the patients had Rai stage 3-4 disease, 76% were IGHV un-mutated, and 57% had del17p. Prevalent co-morbidities by organ system prior to start of acalabrutinib included blood and lymphatic system disorders (48%), infectious issues (52%), cardiac disorders (38%), gastrointestinal disorders (43%) and musculoskeletal disorders (33%). Median time on IB was 13 months (range 1.6-62). Reasons for IB discontinuation are detailed in Table 1. With regards to more severe (grade 3 or 4) IB-attributable AEs, 2 patients experienced grade 4 AEs resulting in discontinuation of IB including neutropenia and subdural hematoma. Both these patients had resolution of their AEs prior to starting subsequent treatment. Among the 14 patients discontinuing IB for grade 3 AEs, 1 had ongoing grade 3 diarrhea, 1 had grade 3 arthralgia, and 2 had grade 2 AEs of diarrhea and fatigue at the time of starting subsequent treatment. Of the 21 included patients, 10 had ongoing IB-attributable grade 1-3 AEs at the time of starting a different BTK inhibitor. Three of these patients experienced resolution of one or more IB-attributable AEs (grade 3 diarrhea, and grade 2 AEs including panniculitis, arthralgia and diarrhea) within 28 days of starting therapy, 6 patients had stable grade 1-2 AEs (grade 1 rash in 2 patients, grade 2 diarrhea, grade 1 arthralgia, grade 1 abdominal pain, grade 2 fatigue) at last follow up while taking alternate BTK inhibitor, and 1 patient with an ongoing grade 2 AE at time of treatment initiation developed CNS involvement with their malignancy and was taken off therapy prior to assessment for resolution of the AE. The remaining 11 patients had either complete resolution or improvement of the AE to grade 1 prior to starting the treatment. No recurrence or worsening of IB-attributable AEs were observed during a median follow-up of 13-months on alternative treatment except for 1 patient with hemophilia A, who developed grade 3 hematoma while on IB therapy and experienced recurrent hematoma after 5 days of treatment with a different BTK inhibitor, at which time the drug was discontinued.Conclusions:For patients who discontinued IB due to non-infectious IB-attributable AEs, subsequent treatment with an alternative BTK inhibitor does not appear to aggravate or precipitate IB-attributable AEs that had previously resulted in discontinuation. This supports attribution of these AEs to off-target effects of ibrutinib and not inhibition of BTK. The use of an alternate BTK inhibitor may be a viable therapeutic strategy for patients who discontinued IB due to intolerable AEs without experiencing clinical progression. [Display omitted] DisclosuresJones:Janssen: Membership on an entity's Board of Directors or advisory committees, Research Funding; Pharmacyclics, LLC, an AbbVie Company: Membership on an entity's Board of Directors or advisory committees, Research Funding; AbbVie: Membership on an entity's Board of Directors or advisory committees, Research Funding. Woyach:Karyopharm: Research Funding; Morphosys: Research Funding; Acerta: Research Funding. Awan:Innate Pharma: Research Funding; Novartis Oncology: Consultancy; Pharmacyclics: Consultancy.

Opposite Effects of Bruton Tyrosine Kinase (BTK) Inhibitor Therapy with Ibrutinib and FCR Chemo-Immunotherapy on the Normal B Lymphocyte Repertoire in Patients with Chronic Lymphocytic Leukemia

Disease-inherent and treatment-related immune dysfunction remain leading causes for morbidity and mortality in patients with chronic lymphocytic leukemia (CLL). The advent of kinase inhibitors that target B cell receptor (BCR) signaling, which lack myelo- and T lymphocyte toxicity, raised hopes that these new agents may be less immunosuppressive and allow for better immune reconstitution when compared to chemo-immunotherapy (CIT). The effects of the BTK inhibitor ibrutinib or CIT with fludarabine, cyclophosphamide and rituximab (FCR) on the normal B cell repertoire have not been well characterized. Here, we used state-of-the-art immunosequencing technology to investigate how ibrutinib treatment affects the regeneration of non-malignant B-cells when compared to patients treated with FCR.Clinical data on infection rates and immunoglobulin levels was analyzed from 40 CLL patients treated with ibrutinib (median number of two pre-treatments) or frontline CIT with FCR at MD Anderson Cancer Center. In a representative subset of 20 patients, flow cytometry and next generation sequencing (NGS) of the immunoglobulin heavy chain (IGH) gene locus was used to monitor non-malignant B-cell immune reconstitution for 24 months after start of treatment with ibrutinib or FCR.Comparison of ibrutinib treatment with CIT revealed that immunoglobulin levels remained stable and relatively low in both cohorts, except for an increase in IgA during ibrutinib treatment, as previously reported. NGS results showed that ibrutinib treatment significantly decreased the non-malignant B-cells count after 24 months of treatment, while the counts were quantitatively stable in the FCR cohort. Next, we determined the dynamics of non-malignant B-cell immune repertoire composition over treatment. Based on the mutational status of the V gene, non-malignant B-cells were classified as IGH hypermutated (<98% identity to the corresponding germline V gene, corresponding to antigen-experienced B-cells) or IGH unmutated (≥98% identity to the corresponding germline V gene, corresponding to antigen-naïve B-cells). Before treatment initiation, the mean percentage of antigen-experienced B-cells did not significantly differ between the groups (ibrutinib 39%, FCR 48%). After 24 months, a significant decrease of antigen-experienced B-cells was observed in the FCR cohort, while the ratio of antigen-experienced and antigen-naïve B-cells remained unchanged in ibrutinib treated patients (ibrutinib 39%, FCR 22%, p=0.01). Analysis of the IGH clonotype repertoire using the Shannon-Wiener and the inverse Simpson diversity indices confirmed these results, showing that the non-malignant IGH repertoire was composed of balanced numbers of antigen-experienced and antigen-naïve medium sized clones before treatment initiation in both cohorts. In line with the IGH repertoire shift towards antigen-naïve B-cells in FCR treated patients, the medium-sized clones disappeared after treatment, with large numbers of small-sized unmutated clones dominating after 24 months (p<0.0001). In ibrutinib treated patients, the repertoire diversity remained stable throughout the course of treatment.Taken together, our data indicate that continuous treatment with ibrutinib preserves preexisting (partially antigen-experienced) B-cells but impairs de-novo generation of naive B-cells. In contrast, FCR leads to a deletion of memory B-cells but also a subsequent substantial renewal of the B-cell repertoire. Both patterns may differentially affect immune-competence towards infections. DisclosuresBokemeyer:Karyopharm: Research Funding. Jain:Pfizer: Consultancy, Honoraria, Research Funding; Incyte: Research Funding; Genentech: Research Funding; Abbvie: Research Funding; Pharmacyclics: Consultancy, Honoraria, Research Funding; Infinity: Research Funding; Novartis: Consultancy, Honoraria; Servier: Consultancy, Honoraria; Novimmune: Consultancy, Honoraria; ADC Therapeutics: Consultancy, Honoraria, Research Funding; BMS: Research Funding; Celgene: Research Funding; Seattle Genetics: Research Funding. Wierda:Gilead: Research Funding; Abbvie: Research Funding; Novartis: Research Funding; Acerta: Research Funding; Genentech: Research Funding. Burger:Pharmacyclics: Research Funding.

Novel Bruton's Tyrosine Kinase Inhibitor Bgb-3111 Demonstrates Potent Activity in Mantle Cell Lymphoma

Background:Aberrant B-cell receptor signaling is an important contributor to lymphomagenesis in mantle cell lymphoma (MCL). Bruton's Tyrosine Kinase (BTK), a component of the BCR signaling axis, has been validated as a clinically relevant target, and BTK inhibitor ibrutinib received FDA approval for treatment of MCL in 2013. Growing concerns that single agent ibrutinib exerts off-target effects that interfere with other treatments such as rituximab-induced antibody-dependent cell cytotoxicity limit its utility in combination treatments. In this study, we assessed the in vitro and in vivo effects of BGB-3111in MCL models.Methods:We performed cell viability assays with BGB-3111 treated MCL cell lines to determine inhibition of cellular proliferation. The same assays were conducted on primary human MCL cells and patient-derived xenograft (PDX) tumor samples. Dose-dependent inhibition of BTK auto-phosphorylation and inhibition of downstream targets such as PLC-γ were determined by phospho-protein immunoblotting and immunoprecipitation. A reverse-phase protein assay (RPPA) was conducted on BGB-3111-treated Mino cells to evaluate changes in MCL oncogenic signaling. Induction of apoptosis in MCL cells treated with increasing doses of BGB-3111 was quantified using flow cytometry. For in vivo experiments, an ibrutinib-sensitive MCL PDX mouse model was treated with 50 mg/kg/day BGB-3111 and monitored for mean tumor burden and survival.Results:BGB-3111 potently inhibited cell viability in a panel of MCL cell lines, with an activity range of 1-10 uM, and induced apoptosis in a dose-dependent manner in several MCL cell lines.BGB-3111 treatment of MCL cells demonstrated a dose-dependent decrease in p-BTK (Y223) and inhibition of downstream effectors without impacting total protein levels, while RPPA revealed upregulation of the PI3K-Akt signaling axes. In addition, BGB-3111 treatment did not impact phosphorylation of off-target kinases affected by ibrutinib treatment. In vivo, BGB-3111 suppressed tumor growth and prolonged tumor survival in BGB-3111 treated mice.Conclusion:The second generation BTK inhibitor BGB-3111 demonstrates selectivity for BTK in vitro and BTK inhibition in vivo. BGB-3111-treated PDX mouse models examining survival, tumor growth, and other factors point to BGB-3111 as an effective single agent BGB-3111 is being investigated in Phase I clinical trials. DisclosuresWang:Beigene: Employment. Wang:Asana BioSciences: Research Funding; Janssen: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Dava Oncology: Honoraria; Acerta: Consultancy, Research Funding; Kite Pharma: Research Funding; BeiGene: Research Funding; Asana biosciences, Beigene, Celgene, Juno, Kite, Onyx, Pharmacyclics: Research Funding; Juno Therapeutics: Research Funding.

HSP90 inhibition overcomes ibrutinib resistance in mantle cell lymphoma

AbstractThe Bruton tyrosine kinase (BTK) inhibitor ibrutinib induces responses in 70% of patients with relapsed and refractory mantle cell lymphoma (MCL). Intrinsic resistance can occur through activation of the nonclassical NF-κB pathway and acquired resistance may involve the BTK C481S mutation. Outcomes after ibrutinib failure are dismal, indicating an unmet medical need. We reasoned that newer heat shock protein 90 (HSP90) inhibitors could overcome ibrutinib resistance by targeting multiple oncogenic pathways in MCL. HSP90 inhibition induced the complete degradation of both BTK and IκB kinase α in MCL lines and CD40-dependent B cells, with downstream loss of MAPK and nonclassical NF-κB signaling. A proteome-wide analysis in MCL lines and an MCL patient-derived xenograft identified a restricted set of targets from HSP90 inhibition that were enriched for factors involved in B-cell receptor and JAK/STAT signaling, the nonclassical NF-κB pathway, cell-cycle regulation, and DNA repair. Finally, multiple HSP90 inhibitors potently killed MCL lines in vitro, and the clinical agent AUY922 was active in vivo against both patient-derived and cell-line xenografts. Together, these findings define the HSP90-dependent proteome in MCL. Considering the disappointing clinical activity of HSP90 inhibitors in other contexts, trials in patients with MCL will be essential for defining the efficacy of and mechanisms of resistance after ibrutinib failure.

Therapeutic Approach to Patients with Chronic Lymphocytic Leukemia and Significant Comorbid Conditions.

Clinical trials in chronic lymphocytic leukemia (CLL) have focused mainly on younger fit patients until recently. However, CLL is a disease of elderly and many patients have significant comorbid conditions which together with advanced age preclude the use of aggressive regimens like FCR (fludarabine, cyclophosphamide, rituximab). Therefore, parameters such as performance status, renal function and number/severity of comorbidities together with clinical judgment should be used to guide the decision-making process regarding intensity of treatment. Two large randomized trials recently demonstrated that addition of monoclonal anti-CD20 antibodies (obinutuzumab, rituximab, and ofatumumab) to chlorambucil in untreated comorbid patients lead to improvement in complete remission rate, progression-free survival and even overall survival (obinutuzumab-chlorambucil and rituximab-chlorambucil), with acceptable toxicity profile. Thus, chemoimmunotherapy combining chlorambucil with an anti-CD20 antibody is the new standard approach for elderly/comorbid CLL patients in the first line. Treatment of relapsed/refractory disease in this patient population is very challenging and data regarding this subpopulation are rather limited. Impressive efficacy of novel targeted small molecules interfering with B-cell receptor signaling, namely Bruton tyrosine kinase inhibitor ibrutinib and phosphatidylinositol-3 kinase delta inhibitor idelalisib, radically changed the treatment paradigms for relapsed/refractory CLL; relatively mild toxicity of these agents make them very good candidates for elderly/comorbid patients. Other options for relapsed/refractory disease include alemtuzumab, ofatumumab, high-dose glucocorticoids+rituximab and bendamustine+rituximab. This review summarizes the current knowledge on prognostication and therapy of elderly and comorbid patients with CLL.

Analysis of Efficacy and Tolerability of Bruton Tyrosine Kinase Inhibitor Ibrutinib in Various B-cell Malignancies in General Community: a Single-Center Experience

Analysis of Efficacy and Tolerability of Bruton Tyrosine Kinase Inhibitor Ibrutinib in Various B-cell Malignancies in General Community: a Single-Center Experience

Abstract 4860: The BTK inhibitor ibrutinib modulates T cell immunity in mouse models and in differentiated human T cells

Abstract Ibrutinib inhibits Bruton's tyrosine kinase (BTK) and effectively treats B-cell malignancies. Ibrutinib also targets IL-2-inducible T-cell kinase (ITK) and modulates the T-cell receptor signaling pathway. Because ibrutinib enhanced antitumor immunity when combined with anti-PD-L1 in various syngeneic solid tumor and lymphoma models (Sajiv-Barfi, PNAS 2015), we sought to determine the effect of ibrutinib on T-cell function and activation. Using in vitro human primary T-cell activation assays, we found that ibrutinib did not suppress T-cell proliferation but slightly inhibited IFNã production in CD4 and CD8 effector T cells when human peripheral blood mononuclear cells (PBMCs) were stimulated with anti-CD3 and anti-CD28 antibodies. Ibrutinib also inhibited PD-1 expression on human CD8 effector T cells. More strikingly, ibrutinib significantly suppressed naïve murine CD4 T-cell differentiation in vitro into Foxp3+ regulatory T cells (Treg) in the presence of TGF-â. Further, we evaluated the immunomodulatory effect of ibrutinib in tumor-bearing mice. In pancreatic (Pan02) and renal cell carcinoma (Renca) models, we found that single-agent ibrutinib, as well as in combination with standard-of-care agents, effectively inhibited tumor growth. In the syngeneic Renca tumor model, combination ibrutinib treatment significantly reduced CD4+Foxp3+ Treg cells both in the spleen (P &amp;lt; 0.001) and tumor microenvironment (P &amp;lt; 0.05), compared with everolimus alone. Ibrutinib also significantly reduced the percentage of PD-1hi cells among CD8 tumor-infiltrating lymphocytes in A20 lymphoma-bearing mice (P &amp;lt; 0.05). These data demonstrate that ibrutinib, a BTK inhibitor, can suppress immune-inhibitory factors and enhance antitumor immunity in vivo. Ibrutinib may serve as a promising immunomodulatory agent in potentiating current cancer immunotherapy in solid tumors. Citation Format: Yujun Huang, Jeff Hsu, Mint Sirisawad, Hsu-Ping Kuo, Betty Y. Chang. The BTK inhibitor ibrutinib modulates T cell immunity in mouse models and in differentiated human T cells. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 4860.

Pathogenic role of B-cell receptor signaling and canonical NF-κB activation in mantle cell lymphoma

AbstractTo interrogate signaling pathways activated in mantle cell lymphoma (MCL) in vivo, we contrasted gene expression profiles of 55 tumor samples isolated from blood and lymph nodes from 43 previously untreated patients with active disease. In addition to lymph nodes, MCL often involves blood, bone marrow, and spleen and is incurable for most patients. Recently, the Bruton tyrosine kinase (BTK) inhibitor ibrutinib demonstrated important clinical activity in MCL. However, the role of specific signaling pathways in the lymphomagenesis of MCL and the biologic basis for ibrutinib sensitivity of these tumors are unknown. Here, we demonstrate activation of B-cell receptor (BCR) and canonical NF-κB signaling specifically in MCL cells in the lymph node. Quantification of BCR signaling strength, reflected in the expression of BCR regulated genes, identified a subset of patients with inferior survival after cytotoxic therapy. Tumor proliferation was highest in the lymph node and correlated with the degree of BCR activation. A subset of leukemic tumors showed active BCR and NF-κB signaling apparently independent of microenvironmental support. In one of these samples, we identified a novel somatic mutation in RELA (E39Q). This sample was resistant to ibrutinib-mediated inhibition of NF-κB and apoptosis. In addition, we identified germ line variants in genes encoding regulators of the BCR and NF-κB pathway previously implicated in lymphomagenesis. In conclusion, BCR signaling, activated in the lymph node microenvironment in vivo, appears to promote tumor proliferation and survival and may explain the sensitivity of this lymphoma to BTK inhibitors.

A randomized, double-blind, placebo-controlled study of ibrutinib, a Bruton tyrosine kinase inhibitor, with nab-paclitaxel and gemcitabine in the first-line treatment of patients with metastatic pancreatic adenocarcinoma (RESOLVE).

TPS2601Background: Advanced pancreatic ductal adenocarcinoma (PDAC) is an aggressive malignancy with poor prognosis and short survival. Nab-paclitaxel/gemcitabine (Nab-P/GCB) is a preferred front-line treatment regimen. The Bruton tyrosine kinase (BTK) inhibitor ibrutinib (ibr) demonstrates antitumor activity in preclinical PDAC models, due in part to modulation of the tumor microenvironment. Ibr treatment inhibits mast-cell degranulation and decreases tumor-associated inflammation and desmoplasia; simultaneously, ibrutinib enhances the presence and activity of cytotoxic T cells in tumors (Affara et al, Cancer Cell, 2014; Masso-Valles et al, Cancer Res, 2015; Gunderson et al, Cancer Discov, 2015). This study will evaluate the efficacy of ibr in combination with Nab-P/GCB. Methods: The randomized, multicenter, double-blind, phase 2/3 RESOLVE trial (NCT02436668; PCYC-1137-CA) compares ibr vs. placebo, in combination with Nab-P/GCB, in approximately 320 patients with previously untreated metastatic pancreati...