Covalent BTK Research Articles

Pacritinib Blocks Key Pro-Survival Signaling Related to Mutated MYD88, Produces High Levels of Apoptosis and Overcomes Mutated BTK Cys481 Related BTK-Inhibitor Resistance

Background: Somatic mutations in MYD88 are prevalent in many B-cell malignancies, including Waldenström Macroglobulinemia (WM; 95-97%), primary CNS lymphoma (70-80%), ABC DLBCL (40%), marginal zone lymphoma (5-10%) and CLL (5-15%). Mutated MYD88 drives oncogenic pro-survival signaling through multiple signaling cascades that include HCK-directed BTK, ERK and SYK, as well as IRAK1/IRAK4 activation. HCK is an SRC family member that mutated MYD88 transcriptionally upregulates, and its activation is triggered by IL-6/IL-6R/gp130/JAK2 signaling. BTK inhibitors are active in MYD88 mutated B-cell malignancies, but fail to produce complete responses, and show varying durations of response that are disease dependent and impacted by depth of response. Acquired resistance to BTK inhibitors is commonly related to a BTK Cys481 mutation. Pacritinib is a kinase inhibitor, FDA-approved for myelofibrosis, that is well tolerated and targets pro-signaling cascades that are relevant to mutated MYD88 signaling, including IRAK1, JAK2, and SRC, a homologue of HCK ( Fig. 1). Pacritinib may therefore represent a more effective option in blocking mutated MYD88 pro-survival signaling versus BTK-inhibitors. Methods: We performed comparative studies to evaluate the activity of pacritinib and the covalent BTK-inhibitors ibrutinib and zanubrutinib on mutated MYD88 relevant pro-survival signaling, as well as proliferation and survival in MYD88 mutated cell lines and primary MYD88-mutated WM cells. Further to these studies, we assessed the activity of pacritinib in well-characterized mutant (BTK Cys481Ser) expressing covalent BTK-inhibitor resistant WM and ABC DLBCL lymphoma cell models. Results: Pacritinib produced higher levels of apoptosis in MYD88-mutated WM (BCWM.1) and ABC DLBCL (TMD-8), and primary (CD19 +) MYD88-mutated WM cells relative to the BTK inhibitors ibrutinib or zanubrutinib.The apoptotic activity of pacritinib showed dose dependence with high levels of apoptosis at 0.5 µM, which is well within its pharmacologically achievable levels. Importantly, pacritinib, in addition to its known effects on JAK2 and IRAK1 activation, also showed robust inhibition of p-HCK (Y411) and its downstream signaling partner p-BTK (Y223) in MYD88 mutated BCWM.1 and TMD-8 cells. By combination index and normalized isobologram analyses, pacritinib showed synergistic interactions with covalent BTK inhibitors, and more so with the BCL-2 inhibitor venetoclax in MYD88 mutated cell lines and primary WM cells. Lastly, pacritinib alone and combined with venetoclax induced high levels of apoptosis in BTK Cys481Ser expressing covalent BTK-inhibitor resistant MYD88 mutated WM and ABC DLBCL lymphoma cells. Conclusions: Pacritinib more broadly extinguishes mutated MYD88 pro-survival signaling cascades ( Fig 1.) and demonstrates high levels of apoptotic activityin mutated MYD88 WM and ABC DLBCL cells versus selective covalent BTK-inhibitors. Pacritinib also synergizes with covalent BTK- and BCL2 inhibitors and can overcome covalent BTK-inhibitor resistance related to mutated BTK Cys481. Our studies provide a framework for investigating pacritinib in MYD88 mutated lymphomas. A clinical trial of pacritinib in relapsed/refractory WM is being initiated.

Clinical outcomes among patients with chronic lymphocytic leukemia (CLL)/small lymphocytic lymphoma (SLL) who received treatment with a covalent BTK and BCL2 inhibitor in the United States: a real-world database study

This retrospective study using the nationwide de-identified Flatiron Health electronic health record-derived database was designed to evaluate clinical outcomes among patients with chronic lymphocytic leukemia (CLL) who previously received both a covalent Bruton’s tyrosine kinase inhibitor (cBTKi) and B-cell lymphoma 2 inhibitor (BCL2i) in a real-world setting. Outcomes for the immediate next line of therapy following the latter of the cBTKi or BCL2i treatment included: real-world response rate of 34.4% (using methods most consistent with clinical trials); median duration of real-world response of 13.3 months; and median real-world progression-free survival of 9.2 months. Median overall survival was 25.5 months from the start of the immediate next line of therapy. There remains a need for more effective therapies after cBTKi and BCL2i therapy for patients with CLL.

Outcomes for Patients With Chronic Lymphocytic Leukemia (CLL) Previously Treated With Both a Covalent BTK and BCL2 Inhibitor in the United States: A Real-World Database Study.

This study describes the treatment patterns and outcomes of patients with CLL/SLL in a de-identified real-world oncology electronic health records database. Adult patients with CLL/SLL were eligible if they had received cBTKi therapy, both a cBTKi and a BCL2i, or all 4 drug classes (cBTKi, BCL2i, rituximab, and chemotherapy) at any time during the first 5 lines of therapy. Time-to-event outcomes were evaluated using Kaplan Meier method. No statistical comparisons were conducted; all analyses were descriptive and conducted using SAS Enterprise. A total of 9578 patients were eligible: 52.0% (n=4983) received at least one cBTKi, 6.1% (n=581) received both a cBTKi and BCL2i, and 2.3% (n=218) received all four therapies (cBTKi, BCL2i, rituximab, and chemotherapy). Of those who discontinued these treatments, only 39.5% (n=1 206/3 577), 59.7% (n=228/382), and 55.0% (n=82/149) received subsequent therapy (post-cBTKi, post-cBTKi/post-BCL2i, and post-all 4 therapies, respectively). Median time from treatment discontinuation of these therapies to the discontinuation of subsequent therapy or death was 9.5 months (all patients who discontinued the cBTKi) 5.6 months (those who discontinued both a cBTKi and BCL2i) and 3.9 months (patients who discontinued all four therapies). The median duration of the next treatment among those who received additional therapy was post-cBTKi treatment duration=4.1 months; post-cBTKi/post-BCL2i treatment duration=5.5 months; and median duration of the immediate next therapy after discontinuation of all 4 therapies=5.1 months. The poor outcomes observed across cohorts in this study demonstrate the need for effective treatments that can improve outcomes in patients with CLL/SLL.

Abstract 792: A novel covalent reversible BTK inhibitor for B cell malignancies with enhanced selectivity and in vivo efficacy

Abstract Bruton's tyrosine kinase (BTK), a member of Tec family kinase, plays a pivotal role in the regulation of BCR signaling, which is important for B cell development and function. Ibrutinib, which has been approved by FDA, is an effective treatment option for multiple types of B cell neoplasms. However, the treatment associated adverse events (AEs) are one of the major concerns due to its poor kinase selectivity . Here, we present a novel covalent, but reversible BTK inhibitor, XNW-1011, which demonstrated enhanced potency, better selectivity, improved drug-like properties and superior in vivo efficacy with a favorable safety profile. XNW-1011 was developed structurally and biochemically differently from current BTK inhibitors, being more potent and highly selective. In the in vitro assay, unlike Ibrutinib, it did not inhibit EGFR, ITK, JAK3, LYN and SRC. It completely suppressed the cell proliferation of a panel of lymphoma cell lines such as TMD-8 and Rec-1. Besides, XNW-1011 inhibited cellular phosphorylation of BTK and calcium efflux, confirming the inhibition of activation of BTK in vitro. We have shown that XNW-1011 formed covalent bond with cysteine in a cysteine reactivity assay. In addition, a washout assay by Western Blot confirmed the reversibility of inhibition of XNW-1011 to BTK. XNW-1011 has a good prolife of ADME, which translated into improved in vivo PK properties over Ibrutinib (low clearance and prolonged half-life). XNW-1011 was further evaluated in several animal models such as REC-1 and TMD-8 tumor xenografts. XNW-1011 inhibited tumor growth with as low as 3.125 mpk by BID and resulted in tumor regression with 12.5 mpk by BID. Interestingly, XNW-1011 completely inhibited the tumor growth of a patient derived xenograft (PDX) with Myd88 mutation. The dose proportionality was confirmed through determination of the plasma concentration and in vivo BTK occupancy in the last day of treatment. In summary, XNW-1011 is a potent, selective, covalent reversible BTK inhibitor with improved ADME prolife and in vivo efficacy, which makes it suitable for further evaluation in clinical setting. Citation Format: Xi Chen, Shaoqiang Huang, Fred Hu, Jun Xian, Jian Yang, Jing Qiang, Meijie Le, Yuchuan Wu. A novel covalent reversible BTK inhibitor for B cell malignancies with enhanced selectivity and in vivo efficacy [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 792.