Drugs in Clinical Pipeline: GDC-0834 | Treatment of Rheumatoid Arthritis | Antiinflammatory Agent | BTK Inhibitor

GDC-0834 [(R)-N-(3-(6-((4-(1,4-dimethyl-3-oxopiperazin-2-yl)phenyl)amino)-4-methyl-5-oxo-4,5-dihydropyrazin-2-yl)-2-methylphenyl)-4,5,6,7-tetrahydrobenzo[b]thiophene-2-carboxamide] is a potent, selective small molecule inhibitor of Bruton's tyrosine kinase (BTK). Its development followed SAR studies on CGI-1746, another potent and selective BTK inhbitor. GDC-0834 maintained the potency and selectivity of CGI-1746, but with much improved PK in preclinical animal models.  GDC-0834 inhibits BTK in vitro with an IC₅₀ of 0.006 uM and has an EC₅₀ of 0.060 uM in the cell based CD86 assay. In human whole blood, GDC-0834 demonstrated potent inhibition of both anti-IgE stimulated CD63 expression (basophils) and anti-IgD stimulated CD69 expression (B-cells) with EC₅₀’s of 0.35 and 0.38 uM, respectively [1].

Appreciating its potential for the treatment of rheumatoid arthritis, a single dose IND was filed and GDC-0834 was taken in to a single dose phase I trial in healthy volunteers to quickly evaluate the human pharmacokinetics. In human, GDC-0834 was found to be highly labile at the exo-cyclic amide bond that links the tetrahydrobenzothiophene moiety to the central aniline ring, resulting in insufficient parent drug exposure.

GDC-0834: 2D and 3D Structure

The activity of GDC-0834 is as follows:

IC₅₀ (BTK enzyme assay) = 0.006 uM

EC₅₀ (Cell Based CD86 assay) = 0.06 uM

Common Name: GDC-0834
Synonyms:  GDC-0834; GDC-0834; GDC 0834
IUPAC Name: (R)-N-(3-(6-((4-(1,4-dimethyl-3-oxopiperazin-2-yl)phenyl)amino)-4-methyl-5-oxo-4,5-dihydropyrazin-2-yl)-2-methylphenyl)-4,5,6,7-tetrahydrobenzo[b]thiophene-2-carboxamide
SMILES:O=C(C1=CC(CCCC2)=C2S1)NC3=CC=CC(C(N=C4NC5=CC=C([C@H]6N(C)CCN(C)C6=O)C=C5)=CN(C)C4=O)=C3C
CAS Number: 1133432-46-8
Mechanism of Action: Kinase Inhibitor; BTK Inhibitor
Indication: Anti-Inflammatory Agent; Treatment of Rheumatoid Arthritis
Development Stage: Pre-Clinical
Company: Gilead Pharmaceutical/Genentech

1H NMR (Estimated) for GDC-0834

References:
1. Young, W. B.; et. al. Potent and selective Bruton's tyrosine kinase inhibitors: discovery of GDC-0834. Bioorg Med Chem Lett 2015, 25(6), 1333-1337 (synthesis and activity).
2. Blomgren, P. A.; et. al. Substituted amides, methods of making, use thereof for the treatment of diseases such as cancer. WO2009039397A2 (synthesis and activity)

Drugs in Clinical Pipeline: ONO-4059

ONO-4059 is a highly potent, oral and selective Bruton’s tyrosine kinase (Btk) inhibitor with an IC₅₀ of 2.2 nM in biochemical assay. The compound covalently binds to BTK, and reversibly blocks BCR signaling and B-cell proliferation and activation. In a selectivity assay, ONO-4059 shows a two-fold potency against TEC kinase (IC₅₀ = 5.3 nM) but it is highly selective against Fyn (IC₅₀ = 2200 nM), Lck (IC₅₀ = 790 nM) and LynA (IC₅₀ = 3500 nM) [1].

In December 2014, Gilead Sciences signed a contract to help expedite development of Ono Pharmaceutical’s Ono-4059 program. The firms plan to work together to develop and commercialize ONO-4059, an oral Bruton's tyrosine kinase (BTK) inhibitor for the treatment of B-cell malignancies and other diseases. Gilead will have exclusive rights to develop and commercialize ONO-4059 in all countries of the world outside of Japan, South Korea, Taiwan, China, and the Association of Southeast Asian Nations countries (ASEAN), where Ono retains development and commercialization rights. Gilead hopes to develop ONO-4059 as a monotherapy or in combination with approved and investigational agents, including combinations with kinase inhibitors in Gilead's portfolio.

The activity of ONO-4059 is as follows:

IC₅₀ (BTK enzyme assay) = 2.2 nM

IC₅₀ (TEC enzyme assay) = 5.3 nM

IC₅₀ (FYN enzyme assay) = 2200 nM

IC₅₀ (LCK enzyme assay) = 790 nM

IC₅₀ (LYNA enzyme assay) = 3500 nM

What's the structure of ONO-4059?

Till date, Ono pharma or Gilead pharma has not revealed the structure for ONO-4059. One of the patent publication in 2015 [3] provides some clues to ONO-4059. The biochemical assays correlates with those reported earlier [1, 2]. The IC₅₀ value for BTK is 0.0021 uM, with 375-fold selectivity against LCK, 1057-fold against FYN and 1662-fold against LYNA.

The stereo-chemistry is a beta position. Interestingly, the base molecule has IC₅₀ value for BTK as 0.007 uM, with 114-fold selectivity against LCK, 762-fold against FYN and 471-fold against LYNA [4].

Common Name: ONO-4059

Synonyms: ONO-4059; ONO 4059; ONO4059; GS 4059; GS-4059; GS4059; ONO-WG-307

IUPAC Name:

CAS Number: 

SMILES:

Mechanism of Action: Kinase Inhibitor; BTK Inhibitor; Bruton’s Tyrosine Kinase Inhibitor

Indication: Various Cancers; Anti-inflammatory Agents

Development Stage: Phase I/II

Company: Ono Pharmaceutical\Gilead Pharma

References:
1. Hendriks, R. W.; et. al. Targeting Bruton's tyrosine kinase in B cell malignancies. Nat Rev Cancer 2014, 14(4), 219-232.
2. Yoshizawa, T.; et. al. Development of a Bruton’s Tyrosine Kinase (Btk) inhibitor, ONO-4059: Efficacy in a Collagen Induced Arthritis (CIA) Model Indicates Potential Treatment for Rheumatoid Arthritis (RA). #1660, ACR, November 12, 2012. (here)
3. Yamamoto, S.; et. al. Purinone derivative hydrochloride. US20150274731A1
4. Yamamoto, S.; et. al. Purinone derivative. US20150094299A1

Drugs in Clinical Pipeline: RN486

RN486 [6-cyclopropyl-8-fluoro-2-(2-hydroxymethyl-3-{1-methyl-5-[5-(4-methyl-piperazin-1-yl)-pyridin-2-ylamino]-6-oxo-1,6-dihydro-pyridin-3-yl}-phenyl)-2H-isoquinolin-1-one] is a potent, selective, reversible inhibitor of Bruton's tyrosine kinase (BTK). In the enzymatic assay, the compound potently inhibited Btk kinase activity with an IC₅₀ of 4.0 nM. RN486 binds the enzyme in a competitive manner as demonstrated in a time-resolved FRET-based competitive binding assay with an IC₅₀ of 0.3 nM. RN486 was shown to be highly selective when tested against a panel of 369 kinases in the Kinomescan. In the assay, the compound exhibited a strong and competitive binding to Btk with a K_d of 0.31 nM and a high degree of selectivity over almost all other kinases, including Syk and Janus kinase (JAK, K_d = 5.1 uM). The enzyme that was most potently inhibited next to Btk was Ste20-like kinase (SLK, K_d = 0.043 uM), for which the compound showed a 139-fold selectivity [1].

The activity of RN486 is as follows:

IC₅₀ (BTK enzyme assay) = 4.0 nM; K_d = 0.31 nM

K_d (JAK binding assay) = 5.1 uM

K_d (SLK binding assay) = 0.043 uM

Common Name: RN486

Synonyms: RN486; RN 486; RN-486

IUPAC Name: 6-cyclopropyl-8-fluoro-2-(2-hydroxymethyl-3-{1-methyl-5-[5-(4-methyl-piperazin-1-yl)-pyridin-2-ylamino]-6-oxo-1,6-dihydro-pyridin-3-yl}-phenyl)-2H-isoquinolin-1-one

CAS Number: 1242156-23-5

SMILES: OCc1c(cccc1n1ccc2c(c1=O)c(F)cc(c2)C1CC1)c1cc(Nc2ccc(cn2)N2CCN(CC2)C)c(=O)n( c1)C

Mechanism of Action: Kinase Inhibitor; BTK Inhibitor

Indication: Various Cancers; Anti-inflammatory Agents; Treatment for Rheumatoid Arthritis

Development Stage: Pre-Clinical

Company: Hoffmann-la-Roche

The production and effector function of antibodies are regulated by distinct immunoreceptors on B cells and innate immune cells. The receptors, termed B cell antigen receptor (BCR) and activating Fc receptor (FcR), belong to a family of Ig-like immunoreceptors containing an intracellular immunoreceptor tyrosine-based activation motif (ITAM). ITAMs act to integrate diverse antigen- or Fc-specific signals into a common pathway regulated by nonreceptor tyrosine kinases including Lyn, spleen tyrosine kinase (Syk), and Bruton's tyrosine kinase (Btk) from the sarcoma kinase, Syk, and Tec kinase families. These kinases relay the signals sequentially from ITAMs to phospholipase Cγ2 (PLCγ2) and thus play a critical and nonredundant role in the signal transduction of BCR and FcR. Consequently, loss-of-function mutation in the Btk gene results in severe B cell immunodeficiency and impaired FcR function in both patients with X-linked agammaglobulinemia and mutant mice with X-linked immunodeficiency.

Together, Btk and Syk regulate the signal transduction of ITAM-containing receptors or adaptors that are critical for autoantibody production, effector function, and osteoclast differentiation. Therefore, pharmacological inhibition of these enzymes may affect multiple steps in the pathogenesis of Rheumatoid arthritis (RA) and represent a useful approach for the treatment of the disease. Rheumatoid arthritis (RA) is an autoimmune joint disease characterized by chronic synovial inflammation and progressive joint destruction. The disease is often associated with the appearance of autoantibodies in both blood and inflamed joints. Several of these autoantibodies have emerged as potential arthritogenic factors. For example, anti-glucose-6 phosphate isomerase and anti-type II collagen antibodies, both of which are highly arthritogenic in mice, can be detected in patients with RA. In addition, anticitrullinated protein autoantibodies, the most prevalent in RA, can bind citrullinated fibrinogen in RA joints to form immune complexes, which stimulate macrophages to produce inflammatory cytokines such as TNFα. Lastly, clinical efficacy of B cell-depleting agents in RA strongly implicates autoantibodies as culprits in the pathogenesis of the disease.

When tested in the rat and mouse, RN486 exhibited an excellent pharmacokinetic profile. In the rat, it reached the maximal concentration of 2.5 µM at 4.5 h when dosed orally at 20 mg/kg and showed a half-life of 9.8 h in the blood when administered intravenously. In the mouse, the compound reached the maximal concentration of 6.0 µM at 3 h and a trough concentration of 1.0 µM at 24 h when dosed orally at 30 mg/kg [1].

Important facts about RN486:

a: RN486 Blocks both BCR and FcR Signaling.

b: RN486 Displays a Selective B Cell Inhibitory Profile in BioMAP Systems.

c: RN486 Displays Efficacy on Immune Arthritis Induced by both Active and Passive Immunization in Mice.

d: RN486 Inhibits Inflammation and Bone Erosions in Adjuvant-Induced Arthritis Either Alone or in Combination with Methotrexate.

e: RN486 Reduces Blood Inflammatory Markers in AIA.

Combination therapy with low-dose methotrexate is an important treatment option for patients with RA. Researchers therefore determined the potential of RN486 for combined therapy with methotrexate. To identify a suboptimal or low dose of methotrexate for combination study, researchers tested methotrexate alone in the AIA model at doses ranging from 0.025 to 0.25 mg/kg. Methotrexate displayed a dose-dependent inhibitory effect on both paw inflammation and splenomegaly in AIA rats, attenuating paw swelling by ~50 and 100%, respectively, at 0.075 and greater or equal 0.15 mg/kg. Researchers then tested RN486 and methotrexate at their respective suboptimal doses, 10 and 0.075 mg/kg, alone or in combination in the AIA model to assess the combined effect. As in the monotherapy studies, both RN486 and methotrexate attenuated paw swelling by approximately 50% when tested alone at the suboptimal doses. When combined, the two compounds completely eradicated paw swelling, splenomegaly, and histopathogical changes of inflammation and bone erosions [1].

References:

1. Xu, D.; et. al. RN486, a selective Bruton's tyrosine kinase inhibitor, abrogates immune hypersensitivity responses and arthritis in rodents. J Pharmacol Exp Ther 2012, 341(1), 90-103.

Drugs in Clinical Pipeline: Spebrutinib

Spebrutinib [N-(3-((5-fluoro-2-((4-(2-methoxyethoxy)phenyl)amino)pyrimidin-4-yl)amino)phenyl)acrylamide] is a covalent, highly selective, orally active small molecule inhibitor of Bruton’s agammaglobulinemia tyrosine kinase (BTK). Spebrutinib forms a covalent bond with Cys481 in Btk and potently inhibits Btk in biochemical (IC_50apparent less than 0.5 nM) and cellular assays (EC₅₀ 1-10 nM) including alpha-IgM stimulation of BCR signaling, B cell proliferation and activation. A quantitative pharmacodynamic assay to determine the level of Spebrutinib bonded to Btk in vitro or in vivo was developed and this drug-target engagement by Spebrutinib was shown to correlate directly with inhibition of Btk enzyme activity and substrate phosphorylation [1].

The activity of Spebrutinib is as follows:

Tec family members sharing a homologous Cys (Btk, BMX, Itk, Tec and Txk)

IC₅₀ (BTK enzyme assay) = 5.9 nM

IC₅₀ (BMX enzyme assay) = 0.7 nM

IC₅₀ (ITK enzyme assay) = 36 nM

IC₅₀ (TEC enzyme assay) = 6.2 nM

IC₅₀ (TXK enzyme assay) = 8.9 nM

Biochemical Activity against Src Family Kinases

IC₅₀ (Brk  enzyme assay) = 2430  nM
IC₅₀ (c-Src  enzyme assay) = 1729  nM
IC₅₀ (Csk  enzyme assay) = greater than 10000  nM
IC₅₀ (Fyn  enzyme assay) = 7146  nM
IC₅₀ (Hck  enzyme assay) = 14460  nM
IC₅₀ (Lck  enzyme assay) = 9079  nM
IC₅₀ (Lyn  enzyme assay) = 4401  nM
IC₅₀ (Yes  enzyme assay) = 723 nM

Common Name: Spebrutinib

Synonyms: CC-292; CC292; CC 292; AVL292; AVL-292; AVL 292

IUPAC Name: N-(3-((5-fluoro-2-((4-(2-methoxyethoxy)phenyl)amino)pyrimidin-4-yl)amino)phenyl)acrylamide

CAS Number: 1202757-89-8

SMILES: 

Mechanism of Action: Kinase Inhibitor; BTK Inhibitor

Indication: Various Cancers; Anti-inflammatory Agents; Treatment for Rheumatoid Arthritis

Development Stage: Phase II

Company: Avila Therapeutics, Inc.\Celegene

Targeted therapies that suppress B cell receptor (BCR) signaling have emerged as promising agents in autoimmune disease and B cell malignancies. Bruton's tyrosine kinase (Btk) plays a crucial role in B cell development and activation through the BCR signaling pathway and represents a new target for diseases characterized by inappropriate B cell activity. Bruton’s tyrosine kinase (Btk) is a kinase expressed exclusively in B cells and myeloid cells and has a well characterized, vital role in B cells highlighted by the human primary immune deficiency disease, X-linked agammaglobulinemia (XLA), which results from mutation in the Btk gene. Specifically, Btk plays an essential role in the B cell receptor (BCR) signaling pathway. Antigen binding to the BCR results in B cell receptor oligomerization, Syk and Lyn kinase activation, followed by Btk kinase activation. While BCR signaling is essential in the normal development and function of B cells, several pathologies have been attributed to dysregulated BCR activity. These include diseases of autoreactivity, such as that observed in lupus, multiple sclerosis, and rheumatoid arthritis, in which B cells inappropriately break self-tolerance to produce antibodies contributing to autoimmune disease. BCR signaling also contributes to several B cell malignancies, such as chronic lymphocytic leukemia (CLL), mantle cell lymphoma, and subsets of diffuse large B cell leukemia [1].

Biochemical kinase assays may overestimate the potency of Spebrutinib due to high ATP concentrations found in the cellular environment, hence cell activity for several of these closely related kinase family members was assessed. Spebrutinib demonstrated a high degree of selectivity against kinases with a cysteine in a homologous position as Cys481 in Btk (Epidermal growth factor receptor (A431 cell line), Itk (Jurkat cells), Janus kinase 3 (CTLL-2 cell line), EC₅₀ = 1-4 uM). To demonstrate specific inhibition of Btk in cells, Spebrutinib was evaluated in Ramos cells, which express an intact BCR signaling pathway that is activated robustly by addition of anti-IgM. Spebrutinib potently inhibited Btk autophosphorylation on Tyr223 (EC₅₀ = 8 nM), phosphorylation of the Btk substrate, PLCγ2, as well as activation of the downstream kinase extracellular signal-regulated kinase, all previously reported to be sensitive to Btk inhibition. It is noteworthy that while Spebrutinib inhibited autophosphorylation of Btk, it had no effect on the phosphorylation of Btk on Tyr551, a site phosphorylated by Lyn and Syk and required for Btk activation. These data demonstrate Spebrutinib is selective for Btk and does not inhibit the Src-family kinases upstream of Btk in the BCR signaling pathway [1].

Moreover, consistent with its covalent mechanism of action, Spebrutinib provided prolonged inhibition of kinase activity hours after the drug was removed from cells. In contrast to reversible inhibition with the potent Btk inhibitor Dasatinib, for which kinase activity had almost completely returned 6 hours after drug removal, recovery of Btk activity following a 1-hour exposure to Spebrutinib continued to be suppressed ~8 hours in drug-free media. Since Btk exposed to Spebrutinib is irreversibly bound and inhibited, the return of Btk-dependent signaling relies on the appearance of new Btk protein as a result of protein synthesis in a Spebrutinib-free environment [1].

Phase I Trials

A phase 1 trial investigated the safety, dose limiting toxicities (DLT), and clinical activity of Spebrutinib monotherapy in subjects with relapsed or refractory (R/R) CLL or non-Hodgkin's lymphoma.  This interim analysis focused on the safety and clinical activity in subjects with CLL and small cell lymphocytic leukemia (SLL). Eligible subjects with R/R (= 1 prior therapy) CLL/SLL were treated with monotherapy Spebrutinib in a dose-escalation study with doses ranging from 125 mg to 1000 mg QD and BID dose levels of 375 mg and 500 mg.  As a maximum tolerated dose was not established, CLL patients have been enrolled in an early dose expansion cohort of 750 mg QD and preliminary recommended phase 2 dose expansion cohort at 500 mg BID. All subjects received continuous dosing in 28-day cycles until progressive disease or intolerable toxicity. Clinical activity was investigator assessed per the 2008 iwCLL criteria.

Although the sample size is small, subjects treated at 375 mg or 500 mg BID showed continued lymph node size reduction over time from cycle 2 (mean reduction of 42% and 45%, respectively) to cycle 7 (mean reduction of 60% and 71%, respectively). The most common treatment-emergent AEs (= 10% of subjects) were diarrhea (59.7%), fatigue (37.5%), neutropenia (26.4%), thrombocytopenia (26.4%), nausea (26.4%), pyrexia (22.2%), headache (19.4%), cough (19.4%), upper respiratory infection (16.7%), peripheral edema (15.3%), abdominal pain (15.3%), dizziness (13.9%), muscle spasms (13.9%), contusion (13.9%), anemia (12.5%), pneumonia (12.5%), sinusitis (12.5%), and urinary tract infection (11.1%) [2]. It was concluded that Spebrutinib is well tolerated as an oral daily therapy. Single-agent therapy with Spebrutinib is sufficient to achieve high nodal and partial response rates in relapsed/refractory CLL subjects, including those with high-risk genomic features.

Researchers report the in vitro effects of the Spebrutinib as a single agent and in combination with several targeted small molecule inhibitors in Diffuse Large B-Cell Lymphoma (DLBCL), T-Cell Lymphoma (TCL), and Hodgkin Lymphoma (HL). DLBCL cell lines (GC: SUDHL6, SUDHL10, Farage, and OCI-LY19; and ABC: OCI-LY3) were treated with increasing concentrations of Spebrutinib (0.1-20 µM) alone and in combination with AKT inhibitor (AZD5363) and dual PI3K/mTOR inhibitor (BEZ235) for 24-72 hours.

Researchers analyzed cell viability with MTT assay and expression of NFκB, AKT, mTOR, MEK, and PARP by Western blot analysis. Treatment with Spebrutinib resulted in a dose dependent decrease in cell viability in all DLBCL lines. The 50% inhibitory concentration (IC₅₀) for Farage, OCI-LY19, SUDHL6, and SUDHL10 were 2.2 µM, 6.3 µM, 7.2 µM, and 3.3 µM respectively. Furthermore, treatment with 2.5 to 10 µM of Spebrutinib effectively reduced phosphorylation of mTOR, AKT, and MEK, while it increased phosphorylation of NFκB (p65) and cleaved PARP in Farage, OCI-LY19, and SUDHL10. Treatment with Spebrutinib in combination with BEZ235 (PI3K/mTOR inhibitor) showed synergistic cell death in Farage cells (CI value at IC₅₀, 75, and 90: 0.842, 0.731, and 0.661, respectively). Further, Spebrutinib treatment in combination with AZD5363 (AKT inhibitor) also resulted in synergistic cell death in Farage cells (CI value at IC₅₀, 75, 90: 0.772, 0.771, 0.698). Additionally, results demonstrated the cytotoxic effect of Spebrutinib in HL and TCL cell lines. Collectively, these results suggests that Spebrutinib is active as a single agent in TCL and HL and novel/novel combinations with PI3K/mTOR or AKT inhibitors may have potential therapeutic value in the treatment of DLBCL [3].

References:
1. Evans, E. K.; et. al. Inhibition of Btk with CC-292 provides early pharmacodynamic assessment of activity in mice and humans. J Pharmacol Exp Ther 2013, 346(2), 219-228.
2. Brown J. R.; et. al. 1630 Phase 1 Study Of Single Agent CC-292, a Highly Selective Bruton’s Tyrosine Kinase (BTK) Inhibitor, In Relapsed/Refractory Chronic Lymphocytic Leukemia (CLL). 55^th ASH Annual Meeting and Exposition, 2013, Dec 7-10, New Orleans LA.
3. Cerulli, R. A.; et. al. The Bruton’s Tyrosine Kinase Inhibitor CC-292 in Diffuse Large B-Cell Lymphoma (DLBCL), T-Cell Lymphoma (TCL), and Hodgkin Lymphoma (HL): Induction of Cell Death and Examination of Rational Novel/Novel Therapeutic Combinations. Blood 2014, 124(12).

4. ClinicalTrials.gov Efficacy and Safety Study of CC-292 Versus Placebo as Co-therapy With Methotrexate in Active Rheumatoid Arthritis. NCT01975610 (retrieved 15-09-2015).

5. ClinicalTrials.gov A Phase IB Study Of The BTKi CC-292 Combined With Lenalidomide In Adults Patients With Relapsed/Refractory B-Cell Lymphoma (CLEAR). NCT01766583 (retrieved 15-09-2015).

6. ClinicalTrials.gov Novel Combinations of CC-122, CC-223, CC-292, and Rituximab in Diffuse Large B-cell Lymphoma. NCT02031419 (retrieved 15-09-2015).

Acalabrutinib | ACP-196 | BTK Inhibitor | Cancer Drug I Treatment for Chronic Lymphocytic Leukemia

Acalabrutinib [(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamide] is a novel, second generation, irreversible Bruton's tyrosine kinase (BTK) inhibitor that may show advantages in terms of binding specificity and drug-drug interactions compared to the first generation BTK inhibitor. It is being developed by Acerta Pharma BV for treatment of Chronic Lymphocytic Leukemia (CLL). The improved selectivity and target coverage of Acalabrutinib has been credited to its covalent bonding with a cysteine residue (Cys481) in the front position of the ATP-binding pocket [1,2].

Acalabrutinib: 2D and 3D Structure

Targeting Bruton's tyrosine kinase (BTK), an essential kinase in the B cell receptor (BCR) pathway in patients with chronic lymphocytic leukemia (CLL) has proven very effective. For the first generation BTK inhibitor Ibrutinib, clinical response rates greater than 70% and 75% and progression free survival greater than 2 years have been reported for previously treated patients. Survival and proliferation of CLL cells is highly dependent on microenvironment interaction, which must be taken into consideration when testing new drugs for CLL.

The activity of Acalabrutinib is as follows:

EC₅₀ (BTK enzyme activity) = less than 10 nM
EC₅₀ (LCK enzyme activity) = greater than 1 uM
EC₅₀ (SRC enzyme activity) = greater than 1 uM
EC₅₀ (FYN enzyme activity) = greater than 1 uM
EC₅₀ (LYN enzyme activity) = greater than 1 uM

Common Name: Acalabrutinib
Synonyms: ACP-196; ACP 196; ACP196
IUPAC Name: (S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamide
CAS Number: 1420477-60-6
SMILES:O=C(NC1=NC=CC=C1)C2=CC=C(C3=C4C(N)=NC=CN4C([C@H]5N(C(C#CC)=O)CCC5)=N3)C=C2
Mechanism of Action: Kinase Inhibitor; BTK Inhibitor
Indication: Chronic Lymphocytic Leukemia
Development Stage: Phase II
Company: Acerta Pharma BV

Acalabrutinib Synthesis

EP2734522A1: First reported synthesis.

Bruton's tyrosine kinase (Btk) is a nonreceptor enzyme in the Tec kinase family expressed among cells of hematopoietic origin including B cells, myeloid cells, mast cells and platelets, but not T cells, where it regulates multiple cellular processes. 

ACP-196 shows in vivo efficacy against human chronic lymphocytic leukemia cells xenografted to the NSG mouse model. Peripheral blood mononuclear cells from previously untreated CLL patients were injected intravenously into NSG mice. Mice received ACP-196 through the drinking water. The effect of ACP-196 on xenografted CLL cells from peripheral blood and spleen was assessed by flow cytometry. At all dose levels tested, ACP-196 significantly inhibited proliferation of human CLL cells in the spleens of NSG mice. As seen with other BCR inhibitors, ACP-196 transiently increased CLL cell counts in the peripheral blood in a dose dependent manner (p = 0.01). ACP-196 inhibited BCR signaling in vivo, as demonstrated by reduced phosphorylation of PLCy2 [1]. 

ACP-196 demonstrated higher selectivity for Btk when profiled against a panel of 395 non-mutant kinases (greater than 1 µM) in a competitive binding assay. IC₅₀ determinations on 9 kinases with a Cys in the same position as Btk showed ACP-196 to be the most selective. The improved selectivity is related to the reduced intrinsic reactivity of ACP-196’s electrophile. Importantly, unlike ibrutinib, ACP-196 did not inhibit EGFR, ITK or TXK. Phosphoflow assays on EGFR expressing cell lines confirmed ibrutinib’s EGFR inhibition (EC₅₀ = 47-66 nM) with no inhibition observed for ACP-196 at 10 µM. These data may explain the Ibrutinib-related incidence of diarrhea and rash [3].

For Canine Lovers 

ACP-196 has been tested in normal dogs and was found to be safe at doses to be used in this clinical trial. Given the demonstrated activity of ibrutinib in dogs with B cell lymphoma, it is expected that ACP-196 will show good activity in this setting as well.

Identifications:

1H NMR (Estimated) for Acalabrutinib

References:
1. Niemann, C. U.; et. al. Abstract 2624: The novel Bruton's tyrosine kinase inhibitor ACP-196 shows in vivo efficacy against human chronic lymphocytic leukemia cells xenografted to the NSG mouse model. Cancer Res 2014, 74, 2624.
2. Lannutti, B. J.; et. al. Abstract 408: ACP-196, an orally bioavailable covalent selective inhibitor of Btk, modulates the innate tumor microenvironment, exhibits antitumor efficacy and enhances gemcitabine activity in pancreatic cancer. Cancer Res 2015, 75, 408.
3. Covey, T.; et. al. Abstract 2596: ACP-196: a novel covalent Bruton's tyrosine kinase (Btk) inhibitor with improved selectivity and in vivo target coverage in chronic lymphocytic leukemia (CLL) patients. Cancer Res 2015, 75, 2596.
4. ClinicalTrials.gov ACP-196, a Novel Bruton Tyrosine Kinase (Btk) Inhibitor, for Treatment of Chronic Lymphocytic Leukemia. NCT02029443 (retrieved on 20-04-2015)
5. ClinicalTrials.gov An Open-label, Phase 1b Study of ACP 196 in Subjects With Waldenström Macroglobulinemia. NCT02180724 (retrieved on 20-04-2015)
6. Barf, T. A.; et. al. 4-imidazopyridazin-1-yl-benzamides and 4- imidazotriazin-1-yl-benzamides as btk-inhibitors. EP2734522A1