Kee Chuan Goh

SIRT1 Modulating Compounds from High-Throughput Screening as Anti-Inflammatory and Insulin-Sensitizing Agents

The nicotinamide adenine dinucleotide (NAD+)-dependent protein deacetylase SIRT1 has been linked to fatty acid metabolism via suppression of peroxysome proliferator-activated receptor gamma (PPAR-γ) and to inflammatory processes by deacetylating the transcription factor NF-κB. First, modulation of SIRT1 activity affects lipid accumulation in adipocytes, which has an impact on the etiology of a variety of human metabolic diseases such as obesity and insulin-resistant diabetes. Second, activation of SIRT1 suppresses inflammation via regulation of cytokine expression. Using high-throughput screening, the authors identified compounds with SIRT1 activating and inhibiting potential. The biological activity of these SIRT1-modulating compounds was confirmed in cell-based assays using mouse adipocytes, as well as human THP-1 monocytes. SIRT1 activators were found to be potent lipolytic agents, reducing the overall lipid content of fully differentiated NIH L1 adipocytes. In addition, the same compounds have anti-inflammatory properties, as became evident by the reduction of the proinflammatory cytokine tumor necrosis factor-alpha (TNF-α). In contrast, a SIRT1 inhibitory compound showed a stimulatory activity on the differentiation of adipocytes, a feature often linked to insulin sensitization.

SB939, a novel potent and orally active histone deacetylase inhibitor with high tumor exposure and efficacy in mouse models of colorectal cancer.

Although clinical responses in liquid tumors and certain lymphomas have been reported, the clinical efficacy of histone deacetylase inhibitors in solid tumors has been limited. This may be in part due to the poor pharmacokinetic of these drugs, resulting in inadequate tumor concentrations of the drug. SB939 is a new hydroxamic acid based histone deacetylase inhibitor with improved physicochemical, pharmaceutical, and pharmacokinetic properties. In vitro, SB939 inhibits class I, II, and IV HDACs, with no effects on other zinc binding enzymes, and shows significant antiproliferative activity against a wide variety of tumor cell lines. It has very favorable pharmacokinetic properties after oral dosing in mice, with >4-fold increased bioavailability and 3.3-fold increased half-life over suberoylanilide hydroxamic acid (SAHA). In contrast to SAHA, SB939 accumulates in tumor tissue and induces a sustained inhibition of histone acetylation in tumor tissue. These excellent pharmacokinetic properties translated into a dose-dependent antitumor efficacy in a xenograft model of human colorectal cancer (HCT-116), with a tumor growth inhibition of 94% versus 48% for SAHA (both at maximum tolerated dose), and was also effective when given in different intermittent schedules. Furthermore, in APCmin mice, a genetic mouse model of early-stage colon cancer, SB939 inhibited adenoma formation, hemocult scores, and increased hematocrit values more effectively than 5-fluorouracil. Emerging clinical data from phase I trials in cancer patients indicate that the pharmacokinetic and pharmacologic advantages of SB939 are translated to the clinic. The efficacy of SB939 reported here in two very different models of colorectal cancer warrants further investigation in patients. Mol Cancer Ther; 9(3); 642–52

Discovery of the Macrocycle 11-(2-Pyrrolidin-1-yl-ethoxy)-14, 19-dioxa-5,7,26-triaza-tetracyclo[19.3.1.1(2,6).1(8,12)]heptacosa-1(25),2(26),3,5,8,10,12(27),16,21,23-decaene (SB1518), a Potent Janus Kinase 2/Fms-Like Tyrosine Kinase-3 (JAK2/FLT3) Inhibitor for the Treatment of Myelofibrosis and Lymphoma

Discovery of the activating mutation V617F in Janus Kinase 2 (JAK2(V617F)), a tyrosine kinase critically involved in receptor signaling, recently ignited interest in JAK2 inhibitor therapy as a treatment for myelofibrosis (MF). Herein, we describe the design and synthesis of a series of small molecule 4-aryl-2-aminopyrimidine macrocycles and their biological evaluation against the JAK family of kinase enzymes and FLT3. The most promising leads were assessed for their in vitro ADME properties culminating in the discovery of 21c, a potent JAK2 (IC(50) = 23 and 19 nM for JAK2(WT) and JAK2(V617F), respectively) and FLT3 (IC(50) = 22 nM) inhibitor with selectivity against JAK1 and JAK3 (IC(50) = 1280 and 520 nM, respectively). Further profiling of 21c in preclinical species and mouse xenograft and allograft models is described. Compound 21c (SB1518) was selected as a development candidate and progressed into clinical trials where it is currently in phase 2 for MF and lymphoma.

Discovery of (2E)-3-{2-Butyl-1-[2-(diethylamino)ethyl]-1H-benzimidazol-5-yl}-N-hydroxyacrylamide (SB939), an Orally Active Histone Deacetylase Inhibitor with a Superior Preclinical Profile

A series of 3-(1,2-disubstituted-1H-benzimidazol-5-yl)-N-hydroxyacrylamides (1) were designed and synthesized as HDAC inhibitors. Extensive SARs have been established for in vitro potency (HDAC1 enzyme and COLO 205 cellular IC(50)), liver microsomal stability (t(1/2)), cytochrome P450 inhibitory (3A4 IC(50)), and clogP, among others. These parameters were fine-tuned by carefully adjusting the substituents at positions 1 and 2 of the benzimidazole ring. After comprehensive in vitro and in vivo profiling of the selected compounds, SB939 (3) was identified as a preclinical development candidate. 3 is a potent pan-HDAC inhibitor with excellent druglike properties, is highly efficacious in in vivo tumor models (HCT-116, PC-3, A2780, MV4-11, Ramos), and has high and dose-proportional oral exposures and very good ADME, safety, and pharmaceutical properties. When orally dosed to tumor-bearing mice, 3 is enriched in tumor tissue which may contribute to its potent antitumor activity and prolonged duration of action. 3 is currently being tested in phase I and phase II clinical trials.

Pacritinib (SB1518), a JAK2/FLT3 inhibitor for the treatment of acute myeloid leukemia.

FMS-like tyrosine kinase 3 (FLT3) is the most commonly mutated gene found in acute myeloid leukemia (AML) patients and its activating mutations have been proven to be a negative prognostic marker for clinical outcome. Pacritinib (SB1518) is a tyrosine kinase inhibitor (TKI) with equipotent activity against FLT3 (IC50=22 n) and Janus kinase 2 (JAK2, IC50=23 n). Pacritinib inhibits FLT3 phosphorylation and downstream STAT, MAPK and PI3 K signaling in FLT3-internal-tandem duplication (ITD), FLT3-wt cells and primary AML blast cells. Oral administration of pacritinib in murine models of FLT3-ITD-driven AML led to significant inhibition of primary tumor growth and lung metastasis. Upregulation of JAK2 in FLT3-TKI-resistant AML cells was identified as a potential mechanism of resistance to selective FLT3 inhibition. This resistance could be overcome by the combined FLT3 and JAK2 activities of pacritinib in this cellular model. Our findings provide a rationale for the clinical evaluation of pacritinib in AML including patients resistant to FLT3-TKI therapy.

Discovery of Kinase Spectrum Selective Macrocycle (16E)-14-Methyl-20-oxa-5,7,14,26-tetraazatetracyclo[19.3.1.1(2,6).1(8,12)]heptacosa-1(25),2(26),3,5,8(27),9,11,16,21,23-decaene (SB1317/TG02), a Potent Inhibitor of Cyclin Dependent Kinases (CDKs), Janus Kinase 2 (JAK2), and Fms-like Tyrosine Kinase-3 (FLT3) for the Treatment of Cancer

Herein, we describe the design, synthesis, and SAR of a series of unique small molecule macrocycles that show spectrum selective kinase inhibition of CDKs, JAK2, and FLT3. The most promising leads were assessed in vitro for their inhibition of cancer cell proliferation, solubility, CYP450 inhibition, and microsomal stability. This screening cascade revealed 26 h as a preferred compound with target IC(50) of 13, 73, and 56 nM for CDK2, JAK2 and FLT3, respectively. Pharmacokinetic (PK) studies of 26 h in preclinical species showed good oral exposures. Oral efficacy was observed in colon (HCT-116) and lymphoma (Ramos) xenograft studies, in line with the observed PK/PD correlation. 26h (SB1317/TG02) was progressed into development in 2010 and is currently undergoing phase 1 clinical trials in advanced leukemias and multiple myeloma.

VS-5584, a Novel and Highly Selective PI3K/mTOR Kinase Inhibitor for the Treatment of Cancer

Dysregulation of the PI3K/mTOR pathway, either through amplifications, deletions, or as a direct result of mutations, has been closely linked to the development and progression of a wide range of cancers. Moreover, this pathway activation is a poor prognostic marker for many tumor types and confers resistance to various cancer therapies. Here, we describe VS-5584, a novel, low-molecular weight compound with equivalent potent activity against mTOR (IC50 = 37 nmol/L) and all class I phosphoinositide 3-kinase (PI3K) isoforms IC50: PI3Kα = 16 nmol/L; PI3Kβ = 68 nmol/L; PI3Kγ = 25 nmol/L; PI3Kδ = 42 nmol/L, without relevant activity on 400 lipid and protein kinases. VS-5584 shows robust modulation of cellular PI3K/mTOR pathways, inhibiting phosphorylation of substrates downstream of PI3K and mTORC1/2. A large human cancer cell line panel screen (436 lines) revealed broad antiproliferative sensitivity and that cells harboring mutations in PI3KCA are generally more sensitive toward VS-5584 treatment. VS-5584 exhibits favorable pharmacokinetic properties after oral dosing in mice and is well tolerated. VS-5584 induces long-lasting and dose-dependent inhibition of PI3K/mTOR signaling in tumor tissue, leading to tumor growth inhibition in various rapalog-sensitive and -resistant human xenograft models. Furthermore, VS-5584 is synergistic with an EGF receptor inhibitor in a gastric tumor model. The unique selectivity profile and favorable pharmacologic and pharmaceutical properties of VS-5584 and its efficacy in a wide range of human tumor models supports further investigations of VS-5584 in clinical trials. Mol Cancer Ther; 12(2); 151–61. ©2012 AACR.

Structure-based design of oxygen-linked macrocyclic kinase inhibitors: discovery of SB1518 and SB1578, potent inhibitors of Janus kinase 2 (JAK2) and Fms-like tyrosine kinase-3 (FLT3)

Macrocycles from our Aurora project were screened in a kinase panel and were found to be active on other kinase targets, mainly JAKs, FLT3 and CDKs. Subsequently these compounds became leads in our JAK2 project. Macrocycles with a basic nitrogen in the linker form a salt bridge with Asp86 in CDK2 and Asp698 in FLT3. This residue is conserved in most CDKs resulting in potent pan CDK inhibition. One of the main project objectives was to achieve JAK2 potency with 100-fold selectivity against CDKs. Macrocycles with an ether linker have potent JAK2 activity with the ether oxygen forming a hydrogen bond to Ser936. A hydrogen bond to the equivalent residues of JAK3 and most CDKs cannot be formed resulting in good selectivity for JAK2 over JAK3 and CDKs. Further optimization of the macrocyclic linker and side chain increased JAK2 and FLT3 activity as well as improving DMPK properties. The selective JAK2/FLT3 inhibitor 11 (Pacritinib, SB1518) has successfully finished phase 2 clinical trials for myelofibrosis and lymphoma. Another selective JAK2/FLT3 inhibitor, 33 (SB1578), has entered phase 1 clinical development for the non-oncology indication rheumatoid arthritis.

Discovery of the Macrocycle (9E)-15-(2-(Pyrrolidin-1-yl)ethoxy)-7,12,25-trioxa-19,21,24-triaza-tetracyclo[18.3.1.1(2,5).1(14,18)]hexacosa-1(24),2,4,9,14(26),15,17,20,22-nonaene (SB1578), a Potent Inhibitor of Janus Kinase 2/Fms-LikeTyrosine Kinase-3 (JAK2/FLT3) for the Treatment of Rheumatoid Arthritis

Herein, we describe the synthesis and SAR of a series of small molecule macrocycles that selectively inhibit JAK2 kinase within the JAK family and FLT3 kinase. Following a multiparameter optimization of a key aryl ring of the previously described SB1518 (pacritinib), the highly soluble 14l was selected as the optimal compound. Oral efficacy in the murine collagen-induced arthritis (CIA) model for rheumatoid arthritis (RA) supported 14l as a potential treatment for autoimmune diseases and inflammatory disorders such as psoriasis and RA. Compound 14l (SB1578) was progressed into development and is currently undergoing phase 1 clinical trials in healthy volunteers.

SB1578, a Novel Inhibitor of JAK2, FLT3, and c-Fms for the Treatment of Rheumatoid Arthritis

SB1578 is a novel, orally bioavailable JAK2 inhibitor with specificity for JAK2 within the JAK family and also potent activity against FLT3 and c-Fms. These three tyrosine kinases play a pivotal role in activation of pathways that underlie the pathogenesis of rheumatoid arthritis. SB1578 blocks the activation of these kinases and their downstream signaling in pertinent cells, leading to inhibition of pathological cellular responses. The biochemical and cellular activities of SB1578 translate into its high efficacy in two rodent models of arthritis. SB1578 not only prevents the onset of arthritis but is also potent in treating established disease in collagen-induced arthritis mice with beneficial effects on histopathological parameters of bone resorption and cartilage damage. SB1578 abrogates the inflammatory response and prevents the infiltration of macrophages and neutrophils into affected joints. It also leads to inhibition of Ag-presenting dendritic cells and inhibits the autoimmune component of the disease. In summary, SB1578 has a unique kinase spectrum, and its pharmacological profile provides a strong rationale for the ongoing clinical development in autoimmune diseases.