Pim Kinase Inhibitors Research Articles

A novel Pim-1 kinase-targeted photosensitizer to combat triple-negative breast cancer with enhanced photodynamic efficacy and reduced metastasis

As one of the most malignant breast cancer types, triple-negative breast cancer (TNBC) treatment is a particularly great challenge in the current era of precision medicine. Due to the limited effectiveness and targeting of current treatment methods, it is urgently need to develop an effective precision strategy to combat TNBC. Herein, a novel TNBC targeted molecular drug based on zinc(II) phthalocyanine (BF-Pc) was rationally designed by conjugating a Pim-1 kinase inhibitor with high TNBC selectivity. In our in vitro investigation, BF-Pc not only exhibits extremely high targeting to TNBC cells overexpressed Pim-1 kinases, but also shows ultra-efficient photocytotoxicity toward TNBC cells far beyond the normal cells. Significantly, BF-Pc-induced PDT can obviously suppress TNBC cell migration and invasion. The proposed manner in our work offers a promising approach to combat TNBC effectively and precisely.

Nicotinonitrile-derived apoptotic inducers: Design, synthesis, X-ray crystal structure and Pim kinase inhibition

Although a plethora of targeted anticancer small molecule drugs became available, the low response rate and drug resistance imply the continuous need for expanding the anticancer chemical space. In this study, a novel series of nicotinonitrile derivatives was designed, synthesized and evaluated for cytotoxic activities in HepG2 and MCF-7 cells. All derivatives showed high to moderate cytotoxic activity against both cell lines, with cell-type and chemotype-dependent cytotoxic potential. The normal HEK-293 T cells were ca. 50-fold less susceptible to the cytotoxic effect of the inhibitors. The in vitro enzyme inhibitory activity of selected active cytotoxic derivatives 8c, 8e, 9a, 9e and 12 showed that they have sub- to one digit micromolar 50 % inhibitory concentration (IC50) against the three Pim kinase isoforms, with 8e being the most potent (IC50 ≤ 0.28 μM against three Pim kinases), comparable to the pan kinase inhibitor, Staurosporine. In HepG2, 8e induced cell cycle arrest at the G2/M phase. Apoptotic mechanistic studies with 8c and 8e in HepG2 cells, indicated a significant upregulation in both P53 and caspase-3 relative gene expression, as well as increased Bax/Bcl-2 protein expression level. Further, docking studies combined with molecular dynamic simulation showed a stable complex with high binding affinity of 8e to Pim-1 kinase; exploiting a negative electrostatic potential surface interaction with the added dimethyl amino group in the new compounds. Moreover, in silico ADME profile prediction indicated that all compounds are orally bioavailable and most of them can penetrate the blood–brain barrier. This study presents novel nicotinonitrile derivatives as auspicious hits for further optimization as antiproliferative agents against liver cancer cells and promising pan Pim kinase inhibitors at submicromolar concentrations.

Development of novel cyanopyridines as PIM-1 kinase inhibitors with potent anti-prostate cancer activity: Synthesis, biological evaluation, nanoparticles formulation and molecular dynamics simulation

Recently, inhibition of PIM-1 enzyme is found as an effective route in the fight against proliferation of cancer. Herein, new cyano pyridines that target PIM-1 kinase were designed, synthesized, and biologically evaluated. Two prostate cell lines were used to examine each of the new compounds in vitro for anticancer activity, namely, PC-3 and DU-145. The cyanopyridine derivatives 2b, 3b, 4b, and 5b with an N,N-dimethyl phenyl group at the pyridine ring's 4-position showed considerable antitumor effect on the tested cell lines. Additionally, the high selectivity index revealed that these compounds were less cytotoxic to normal WI-38 cells. Furthermore, they exhibited strong inhibitory effect on PIM-1 having IC50 = 0.248, 0.13, 0.326 and 0.245 μM, respectively. The most powerful derivatives2b, 3b, 4b, and 5b, were chosen for further examination of their inhibitory potential on both kinases (PIM-2 and PIM-3). Interestingly, upon loading compound 3b in a cubosomes formulation with nanometric size, improvements in cytotoxicity and inhibitory effect on PIM-1 kinase were observed. In silico ADME parameters study revealed that compound 3b is orally bioavailable without penetration to the blood-brain barrier. Further, the docking simulations revealed the ability of our potent compounds to well accommodate the PIM-1 kinase active site forming stable complexes. In a 150 ns MD simulation, the most powerful PIM-1 inhibitor 3b produced stable complex with the PIM-1 enzyme (RMSD = 1.76). Furthermore, the 3b–PIM-1 complex has the low binding free energy (−242.2 kJ/mol) according to the MM-PBSA calculations.

Computational modelling strategies in exploring triazolopyridazine PIM1 kinase inhibitors as anticancer agents.

PIM (Proviral Integration site for Moloney Murine Leukemia virus) kinases are members of the class of kinase family serine/ threonine kinases, which play a crucial role in cancer development. As there is no drug in the market against PIM-1, kinase has transpired as a budding and captivating target for discovering new anticancer agents targeting PIM-1 kinase. The current research pondered the development of new PIM-1 kinase inhibitors by applying a ligand-based and structure-based drug discovery approach involving 3D QSAR, molecular docking, and dynamics simulation. In this study, association allying the structural properties and biological activity was undertaken using 3D-QSAR analysis. The 3D-QSAR model was generated with the help of 35 compounds from which the best model manifested an appreciated cross-validation coefficient (q2) of 0.8866 and conventional correlation coefficient (r2) of 0.9298, respectively and predicted correlation coefficient (r2 pred) was obtained as 0.7878. The molecular docking analysis demonstrated that the analogs under analysis occupied the active site of PIM-1 kinase receptor and interactions with Lys67 in the catalytic region, Asp186 in the DFG motif, and Glu171 were noticed with numerous compounds. Furthermore, the molecular dynamics simulation study stated that the ligand portrayed the strong conformational stability within the active site of PIM-1 kinase protein, forming of two hydrogen bonds until 100 ns, respectively. Overall outcomes of the study revealed that applications of the ligand-based drug discovery approach and structure-based drug discovery strategy conceivably applied to discovering new PIM-1 kinase inhibitors as anticancer agents.

Stereoselective synthesis of 3,4-dihydropyrrolo[1,2-a]pyrazin-1(2H)-one derivatives as PIM kinase inhibitors inspired from marine alkaloids.

We previously demonstrated that natural product-inspired 3,4-dihydropyrrolo[1,2-a]pyrazin-1(2H)-ones derivatives delivered potent and selective PIM kinases inhibitors however with non-optimal ADME/PK properties and modest oral bioavailability. Herein, we describe a structure-based scaffold decoration and a stereoselective approach to this chemical class. The synthesis, structure-activity relationship studies, chiral analysis, and pharmacokinetic data of compounds from this inhibitor class are presented herein. Compound 20c demonstrated excellent potency on PIM1 and PIM2 with exquisite kinases selectivity and PK properties that efficiently and dose-dependently promoted c-Myc degradation and appear to be promising lead compounds for further development.

Synthesis of new pyrazolo[4,3-a]phenanthridine Pim-1 inhibitors and evaluation of their cytotoxic activity towards the MOLM-13 acute myeloid leukemia cell line

We synthesized new analogues of the anti-AML agent VS-II-173. We studied the effect of the substitution at the 1- and 5-positions of the pyrazolo[4,3-a]phenanthridine scaffold on Pim-1 kinase inhibition and cytotoxicity against AML MOLM-13 cells. We found that compounds 20 and 21, substituted at the 1-position exhibited stronger Pim-1 inhibition together with a high potency toward MOLM-13 cells, associated with apoptosis induction and selectivity over non-cancerous NRK cells.

Design, synthesis, and bioactivity evaluation of macrocyclic benzo[b]pyrido[4,3-e][1,4]oxazine derivatives as novel Pim-1 kinase inhibitors

Pim-1 kinase is a serine/threonine kinase which is vital in many tumors. The Pim-1 inhibitor 10-DEBC and its derivatives discovered in our previous work were modified through macrocyclization strategy. A series of benzo[b]pyridine[4,3-e][1,4]oxazine macrocyclic compounds were designed, synthesized, and evaluated as novel Pim-1 kinase inhibitors. Among these compounds, compound H5 exhibited the highest activity with an IC50 value of 35 nM. In addition, the crystal complex structure of Pim-1 kinase bound with compound H3 was determined, and the structure–activity relationship of these macrocyclic compounds was analyzed, which provides the structural basis of further optimization of novel macrocyclic Pim-1 kinase inhibitors..

Abstract 3338: Concurrent treatment with Pim kinase inhibitor enhances efficacy of FLT3 inhibitors in AML with FLT3-ITD through GSK-3βactivation and GSK-3β-mediated enhanced proteasomal degradation of c-Myc and Mcl-1

Abstract BACKGROUND: fms-like tyrosine kinase 3 internal tandem duplication (FLT3-ITD) is present in acute myeloid leukemia (AML) cells in 30% of patients, with poor patient outcomes. FLT3 inhibitors are in clinical use, but with incomplete responses and onset of resistance. New targeting strategies are needed. The oncogenic serine/threonine kinase proviral integration site for Moloney murine leukemia virus (Pim-1) is upregulated downstream of FLT3-ITD, directly stimulates cell growth and inhibits apoptosis, and also phosphorylates and stabilizes FLT3 in a positive feedback loop. We previously showed that concurrent treatment of FLT3-ITD cells with Pim kinase and FLT3 inhibitors enhanced apoptosis induction through post-translational downregulation of the master transcription factor c-Myc and the anti-apoptotic protein Mcl-1. The serine/threonine kinase GSK-3β regulates c-Myc and Mcl-1 protein via phosphorylation at T58 and S159, respectively; we hypothesized downregulation of both proteins by combination treatment through phosphorylation by GSK-3β. METHODS: FLT3-ITD cells lines (Ba/F3-ITD, MV4-11, and MOLM-14) and primary FLT3-ITD AML patient blasts were cultured with the pan-Pim kinase inhibitor AZD1208, the FLT3 inhibitors gilteritinib or quizartinib and the GSK-3β inhibitor TCG-24. Protein expression was measured by immunoblotting. To measure protein half-lives, cells were pretreated with cycloheximide with and without the proteasome inhibitor MG-132. Ba/F3-ITD cells were infected with Myc-T58A plasmid, preventing c-Myc T58 phosphorylation, or Mcl-1 S159A plasmid, preventing Mcl-1 S159 phosphorylation, or empty vector controls. Apoptosis was measured by Annexin V labeling, measured by flow cytometry. RESULTS: Combined treatment of FLT3-ITD-expressing cell lines and AML patient blasts with Pim and FLT3 inhibitors caused rapid downregulation of c-Myc, and then Mcl-1, protein expression, relative to single drugs. Combined Pim and FLT3 inhibitor treatment caused marked decrease in half-lives of both proteins, which was rescued by proteasome inhibition, as the mechanism of protein downregulation. Combined Pim and FLT3 inhibitor treatment rapidly activated (dephosphorylated) GSK-3β and culture with GSK-3β inhibitor prevented induction of c-Myc and Mcl-1 downregulation, increased c-Myc and Mcl-1 protein turnover by concurrent Pim and FLT3 inhibitor treatment. Finally, combination treatment of Ba/F3-ITD cells infected with T58A c-Myc or S159A Mcl-1 plasmids did not downregulate these proteins or increase their turnover, with reduced apoptosis induction. CONCLUSION: Pim inhibitors enhance efficacy of FLT3 inhibitors via GSK-3β activation and GSK-3β-mediated enhanced proteasomal degradation of c-Myc and Mcl-1, highlighting GSK-3β as a key target in cells with FLT3-ITD. Citation Format: Jonelle Lee, Mario Scarpa, Aditi Chatterjee, Mooath Mustafa Ali, Prerna Singh, Shivani Kapoor, Rossana Trotta, Maria Baer. Concurrent treatment with Pim kinase inhibitor enhances efficacy of FLT3 inhibitors in AML with FLT3-ITD through GSK-3βactivation and GSK-3β-mediated enhanced proteasomal degradation of c-Myc and Mcl-1 [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 3338.

Abstract 263: PIM kinase regulates actin dynamics in hypoxia

Abstract One of the leading factors in prostate cancer mortality is the onset of metastatic events. This is seen by the drastic decline in survival rates following tumor escape from the primary site. A key promoter of metastasis is tumor hypoxia, or low oxygen concentration. Our lab previously demonstrated that Proviral Integration site for Moloney murine leukemia virus (PIM)-1, a serine-threonine kinase, is upregulated in hypoxia and that inhibition of PIM1 reduced metastatic occurrence in orthotopic mouse models. Here, we identify a novel signaling pathway that links hypoxia and PIM kinase to the actin cytoskeleton and cell motility. An unbiased screen to discover new PIM substrates in hypoxia identified Abl-Interactor 2 (Abi2), an integral member of the actin regulating Wave Regulatory Complex (WRC), as a novel PIM kinase substrate. Biochemical analysis demonstrates that phosphorylation of Abi2 at our identified PIM sites increases Abi2 stability. Overexpression of PIM1 increases Abi2 and other WRC members at the protein level, resulting in increased actin protrusions and cell motility. CRISPR-generated Abi2 knockout cell lines showed diminished protrusive activity and negated both the necessity and sufficiency of PIM kinase to increases protrusive activity in cancer cells. Lastly, in vivo experiments show that overexpression of PIM1 increases tumor invasion into the surrounding tissue in an Abi2-dependent manner, and small molecule PIM inhibitors significantly reduce invasion. In summary, we uncovered a novel role for PIM1 in promoting cellular motility and protrusion, identified novel phosphorylation sites that control the stability of Abi2 in normoxic and hypoxic conditions, and highlight a novel PIM1/Abi2 signaling axis that regulates hypoxia-induced tumor invasion. This research provides important insight in the role of PIM signaling in prostate tumors and yields preclinical evidence for further use of PIM kinase inhibitors as a means to inhibit tumor invasion, a key driving step in the formation of lethal prostate cancer metastasis. Citation Format: Corbin C. Jensen, Noel A. Warfel. PIM kinase regulates actin dynamics in hypoxia [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 263.

Identification of Pim-1 Kinase Inhibitors by Pharmacophore Model, Molecular Docking-based Virtual Screening, and Biological Evaluation.

This study aimed at screening and development of Pim-1 inhibitors as anticancer agent. Pim-1, a member of the Ser/Thr kinase family, plays a crucial role in cell proliferation and is being regarded as a promising target for cancer therapeutics. The present work focused on screening more potent Pim-1 inhibitors by in-silico method and biological evaluation. To identify more potent Pim-1 inhibitors, a GALAHAD pharmacophore model was constructed based on nine known Pim-1 inhibitors and followed by in silico screening including pharmacophore and molecular docking-based virtual screening. The hit compounds were further assessed the Pim-1, 2, and 3 kinase activities and the anticancer inhibition property against human myeloma RPMI-8226 and U266 cells using cytotoxicity studies. Based on Qfit value (from pharmacophore), docking score and clustering analysis, six compounds including C445_0268, C470_0769, 4456_0744, 0806_0325, G395_1510 and V023_3227 were hit. Binding mode analysis showed that hydrogen bond, hydrophobic and π-π stacking interactions dominated the bindings of these compounds to Pim-1. The further biological evaluation indicated that compounds C445_0268 and C470_0769 possessed excellent pan-Pim kinase activities and inhibited the growths of RPMI-8226 and U266 cell lines with IC50 values lower than 3.75 μM. We reported a series of Pim-1 small molecule inhibitors that could serve as the lead compounds to develop new targeted anticancer therapeutics.

Synthesis of 2-Oxoquinoline Derivatives as Dual Pim and mTORC Protein Kinase Inhibitors.

Compound VBT-5445 was identified as an inhibitor to block the association of Pim and the protein Enhancer of Decapping 3 (EDC3), a Pim substrate, which normally functions to enhance the decapping of messenger RNA (mRNA). It was also shown to inhibit both the Pim and mTORC protein kinases. The activity of this compound class can be fine-tuned by structural modification. A series of VBT analogs were designed, synthesized, and evaluated. These compounds decrease the growth of multiple cancer types, including pancreas, prostate, breast, lung, and leukemia. Notably, 6-methyl (GRG-1-31, 6d), 4-Bromo (GRG-1-34, 6e), 4-Chloro (GRG-1-35, 6f), and phenylthio substituted (GRG-1-104, 6n) derivatives are highly potent at inhibiting tumor growth. The ability of these compounds to block cancer growth in vitro is highly correlated with their activity as mTORC inhibitors. The toxicity of GRG 1-34 is low in mice treated with twice-daily gavage for 30 days and did not induce weight loss. Pharmacokinetics of a single oral dose demonstrated a peak concentration at 0.5 hours after gavage. In summary, further development of this compound class has the potential to inhibit important signaling pathways and impact cancer treatment.

P602. Do Antipsychotics Suppress Cancer initiation? Identification of Novel off Target Effects of Typical Antipsychotics Using Epidemiology, Bioinformatics, and Protein-Kinase Activity

Schizophrenia is a devastating psychiatric disorder that has a global prevalence of 1%. Interestingly, cancer rates in schizophrenia are lower than the general population. Previous epidemiological studies of cancer rates in schizophrenia did not fully account for confounding variables such as smoking and substance abuse. We sought to confirm lower cancer rates in schizophrenia and explore mechanisms responsible for this unexpected observation. We hypothesize that typical antipsychotics are competitive inhibitors of PIM kinases.

Synthesis of Fused Quinoline Derivatives With Antiproliferative Activities and Tyrosine Kinases, Pim-1 Kinase Inhibitions.

Cyclohexan-1,3-dione (1) reacted with either 2-aminoprop-1-ene-1,1,3-tricarbonitrile (2a) or diethyl 3-amino-2-cyanopent-2-enedioate (2b) to give the 5,6,7,8-tetrahydronaphthalene derivatives 3a and 3b, respectively. The latter compounds underwent further heterocyclization reactions to give the thieno[2',3':5,6]benzo[1,2-e][1,3]oxazine derivatives. On the other hand, the reaction of compound 1 with trichloroacetonitrile afforded the (2,2,2-trichloroethylidene)cyclohexane derivative 14. The latter underwent a series of reactions to produce 2,3,6,7-tetrahydroquinazoline, dihydrothieno[2,3-h]isoquinoline, octahydrobenzo[h]quinazoline and dihydrothieno[2,3-h]isoquinoline derivatives. The synthesized compounds were tested toward six cancer cell lines where most of them gave high inhibitions with c-Met enzymatic activity, with tyrosine kinases and Pim-1 inhibitions. The results obtained will encourage further work through such compounds to produce optimized anticancer agents.

Targeting Host PIM Protein Kinases Reduces Mayaro Virus Replication.

Mayaro virus (MAYV) manipulates cell machinery to successfully replicate. Thus, identifying host proteins implicated in MAYV replication represents an opportunity to discover potential antiviral targets. PIM kinases are enzymes that regulate essential cell functions and also appear to be critical factors in the replication of certain viruses. In this study we explored the consequences of PIM kinase inhibition in the replication of MAYV and other arboviruses. Cytopathic effects or viral titers in samples from MAYV-, Chikungunya-, Una- or Zika-infected cells treated with PIM kinase inhibitors were evaluated using an inverted microscope or plaque-forming assays. The expression of viral proteins E1 and nsP1 in MAYV-infected cells was assessed using an immunofluorescence confocal microscope or Western blot. Our results revealed that PIM kinase inhibition partially prevented MAYV-induced cell damage and also promoted a decrease in viral titers for MAYV, UNAV and ZIKV. The inhibitory effect of PIM kinase blocking was observed for each of the MAYV strains tested and also occurred as late as 8 h post infection (hpi). Finally, PIM kinase inhibition suppressed the expression of MAYV E1 and nsP1 proteins. Taken together, these findings suggest that PIM kinases could represent an antiviral target for MAYV and other arboviruses.

In silico design of novel anticancer drugs with amino acid and carbohydrate building blocks to inhibit PIM kinases

ABSTRACT PIM-1 is a serine-threonine kinase mainly expressed in tissues like the Thymus, spleen, bone marrow, and liver. Overexpression of PIM kinases occurs in various types of human tumours, such as lymphomas, prostate cancer, and oral cancer. As a result, the design of drugs to inhibit PIM-1 in cancerous cells has attracted much attention in recent years. This study aimed to design the alternative inhibitors for PIM-1 kinase, which are based on carbohydrates and amino acids and are expected to be non-toxic with the same chemotherapeutic effects as the traditional known anticancer drugs. The combinatorial use of quantum mechanics (QM) and molecular dynamic simulation (MD) has enabled us to precisely predict the inhibition of PIM-1 kinase by the novel designed drugs and compare them with the recently synthesized chemotherapeutic drugs such as DBC (Table 1). All designed structures were optimized at the B3LYP/6-311++G(d,p) level. The designed structure UNK4 (Table 1) was the most similar structure to DBC in terms of volumes, areas, and electrostatic potential map to DBC. The results obtained in this research represented that UNK4, with the building blocks of amino acid and cyclic carbohydrates could be considered an alternative anticancer drug for DBC.

In silico study for the identification of potential compounds as PIM-1 kinase inhibitors

PIM kinases are a group of serine/threonine kinases that are classified into three isoforms: PIM1, PIM2, and PIM3. Pim-1 kinase is a critical enzyme that is involved in cell growth, cell survival, differentiation, apoptosis, senescence and drug resistance. The PUBMED database has been taken for the screening of PIM-1 kinase inhibitor. This database, further, screened by Lipinski Rule of five, HTVS, standard precision (SP), and extra precision (XP) methodologies. 2OJF protein of PIM-1 kinase was taken for molecular docking. The compound 1a showed good docking scores, SP = −7.244 and XP = −8.6, whereas 1i showed minimal SP and XP scores. These studies may be used for the further development of potential compounds against PIM-1 kinase.

Multi-Component Reactions of Cyclohexan-1,3-dione: Synthesis of Fused Pyran, Pyridine, Thiophene and Pyrazole Derivatives with c-Met, Anti-Proliferative Activities.

Recently, products of Multi-Component Reactions (MCR's) acquired special attention due to their wide range of pharmacological activities especially therapeutic activities. In the market it was found that many pharmacological drugs containing the pyran and pyridine nucleus that were produced through MCR's were found. This work aims to synthesize target molecules not only possess anti-tumour activities but also c-Met and prostate cancer inhibitors. The target molecules were obtained starting from cyclohexan-1,3-dione through its multi-component reactions to produce anticancer target molecules. Cyclohexane-1,3-dione underwent different multi-component reactions to produce fused pyran, pyridine and thiophene derivatives. The anti-proliferative activity of the newly synthesized compounds among the synthesized compounds toward the six cancer cell lines, namely A549, H460, HT-29, MKN-45, U87MG, and SMMC-7721 was studied. In addition, inhibitions toward c-Met kinase and prostate cancer cell line were studied. Antitumor evaluations toward seventeen cancer cell lines subpanel, for certain compounds, were also demonstrated according to the diseases. Pim-1 kinase inhibitions of the most active compounds were also measured. Anti-proliferative evaluations, c-Met and Pim-1 kinase inhibitions were performed for most of the synthesized compounds where the varieties of substituent through the aryl ring and the heterocyclic ring afforded compounds with high activities. Compounds 4b, 6b, 8b, 9a, 11b, 12b, 17b, 18b, 19, 22c, 23b, 25b and 26b were the most cytotoxic compounds toward the six cancer cell lines. Inhibitions toward c-Met kinase and prostate cancer cells showed that the presence of the electronegative Cl group within the molecule were responsible for its high activity. In addition, inhibitions toward Pim-1 kinase exhibited that most of tested compounds showed high inhibitions.

Abstract P223: Phase 1, first-in-human, dose-escalation, safety, pharmacokinetic, and pharmacodynamic study of oral TP-3654, a PIM kinase inhibitor, in patients with advanced solid tumors

Abstract Background: Elevated expression of PIM kinases in various solid and hematologic malignancies is correlated with increased cell survival and reduced apoptosis, implicating inhibition of PIM kinases as attractive targets for disruptive therapy. TP-3654 is an oral, small molecule, investigational PIM kinase inhibitor with favorable selectivity against PIM-1 over other kinases. TP-3654 is currently being evaluated in a phase 1 first-in-human study in patients (pts) with advance solid tumors (NCT03715504). Methods: Pts with advanced metastatic, progressive, or unresectable solid tumors refractory or intolerant of established therapies were treated with TP-3654. Pts took TP-3645 orally once (QD) or twice (BID) daily in a 28-day dosing cycle until unacceptable toxicity or disease progression. Escalating dose levels of TP-3654 were evaluated using a 3+3 design. The primary objective was to determine maximum tolerated dose (MTD) and dose-limiting toxicities (DLTs). Secondary objectives included establishing the pharmacokinetic (PK) profile of oral TP-3654 and any evidence of antitumor activity. An exploratory objective was to study predictive biomarkers in tumor tissues and mechanism-based pharmacodynamic activity in peripheral blood mononuclear cells (PBMCs). Results: As of 31 May 2021, 22 pts were treated with TP-3654 at 480 mg QD (n=3), 720 mg QD (n=3), 1080 mg QD (n=4), 1440 mg QD (n=3), 720 mg BID (n=3), and 960 mg BID (n=6). Median age of 58 years (range: 20–72), 59% (13/22) pts male, and 86% (19/22) pts received ≥3 lines of prior therapy. Median treatment duration is 2 cycles (range: 0.1–24) and 6 pts received treatment for ≥4 cycles. Two pts remain on active study treatment. No DLT was observed. The most common treatment-emergent adverse events (TEAEs) reported in ≥20% of pts were vomiting, diarrhea, nausea, and abdominal pain. No related myelosuppressive AEs were reported. Fifty percent (11/22) of pts reported 18 Grade ≥3 TEAEs, of which only one (alanine aminotransferase increased) was considered related to TP-3654. No related, serious AEs were reported. Two TEAEs resulted in a fatal outcome (acute kidney injury, completed suicide); neither were related to TP-3654. Best response is stable disease (SD) in 67% (10/15) evaluable pts, SD for ≥ 16 weeks in 5 pts, best reduction of tumor size from baseline is -22%. TP-3654 plasma exposure increased with doses up to 1080 mg QD; the BID regimen at the 720 mg dose achieved greater plasma exposure than the 1440 mg QD dosing regimen. Reduced phosphorylation of PIM-1 downstream signal protein pBAD was observed in isolated PBMCs, predominantly in pts with prolonged clinical benefit. Conclusions: Preliminary data suggest that TP-3654 is tolerated as a monotherapy up to 960 mg BID in pts with heavily pretreated, relapsed, and refractory solid tumors. No DLT was observed and MTD was not reached. PK data indicate that the BID dose regimen achieved better plasma exposure than the QD regimen at higher doses. TP-3654 has shown target-specific inhibition of PIM-1 in pts. Updated data will be presented. Citation Format: Ignacio Garrido-Laguna, Patrick M. Dillon, Sujan Kabir, Jian Mei, Mark L. Wade, Huyuan Yang, Carl Stapinski, Jason M. Foulks, Steven L. Warner, Clifford Whatcott, Claudia Lebedinsky, Siqing Fu. Phase 1, first-in-human, dose-escalation, safety, pharmacokinetic, and pharmacodynamic study of oral TP-3654, a PIM kinase inhibitor, in patients with advanced solid tumors [abstract]. In: Proceedings of the AACR-NCI-EORTC Virtual International Conference on Molecular Targets and Cancer Therapeutics; 2021 Oct 7-10. Philadelphia (PA): AACR; Mol Cancer Ther 2021;20(12 Suppl):Abstract nr P223.

Inhibition of PIM Kinases in DLBCL Targets MYC Transcriptional Program and Augments the Efficacy of Anti-CD20 Antibodies.

The family of PIM serine/threonine kinases includes three highly conserved oncogenes, PIM1, PIM2, and PIM3, which regulate multiple prosurvival pathways and cooperate with other oncogenes such as MYC. Recent genomic CRISPR-Cas9 screens further highlighted oncogenic functions of PIMs in diffuse large B-cell lymphoma (DLBCL) cells, justifying the development of small-molecule PIM inhibitors and therapeutic targeting of PIM kinases in lymphomas. However, detailed consequences of PIM inhibition in DLBCL remain undefined. Using chemical and genetic PIM blockade, we comprehensively characterized PIM kinase-associated prosurvival functions in DLBCL and the mechanisms of PIM inhibition-induced toxicity. Treatment of DLBCL cells with SEL24/MEN1703, a pan-PIM inhibitor in clinical development, decreased BAD phosphorylation and cap-dependent protein translation, reduced MCL1 expression, and induced apoptosis. PIM kinases were tightly coexpressed with MYC in diagnostic DLBCL biopsies, and PIM inhibition in cell lines and patient-derived primary lymphoma cells decreased MYC levels as well as expression of multiple MYC-dependent genes, including PLK1. Chemical and genetic PIM inhibition upregulated surface CD20 levels in an MYC-dependent fashion. Consistently, MEN1703 and other clinically available pan-PIM inhibitors synergized with the anti-CD20 monoclonal antibody rituximab in vitro, increasing complement-dependent cytotoxicity and antibody-mediated phagocytosis. Combined treatment with PIM inhibitor and rituximab suppressed tumor growth in lymphoma xenografts more efficiently than either drug alone. Taken together, these results show that targeting PIM in DLBCL exhibits pleiotropic effects that combine direct cytotoxicity with potentiated susceptibility to anti-CD20 antibodies, justifying further clinical development of such combinatorial strategies. SIGNIFICANCE: These findings demonstrate that inhibition of PIM induces DLBCL cell death via MYC-dependent and -independent mechanisms and enhances the therapeutic response to anti-CD20 antibodies by increasing CD20 expression.

Genetic ablation of Pim1 or pharmacologic inhibition with TP-3654 ameliorates myelofibrosis in murine models.

Myelofibrosis (MF) is the deadliest form of myeloproliferative neoplasm (MPN). The JAK inhibitor Ruxolitinib can reduce constitutional symptoms but it does not substantially improve bone marrow fibrosis. Pim1 expression is significantly elevated in MPN/MF hematopoietic progenitors. Here, we show that genetic ablation of Pim1 blocked the development of myelofibrosis induced by Jak2V617F and MPLW515L. Pharmacologic inhibition of Pim1 with a second-generation Pim kinase inhibitor TP-3654 significantly reduced leukocytosis and splenomegaly, and attenuated bone marrow fibrosis in Jak2V617F and MPLW515L mouse models of MF. Combined treatment of TP-3654 and Ruxolitinib resulted in greater reduction of spleen size, normalization of blood leukocyte counts and abrogation of bone marrow fibrosis in murine models of MF. TP-3654 treatment also preferentially inhibited Jak2V617F mutant hematopoietic progenitors in mice. Mechanistically, we show that TP-3654 treatment significantly inhibits mTORC1, MYC and TGF-β signaling in Jak2V617F mutant hematopoietic cells and diminishes the expression of fibrotic markers in the bone marrow. Collectively, our results suggest that Pim1 plays an important role in the pathogenesis of MF, and inhibition of Pim1 with TP-3654 might be useful for treatment of MF.