CTX-712, a Novel Clk Inhibitor Targeting Myeloid Neoplasms with SRSF2 Mutation

Abstract Splicing factors (SFs) are among the most frequent mutational targets in myeloid neoplasms, particularly in myelodysplastic syndromes (MDS) and a subset of acute myeloid leukemia (AML), designated as chromatin/spliceosome-mutated AML, where major SFs mutated include SF3B1, SRSF2, U2AF1, and ZRSR2. SF mutations are largely mutually exclusive and except for ZRSR2 mutations, heterozygous, suggesting synthetic lethality of homozygous or multiple mutations. Thus, SF functions might be a plausible target of therapy for MDS/AML. Of potential interest in this regard is serine/arginine-rich (SR) domains ubiquitously shared by many SFs, including U2AF1, SRSF2, and ZRSR2, which need to be phosphorylated for their nuclear translocation by evolutionally conserved kinases, known as CLK family of proteins. CLK family kinases regulate mRNA splicing by phosphorylating various SR proteins, and inhibition of CLK family kinases resulted in reduction of phosphorylation levels of SR proteins, induction of splicing alterations and protein depletion for multiple genes. In addition, a recent report showed that CLK inhibition can induce skipped exons, cell death, and cell growth suppression. Thus, CLK inhibitors might have a role in the therapeutics of SF-mutated MDS/AML, by further compromising RNA splicing. We have recently developed an orally available and highly potent CLK inhibitor, CTX-712, and evaluated its anti-leukemic activities both in vitro and in vivo. When tested in human myeloid cell lines (K562 and MV-4-11), CTX-712 showed a strong inhibitory effect on cell proliferation (IC 50=0.15 and 0.036 μM, respectively). The anti-leukemic effect was also confirmed by survival assay using a total of 79 primary AML cells (the average of IC 50 was 0.078 μM). In addition, CTX-712 suppressed phosphorylation of multiple SR proteins including SRSF3/4/6, all of which bind to SRSF2. RNA-seq analysis revealed that CTX-712 induced global splicing changes, which typically resulted in skipped exon. Notably, the degree of splicing (percent spliced-in value) in skipped exon events induced by the drug was positively correlated with the sensitivity to the drug (IC 50) in primary AML cells (n=32, R=0.61, P=0.00018). To further investigate the effect of CTX-712 on tumor growth in vivo, we established 13 MDS/AML-derived xenografts (PDXs), which were treated with varying doses of CTX-712. Among the 13 PDX models, SRSF2 mutation was found in 2 cases, which had the SRSF2 P95H or P95L mutation. The SRSF2 P95H PDX showed a significant response to CTX-712 in a dose-dependent manner. Of note, 4 out of 5 mice treated using a high dose protocol (12.5 mg/kg) achieved complete remission (the tumor shrank completely to unmeasurable size). Two weeks after treatment, tumor volumes (mm 3) were 762 ± 147 (vehicle), 331 ± 64 (low dose of CTX-712: 6.25mg/kg, P=0.0395), and 39 ± 39 (high dose, P=0.0064) (N=5 each, mean ± SEM). Interestingly CLK inhibition induced aberrant splicing events including skipped exons in vivo, which were more strongly affected in the SRSF2-mutated model. In addition, CTX-712 efficacy was also confirmed in the model with the SRSF2 P95L mutation. The SRSF2 P95L model showed a significant reduction in tumor volumes (mm 3) 2 weeks after CTX-712 treatment; 406 ± 94 (vehicle) and 75 ± 17 (high dose, P=0.0162) (N=6 each, mean ± SEM). CTX-712 also significantly improved the survival of the SRSF2 P95L-mutated model (high dose, P=0.0030) (N=6 each). Overall, 10 out of 13 PDX AML/MDS models, including 2 SRSF2-mutated models, showed anti-tumor effect of CTX-712. Complete disappearances of tumors were obtained in the SRSF2 P95H mutation model. These results provide mechanistic insights of CLK inhibition and a rationale for further investigation of the novel CLK inhibitor in MDS/AML. Disclosures Yoda: Chordia Therapeutics Inc.: Research Funding. Morishita: Chordia Therapeutics Inc.: Current Employment, Current equity holder in publicly-traded company. Mizutani: Chordia Therapeutics Inc.: Current Employment, Current equity holder in publicly-traded company. Tozaki: Chordia Therapeutics Inc.: Current Employment, Current equity holder in publicly-traded company. Satoh: Chordia Therapeutics Inc.: Current Employment, Current equity holder in publicly-traded company. Nannya: Otsuka Pharmaceutical Co., Ltd.: Consultancy, Speakers Bureau; Astellas: Speakers Bureau. Miyake: Chordia Therapeutics Inc.: Current Employment, Current equity holder in publicly-traded company. Ogawa: Kan Research Laboratory, Inc.: Consultancy, Research Funding; Dainippon-Sumitomo Pharmaceutical, Inc.: Research Funding; Eisai Co., Ltd.: Research Funding; Otsuka Pharmaceutical Co., Ltd.: Research Funding; ChordiaTherapeutics, Inc.: Consultancy, Research Funding; Ashahi Genomics: Current holder of individual stocks in a privately-held company.

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CTX-712, a Novel Clk Inhibitor Targeting Myeloid Neoplasms with SRSF2 Mutation

Blood ◽

10.1182/blood-2019-131559 ◽

2019 ◽

Vol 134 (Supplement_1) ◽

pp. 404-404

Author(s):

Akinori Yoda ◽

Daisuke Morishita ◽

Akio Mizutani ◽

Yoshihiko Satoh ◽

Yotaro Ochi ◽

...

Keyword(s):

Low Dose ◽

Synthetic Lethality ◽

Sr Proteins ◽

High Dose ◽

Dependent Manner ◽

Employment Equity ◽

Myeloid Neoplasms ◽

Equity Ownership ◽

Dose Dependent

Splicing factors (SFs) are among the most frequent mutational targets in myeloid neoplasms, particularly in myelodysplastic syndromes (MDS) and a subset of acute myeloid leukemia (AML), designated as 'chromatin/spliceosome-mutated AML, where major SFs mutated include SF3B1, SRSF2, U2AF1, and ZRSR2. These SF mutations are largely mutually exclusive and except for ZRSR2 mutations, are invariably heterozygous, showing prominent hotspots, suggesting that mutations have neomorphic functions and might cause a synthetic lethality when they are homozygous or two SFs are mutated. Thus, SF functions might be a plausible target of therapy for MDS/AML. Of potential interest in this regard is serine/arginine-rich (SR) domains ubiquitously shared by many SFs, including U2AF1, SRSF2, and ZRSR2, which need to be phosphorylated for their nuclear translocation by evolutionally conserved kinases, known as CLK family of proteins. CLK family kinases regulate mRNA splicing by phosphorylating various SR proteins, and inhibition of CLK family kinases resulted in reduction of phosphorylation levels of SR proteins, induction of splicing alterations and protein depletion for multiple genes, including those involved in growth and survival pathways such as S6K, EGFR, EIF3D, and PARP. In addition, a recent report showed that CLK inhibition can induce skipped exons, cell death, and cell growth suppression, which are dependent of CLK2 expression levels. Thus, CLK family kinases are possible targets of inhibition by small molecules to induce synthetic lethality in SF-mutated MDS/AML and for this purpose, we have recently developed an orally available and highly potent CLK inhibitor, CTX-712 and evaluated its anti-leukemic activities both in vitro and in vivo. When tested in human myeloid cell lines (K562 and THP1), CTX-712, strongly inhibited phosphorylation of multiple SR proteins including SRSF3, SRSF4, SRSF5, and SRSF6 that bind to SRSF2. To further investigate the efficacy of CTX-712 in vivo, we established 5 AML-derived xenograft (PDX) models, which treated with varying doses of CTX-712. Among these 5 PDX models, SRSF2 mutation was found in only one case, which had a SRSF2 p.P95H, mutation, while others (a subcutaneous and 3 leukemia model) were negative for SRSF2 mutations. The SRSF2-mutated model showed a significant response to CTX-712 in a dose-dependent manner. Of note, 4 out of 5 mice treated using a high dose protocol (12.5 mg/kg) achieved complete remission (the tumor shrank completely to unmeasurable size). Two-week after treatment, tumor volumes (mm3) were 762 ± 147 (vehicle), 331 ± 64 (low dose of CTX-712: 6.25mg/kg, P=0.028), and 39 ± 39 (high dose, P=0.0014) (N=5 each, mean ± SEM). CTX-712 also significantly improved the survival of PDX #1. Median survivals (days after engraftment) were 34.5 (vehicle) vs. 93.5 (12.5mg/kg, P=0.015) (N=2 each). Interestingly, another leukemic model carrying KRAS, NF1, and TP53 but not SRSF2 mutations also showed a significant reduction of leukemic burden 2 weeks after CTX-712 treatment; leukemic burden after therapy, as measured by frequency of hCD45+cells in PB (%), were 82 ± 2.2 (vehicle), 17 ± 3.6 (low dose, P<0.0001), and 0.89 ± 0.43 (complete remission, high dose, P<0.0001), (N=4 each, mean ± SEM). In the third PDX model with mutations of FLT3, RAD21, RUNX1, and WT1, CTX-712 administration reduced subcutaneous AML tumors in a dose-dependent manner and achieved partial remission (high dose, P=0.0008) (N=6 each). CTX-712 also significantly improved the survival of the PDX #3 model (high dose, P=0.0069) (N=6 each). In PDX #4, leukemic model with mutations of ASXL1, BCOR, and TET2, high dose CTX-712 therapy strongly reduced the leukemic cell burden than vehicle control (P=0.0027), (N=4 each). CTX-712 also significantly improved the survival of this model (P=0.016) (N=5 each). The last AML PDX #5 model with U2AF1, BCOR, DNMT3A, IDH1, KDM6A, RUNX1, and TET2 mutations was refractory for CTX-712 therapy. Overall, 4 out of 5 PDX AML models showed anti-tumor effect of CTX-712. Complete disappearances of tumors were obtained in 2 cases, including an SRSF2-mutated model. These results provide mechanistic insights of CLK inhibition and a rationale for further investigation of the novel CLK inhibitor in MDS/AML. CTX-712 is currently in clinical phase 1 trials for relapsed and refractory malignancies. Disclosures Yoda: Chordia Therapeutics Inc.: Research Funding. Morishita:Chordia Therapeutics Inc.: Employment, Equity Ownership. Mizutani:Chordia Therapeutics Inc.: Employment, Equity Ownership. Satoh:Chordia Therapeutics Inc: Employment, Equity Ownership. Miyake:Chordia Therapeutics Inc.: Employment, Equity Ownership. Ogawa:Dainippon-Sumitomo Pharmaceutical, Inc.: Research Funding; ChordiaTherapeutics, Inc.: Consultancy, Equity Ownership; Kan Research Laboratory, Inc.: Consultancy; RegCell Corporation: Equity Ownership; Asahi Genomics: Equity Ownership; Qiagen Corporation: Patents & Royalties.

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Selective Targeting of Splicing Factor Mutant Hematopoietic Stem and Progenitor Cells Via STAT3 Inhibition

Blood ◽

10.1182/blood-2021-151338 ◽

2021 ◽

Vol 138 (Supplement 1) ◽

pp. 1509-1509

Author(s):

Kathryn S Potts ◽

Rosannah C. Cameron ◽

Noura Ghazale ◽

Varun Gupta ◽

Juan Martin Barajas ◽

...

Keyword(s):

Research Funding ◽

Synthetic Lethality ◽

K562 Cells ◽

Splicing Factor ◽

Publicly Traded ◽

Hematopoietic Stem ◽

Stem And Progenitor Cells ◽

Mutant Cells ◽

Stat3 Inhibition

Abstract Myelodysplastic syndrome (MDS) is a bone marrow failure disorder driven by dysfunction of hematopoietic stem and progenitor cells (HSPCs). Patient sequencing studies over the last decade have revealed that mutations in splicing machinery predominate in MDS, thus selective targeting of these cells is therapeutically attractive. STAT3 inhibition has been explored previously as a means to eradicate HSPCs in MDS. While efficacy was demonstrated in a subset of samples, the underlying mechanism for this selectivity remains unknown. We examined RNAseq of MDS CD34+ HSPCs with splicing factor mutations versus wildtype, finding alternative splicing and differential expression of STAT3 pathway components. Functionally, we explored if STAT3 signaling represents a novel vulnerability in SF3B1 mutant HSPCs using a multi-model approach of in vivo zebrafish and mouse systems, and in vitro assays of CRISPR-engineered human leukemia K562 cells and primary MDS samples. Utilizing the small molecule STAT3 inhibitor STATTIC, we found that human cells carrying MDS-associated SF3B1 point mutations had heightened sensitivity to STAT3 inhibition compared to wildtype controls. To evaluate the activity of STAT3 inhibition in vivo, we utilized an Mx1-cre conditional knock-in mouse model of mutant SF3B1 (Sf3b1+/K700E). We demonstrated that in vivo STATTIC treatment selectively depleted Sf3b1 mutant cells over wildtype in vivo. RNAseq of sf3b1 homozygous mutantzebrafish cells revealed conserved dysregulation of STAT3 pathway splicing and target expression. Diminishing Stat3 (via morpholino knockdown, stable mutants, or STATTIC treatment) decreased HSPCs in sf3b1 heterozygotes but not wildtype embryos, demonstrating synthetic lethality between Sf3b1 and Stat3. Our data indicate that SF3B1 heterozygosity, regardless of the type of mutation, confers a heightened sensitivity to STAT3 inhibition in zebrafish, mouse, and human HSPCs. Critically, our data indicate that SF3B1-mutant cells can be selectively killed in vivo while sparing wildtype cells. We sought to rescue HSPCs in sf3b1 homozygous mutant zebrafish, however overexpression of ligands Osm and Il6 or wildtype Stat3 was insufficient. Instead, overexpression of constitutively-active Stat3 partially restored HSPCs, indicating that functional Stat3 signaling downstream of Sf3b1 is critical for HSPC formation. To investigate the specificity of the synthetic lethality for SF3B1, we assessed the STAT3 synthetic lethal interaction with other mutated splicing factors in MDS. Similar to SF3B1, we demonstrated STAT3 synthetic lethality with U2AF1 and SRSF2 heterozygosity in zebrafish and human cells. RNA-sequencing analysis of STATTIC-treated K562 cells revealed an exacerbation of splicing alterations upon STAT3 inhibition that was more pronounced in SF3B1+/K666N cells compared to wildtype. Even more strikingly, we demonstrated that constitutive activation of STAT3 could partially reverse defective splicing in zebrafish sf3b1 homozygous mutant cells. Mechanistically, these data strongly support coordinated splicing dysfunction as the underlying cause for STAT3-SF3B1 synthetic lethality. Together, we demonstrated a conserved and selective synthetic lethal interaction between STAT3 function and splicing factor defects that represents a novel liability for mutant HSPCs with important implications for MDS treatment. Disclosures Shastri: Guidepoint: Consultancy; Kymera Therapeutics: Research Funding; Onclive: Honoraria; GLC: Consultancy. Verma: Curis: Research Funding; BMS: Research Funding; Stelexis: Consultancy, Current equity holder in publicly-traded company; Eli Lilly: Research Funding; Medpacto: Research Funding; Incyte: Research Funding; GSK: Research Funding; Novartis: Consultancy; Acceleron: Consultancy; Celgene: Consultancy; Stelexis: Current equity holder in publicly-traded company; Throws Exception: Current equity holder in publicly-traded company.

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Phaseolin Attenuates Lipopolysaccharide-Induced Inflammation in RAW 264.7 Cells and Zebrafish

Biomedicines ◽

10.3390/biomedicines9040420 ◽

2021 ◽

Vol 9 (4) ◽

pp. 420

Author(s):

Su-Jung Hwang ◽

Ye-Seul Song ◽

Hyo-Jong Lee

Keyword(s):

Nitric Oxide ◽

Nuclear Translocation ◽

Raw 264.7 Cells ◽

Network Pharmacology ◽

Raw 264.7 ◽

Dependent Manner ◽

Bioactive Constituents

Kushen (Radix Sophorae flavescentis) is used to treat ulcerative colitis, tumors, and pruritus. Recently, phaseolin, formononetin, matrine, luteolin, and quercetin, through a network pharmacology approach, were tentatively identified as five bioactive constituents responsible for the anti-inflammatory effects of S. flavescentis. However, the role of phaseolin (one of the primary components of S. flavescentis) in the direct regulation of inflammation and inflammatory processes is not well known. In this study, the beneficial role of phaseolin against inflammation was explored in lipopolysaccharide (LPS)-induced inflammation models of RAW 264.7 macrophages and zebrafish larvae. Phaseolin inhibited LPS-mediated production of nitric oxide (NO) and the expression of inducible nitric oxide synthase (iNOS), without affecting cell viability. In addition, phaseolin suppressed pro-inflammatory mediators such as cyclooxygenase 2 (COX-2), interleukin-1β (IL-1β), tumor necrosis factor α (TNF-α), monocyte chemoattractant protein-1 (MCP-1), and interleukin-6 (IL-6) in a dose-dependent manner. Furthermore, phaseolin reduced matrix metalloproteinase (MMP) activity as well as macrophage adhesion in vitro and the recruitment of leukocytes in vivo by downregulating Ninjurin 1 (Ninj1), an adhesion molecule. Finally, phaseolin inhibited the nuclear translocation of nuclear factor-kappa B (NF-κB). In view of the above, our results suggest that phaseolin could be a potential therapeutic candidate for the management of inflammation.

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Effect of Synchronous Versus Sequential Regimens on the Pharmacokinetics and Biodistribution of Regorafenib with Irradiation

Pharmaceutics ◽

10.3390/pharmaceutics13030386 ◽

2021 ◽

Vol 13 (3) ◽

pp. 386

Author(s):

Tung-Hu Tsai ◽

Yu-Jen Chen ◽

Li-Ying Wang ◽

Chen-Hsi Hsieh

Keyword(s):

Stereotactic Body Radiation Therapy ◽

In Vivo Studies ◽

Control Group ◽

High Dose ◽

Dependent Manner ◽

Hep G2 ◽

Time Curve ◽

Dose Dependent

This study was performed to evaluate the interaction between conventional or high-dose radiotherapy (RT) and the pharmacokinetics (PK) of regorafenib in concurrent or sequential regimens for the treatment of hepatocellular carcinoma. Concurrent and sequential in vitro and in vivo studies of irradiation and regorafenib were designed. The interactions of RT and regorafenib in vitro were examined in the human hepatoma Huh-7, HA22T and Hep G2 cell lines. The RT–PK phenomenon and biodistribution of regorafenib under RT were confirmed in a free-moving rat model. Regorafenib inhibited the viability of Huh-7 cells in a dose-dependent manner. Apoptosis in Huh-7 cells was enhanced by RT followed by regorafenib treatment. In the concurrent regimen, RT decreased the area under the concentration versus time curve (AUC)regorafenib by 74% (p = 0.001) in the RT2 Gy × 3 fraction (f’x) group and by 69% (p = 0.001) in the RT9 Gy × 3 f’x group. The AUCregorafenib was increased by 182.8% (p = 0.011) in the sequential RT2Gy × 1 f’x group and by 213.2% (p = 0.016) in the sequential RT9Gy × 1 f’x group. Both concurrent regimens, RT2Gy × 3 f’x and RT9Gy × 3 f’x, clearly decreased the biodistribution of regorafenib in the heart, liver, lung, spleen and kidneys, compared to the control (regorafenib × 3 d) group. The concurrent regimens, both RT2Gy × 3 f’x and RT9Gy × 3 f’x, significantly decreased the biodistribution of regorafenib, compared with the control group. The PK of regorafenib can be modulated both by off-target irradiation and stereotactic body radiation therapy (SBRT).

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Therapy-Related Myeloid Neoplasm Has a Distinct Pro-Inflammatory Bone Marrow Microenvironment and Delayed DNA Damage Repair

Blood ◽

10.1182/blood-2020-137277 ◽

2020 ◽

Vol 136 (Supplement 1) ◽

pp. 37-38

Author(s):

Monika M Kutyna ◽

Li Yan A Wee ◽

Sharon Paton ◽

Dimitrios Cakouros ◽

Agnieszka Arthur ◽

...

Keyword(s):

Dna Damage ◽

Board Of Directors ◽

Research Funding ◽

High Dose ◽

Cytotoxic Therapy ◽

Advisory Committees ◽

Myeloid Neoplasm ◽

Myeloid Neoplasms ◽

Damage Repair

Introduction: Therapy-related myeloid neoplasms (t-MN) are associated with extremely poor clinical outcomes in otherwise long-term cancer survivors. t-MN accounts for ~20% of cases of myeloid neoplasms and is expected to rise due to the increased use of chemotherapy/radiotherapy (CT/RT) and improved cancer survivorship. Historically, t-MN was considered a direct consequence of DNA damage induced in normal hematopoietic stem cells (HSC) by DNA damaging cytotoxics. However, these studies have largely ignored the bone marrow (BM) microenvironment and the effects of age and concurrent/previous cancers. Aim: We performed an exhaustive functional study of mesenchymal stromal cells (MSC) obtained from a comparatively large cohort of t-MN patients and carefully selected control populations to evaluate the long-term damage induced by cytotoxic therapy to BM microenvironment and its impact on malignant and normal haematopoiesis. Methods: Four different cohorts were used: (1) t-MN, in which myeloid malignancy occurred after CT/RT for a previous cancer (n=18); (2) patients with multiple cancer and in which a myeloid neoplasm developed following an independent cancer which was not treated with CT/RT (MC-MN; n=10); (3) primary MN (p-MN; n=7) untreated and without any prior cancer or CT/RT; (4) age-matched controls (HC; n=17). Morphology, proliferation, cellular senescence, differentiation potential and γH2AX DNA damage response was performed. Stem/progenitor supportive capacity was assessed by co-culturing haematopoietic stem cells on MSC feeder-layer in long-term culture initiating assay (LTC-IC). Cytokine measurements were performed using 38-plex magnetic bead panel (Millipore) and RNA sequencing libraries were prepared with Illumina TruSeq Total RNA protocol for 150bp paired-end sequencing on a NextSeq500 instrument. Functional enrichment analysis was performed using EnrichR software. Results: MSC cultured from t-MN patients were significantly different from HC, p-MN and MC-MN MSC according to multiple parameters. They exhibited aberrant morphology consisting of large, rounded and less adhesive cells compared to typical spindle-shaped morphology observed with controls. MSC from myeloid neoplasm also showed impaired proliferation, senescence, osteo- and adipogenic differentiation with t-MN MSC showing the greatest differences. DNA repair was dramatically impaired compared to p-MN and HC (Fig.1A). Importantly, these aberrant t-MN MSC were not able to support normal or autologous in vitro long-term haematopoiesis (Fig.1B). The biological characteristic and poor haematopoietic supportive capacity of MSC could be "cell-intrinsic" or driven by an altered paracrine inflammatory microenvironment. Interestingly, several inflammatory cytokines were higher in t-MN compared with marrow interstitial fluid obtained from p-MN patients (Fig.1Ci) and many of these including Fractalkine, IFNα2, IL-7 and G-CSF were also significantly higher in t-MN MSC conditional media (Fig.1Cii). Together, this data suggest that t-MN microenvironment is distinct from p-MN with paracrine production of pro-inflammatory milieu that may contribute to poor HSC supportive capacity. Preliminary whole transcriptome analysis revealed differential gene expression between t-MN and HC (Fig.1Di) and p-MN MSC. Importantly, the deregulated genes play critical role in cell cycle, DNA damage repair, and cellular senescence pathways explaining phenotypical characteristic of t-MN MSC (Fig.1Dii). Moreover CXCL12 expression, a key regulator of haematopoiesis, was significantly lower in t-MN compared to HC (p=0.002) and p-MN MSC (p=0.009), thus explaining poor HSC supportive capacity. The key difference between the p-MN, MC-MN and t-MN is prior exposure to CT/RT. To study this we obtained MSC from two t-MN patients for whom we had samples at the time of their primary cancer, post high-dose chemotherapy and at the time of t-MN. MSC displayed aberrant proliferation and differentiation capacity after high-dose cytotoxic therapy (2 to 4 years prior to developing t-MN) and remained aberrant at t-MN diagnosis (Fig.1E). Conclusions: BM-MSC from t-MN patients are significantly abnormal compared with age-matched controls and typical myeloid neoplasm. Importantly, prior CT/RT leads to long-term irreversible damage to the BM microenvironment which potentially contributes to t-MN pathogenesis. Disclosures Hughes: Novartis: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; BMS: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding. Hiwase:Novartis Australia: Research Funding.

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Targeting Antibody Checkpoint Fcgriib Using Monoclonal Antibody BI-1206 to Overcome Therapeutic Resistance in Mantle Cell Lymphoma

Blood ◽

10.1182/blood-2020-140849 ◽

2020 ◽

Vol 136 (Supplement 1) ◽

pp. 35-35

Author(s):

Alexa A Jordan ◽

Joseph McIntosh ◽

Yang Liu ◽

Angela Leeming ◽

William Lee ◽

...

Keyword(s):

Monoclonal Antibody ◽

Tumor Growth ◽

Research Funding ◽

Cell Lymphoma ◽

Single Agent ◽

Publicly Traded ◽

In Vivo Efficacy ◽

Mantle Cell ◽

Xenograft Models

Mantle cell lymphoma (MCL) is a rare but aggressive B-cell non-Hodgkin's lymphoma that represents 6% of all lymphomas in the United States. Recent therapies including anti-CD20 antibody rituximab, BTK inhibitors, and BCL-2 inhibitors alone or in combination have shown great anti-MCL efficacy. However, primary and acquired resistance to one or multiple therapies commonly occurs, resulting in poor clinical outcome. Therefore, resistance to such therapies is currently an unmet clinical challenge in MCL patients. Therapeutic strategies to overcome this resistance holds promise to significantly improve survival of refractory/relapsed MCL patients. Recent studies showed Fc gamma receptors (FcγRs) play important roles in enhancing the efficacy of antibody-based immunotherapy. In particular, FcgRIIB (CD32B), an inhibitory member of the FcγR family, is implicated in the immune cell desensitization and tumor cell resistance through the internalization of therapeutic antibodies such as rituximab. Based on our flow cytometry analysis, we demonstrated that FcgRIIB is highly expressed on the cell surface of MCL cell lines (n=10) and primary MCL patient samples (n=22). This indicates that FcgRIIB may play an important role in MCL malignancy and identifies FcgRIIB is a potential therapeutic target for the treatment of MCL. To address this, we tested the in vivo efficacy of BI-1206, a fully humanized monoclonal antibody targeting FcgRIIB, alone, or in combination with clinically approved or investigational drugs including rituximab, ibrutinib and venetoclax. In the first in vivo cohort, BI-1206, as a single agent, dramatically inhibited the tumor growth of ibrutinib-venetoclax dual-resistant PDX tumor models, suggesting that targeting FcgRIIB by BI-1206 alone has high anti-MCL activity in vivo. Next, we assessed whether BI-1206 can boost anti-MCL activity of antibody-based therapy such as rituximab in combination with ibrutinib or venetoclax using additional mice cohorts of cell line-derived xenograft and patient-derived xenograft models. BI-1206 significantly enhanced the in vivo efficacy of ibrutinib plus rituximab, and venetoclax plus rituximab, on tumor growth inhibition, including the JeKo-1 derived xenograft models, previously proven to be partially resistant to ibrutinib and venetoclax in vivo. This tumor-sensitizaton effect was further confirmed in the ibrutinib and venetoclax dual-resistant PDX models of MCL where BI-1206 was combined with venetoclax and rituximab. More detailed mechanistic studies are currently ongoing to reveal the mechanism of action of BI-1206-based combinations or as single therapy with the possibility that BI-1206 itself may have a cytotoxic anti-tumor direct activity in MCL. In conclusion, BI-1206 as single agent showed potent efficacy in overcoming ibrutnib-venetoclax dual resistance. Moveover, BI-1206 enhanced the in vivo efficacy of ibrutinib plus rituximab and venetoclax plus rituximab and overcomes resistance to these treatments resulting in enhanced anti-tumor effects. Disclosures Karlsson: BioInvent International AB: Current Employment. Mårtensson:BioInvent International AB: Current Employment, Current equity holder in publicly-traded company. Kovacek:BioInvent International AB: Current Employment, Current equity holder in publicly-traded company. Teige:BioInvent International AB: Current Employment, Current equity holder in publicly-traded company. Frendéus:BioInvent International AB: Current Employment, Current equity holder in publicly-traded company. Wang:Pulse Biosciences: Consultancy; Loxo Oncology: Consultancy, Research Funding; Kite Pharma: Consultancy, Other: Travel, accommodation, expenses, Research Funding; BioInvent: Research Funding; Juno: Consultancy, Research Funding; Beijing Medical Award Foundation: Honoraria; OncLive: Honoraria; Verastem: Research Funding; Molecular Templates: Research Funding; Dava Oncology: Honoraria; Guidepoint Global: Consultancy; Nobel Insights: Consultancy; Oncternal: Consultancy, Research Funding; InnoCare: Consultancy; Acerta Pharma: Research Funding; VelosBio: Research Funding; MoreHealth: Consultancy; Targeted Oncology: Honoraria; OMI: Honoraria, Other: Travel, accommodation, expenses; Celgene: Consultancy, Other: Travel, accommodation, expenses, Research Funding; AstraZeneca: Consultancy, Honoraria, Other: Travel, accommodation, expenses, Research Funding; Pharmacyclics: Consultancy, Honoraria, Other: Travel, accommodation, expenses, Research Funding; Janssen: Consultancy, Honoraria, Other: Travel, accommodation, expenses, Research Funding; Lu Daopei Medical Group: Honoraria.

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Metformin Suppressed CXCL8 Expression and Cell Migration in HEK293/TLR4 Cell Line

Mediators of Inflammation ◽

10.1155/2017/6589423 ◽

2017 ◽

Vol 2017 ◽

pp. 1-11 ◽

Cited By ~ 4

Author(s):

Zhihui Xiao ◽

Wenjun Wu ◽

Vladimir Poltoratsky

Keyword(s):

Cell Proliferation ◽

Cell Migration ◽

Tumor Cell ◽

Nuclear Translocation ◽

High Dose ◽

Dependent Manner ◽

Tumor Cell Proliferation ◽

Cell Proliferation And Migration ◽

And Migration ◽

Proliferation And Migration

Chronic inflammation is associated with cancer. CXCL8 promotes tumor microenvironment construction through recruiting leukocytes and endothelial progenitor cells that are involved in angiogenesis. It also enhances tumor cell proliferation and migration. Metformin, type II diabetes medication, demonstrates anticancer properties via suppressing inflammation, tumor cell proliferation, angiogenesis, and metastasis. This study intended to address the role of metformin in regulation of CXCL8 expression and cell proliferation and migration. Our data indicated that metformin suppressed LPS-induced CXCL8 expression in a dose-dependent manner through inhibiting NF-κB, but not AP-1 and C/EBP, activities under the conditions we used. This inhibitory effect of metformin is achieved through dampening LPS-induced NF-κB nuclear translocation. Cell migration was inhibited by metformin under high dose (10 mM), but not cell proliferation.

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Suicide inactivation of aromatase in human placenta and uterine leiomyoma by 5α-dihydronorethindrone, a metabolite of norethindrone, and its effect on steroid-producing enzymes

Acta Endocrinologica ◽

10.1530/eje.0.1300634 ◽

1994 ◽

Vol 130 (6) ◽

pp. 634-640 ◽

Cited By ~ 5

Author(s):

Takara Yamamoto ◽

Takaya Tamura ◽

Jo Kitawaki ◽

Yoshio Osawa ◽

Hiroji Okada

Keyword(s):

Human Placenta ◽

Uterine Leiomyoma ◽

High Dose ◽

Aromatase Activity ◽

Dependent Manner ◽

Chain Cleavage ◽

Time Period ◽

Suicide Inactivation

Yamamoto T, Tamura T, Kitawaki J, Osawa Y, Okada H. Suicide inactivation of aromatase in human placenta and uterine leiomyoma by 5α-dihydronorethindrone, a metabolite of norethindrone, and its effect on steroid-producing enzymes. Eur J Endocrinol 1994;130:634–40. ISSN 0804–4643 Norethindrone (NET; 17α-ethynyl-19-nortestosterone), a progestogen component of the contraceptive pill, irreversibly inhibits aromatase activity in human placental microsomes. However, it is known also to be aromatized in vitro and in vivo to produce a biologically very active estrogen called ethynylestradiol (EE2). It is therefore inappropriate to administer a high dose of NET to estrogendependent cancer patients for a prolonged time period. In this study, we focused on 5α-dihydronorethindrone (5α-DHNET), a metabolite of NET that is not aromatizable, and the inhibitory effects of 5α-DHNET on human placental and uterine leiomyoma microsomal aromatase and other steroid synthetases. 5α-Dihydronorethindrone showed weak affinity for both estrogen and progestogen receptors. It inhibited significantly human placental aromatase activity in a dose-dependent manner (Ki = 9.0 μmol/l; Kinact = 0.024/min), as well as that of uterine leiomyoma, but did not influence cholesterol side-chain cleavage or 17α-hydroxylase, 21-hydroxylase or 11β-hydroxylase activities. These results suggest that 5α-DHNET may be useful as an aromatase inhibitor, whose use in large doses is expected to reduce the size of estrogen-dependent tumors. Takara Yamamoto, Department of Obstetrics and Gynecology, Kawaramachi-Hirokoji, Kamikyo-Ku, Kyoto 602, Japan

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The New CXCR4 Inhibitor MDX-1338 Exerts Anti-Tumor Activity in Multiple Myeloma

Blood ◽

10.1182/blood.v118.21.1844.1844 ◽

2011 ◽

Vol 118 (21) ◽

pp. 1844-1844 ◽

Cited By ~ 1

Author(s):

Aldo M Roccaro ◽

Antonio Sacco ◽

Michelle Kuhne ◽

AbdelKareem Azab ◽

Patricia Maiso ◽

...

Keyword(s):

Multiple Myeloma ◽

Bone Marrow ◽

Tumor Cells ◽

Mouse Models ◽

Research Funding ◽

Therapeutic Agents ◽

Dependent Manner ◽

Apoptotic Pathways ◽

Dose Dependent

Abstract Abstract 1844 Background. We have previously shown the SDF1/CXCR4 axis plays a major role in homing and trafficking of multiple myeloma (MM) to the bone marrow (BM), and disruption of the interaction of tumor cells with the BM leads to enhanced sensitivity to therapeutic agents. We hypothesize that the novel anti-CXCR4 antibody, BMS936564/MDX-1338, may prevent the homing and adhesion of MM cells to the BM and will sensitize them to therapeutic agents. Methods. Primary MM cells (CD138+); MM cell lines (MM.1S, RPMI.8226); and primary MM bone marrow stromal cells (BMSCs) were used. Migration towards SDF-1 and BMSCs has been evaluated. Cytotoxicity and DNA synthesis were measured by MTT and thymidine uptake, respectively. Cell signaling and apoptotic pathways were studied by Western Blot. Synergism was calculated using the Chou-Talalay method. In vivo MM tumor growth was evaluated with xenograft mouse models. Results. MDX-1338 inhibited migration of MM cells toward SDF-1a and primary MM BMSCs, in a dose-dependent manner. Adhesion of primary MM cells to BMSCs was also inhibited by BMS936564/MDX-1338 in a dose-dependent manner, while also inducing cytotoxicity on primary BM-derived CD138+ cells. BMS936564/MDX-1338 targeted MM cells in the context of BM milieu by overcoming BMSC-induced proliferation of tumor cells. In addition, BMS936564/MDX-1338 synergistically enhanced bortezomib-induced cytotoxicity in MM cells. BMS936564/MDX-1338-dependent activation of apoptotic pathways in MM cells was documented, as shown by cleavage of caspase-9 and PARP. SDF-1a-induced ERK-, Akt-, and Src-phosphorilation was inhibited by BMS936564/MDX-1338 in a dose-dependent manner. Importantly, BMS936564/MDX-1338 inhibited MM cell proliferation in vivo in xenograft mouse models. Conclusion. These studies therefore show that targeting CXCR-4 in MM by using BMS936564/MDX-1338 represents a valid therapeutic strategy in this disease. Disclosures: Roccaro: Roche:. Kuhne:BMS: Employment. Pan:Bristol-Myers Squibb: Employment. Cardarelli:Bristol-Myers Squibb: Employment. Ghobrial:Noxxon: Research Funding; Bristol-Myers Squibb: Research Funding; Millennium: Research Funding; Noxxon:; Millennium:; Celegene:; Novartis:.

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Vialinin A, an Edible Mushroom-Derived p-Terphenyl Antioxidant, Prevents VEGF-Induced Neovascularization In Vitro and In Vivo

Oxidative Medicine and Cellular Longevity ◽

10.1155/2018/1052102 ◽

2018 ◽

Vol 2018 ◽

pp. 1-10 ◽

Cited By ~ 4

Author(s):

Himangshu Sonowal ◽

Kirtikar Shukla ◽

Sumedha Kota ◽

Ashish Saxena ◽

Kota V. Ramana

Keyword(s):

Nuclear Translocation ◽

Vascular Endothelial Cell ◽

Natural Antioxidants ◽

Edible Mushroom ◽

Tube Formation ◽

Dependent Manner ◽

Vascular Endothelial ◽

In Vivo Models

Increased side toxicities and development of drug resistance are the major concern for the cancer chemotherapy using synthetic drugs. Therefore, identification of novel natural antioxidants with potential therapeutic efficacies is important. In the present study, we have examined how the antioxidant and anti-inflammatory activities of vialinin A, a p-terphenyl compound derived from Chinese edible mushroomT. terrestrisandT. vialis, prevents human umbilical vascular endothelial cell (HUVEC) neovascularization in vitro and in vivo models. Pretreatment of HUVECs with vialinin A prevents vascular endothelial growth factor- (VEGF) induced HUVEC cell growth in a dose-dependent manner. Further, vialinin A also inhibits VEGF-induced migration as well as tube formation of HUVECs. Treatment of HUVECs prevents VEGF-induced generation of reactive oxygen species (ROS) and malondialdehyde (MDA) and also inhibits VEGF-induced NF-κB nuclear translocation as well as DNA-binding activity. The VEGF-induced release of various angiogenic cytokines and chemokines in HUVECs was also significantly blunted by vialinin A. Most importantly, in a mouse model of Matrigel plug assay, vialinin A prevents the formation of new blood vessels and the expression of CD31 and vWF. Thus, our results indicate a novel role of vialinin A in the prevention of neovascularization and suggest that anticancer effects of vialinin A could be mediated through its potent antioxidant and antiangiogenic properties.

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