bluuebunny/crossref_metadata_2025 huggingface dataset:
Data from NEI-01-Induced Arginine Deprivation Has Potent Activity Against Acute Myeloid Leukemia Cells Both In Vitro and In Vivo
Authors: Cai Yijun , Chow Jeremy P.H. , Leung Yu-On , Lu Xiaoxu , Yuen Chak-Ho , Lee Wing Lun , Chau Ka-Chun , Yang Liz L. , Wong Raymond M.H. , Lam Justin Y.T. , Chow Daniel T.L. , Chung Steven H.K. , Kwok Sui-Yi , Leung Yun-chung
Published: April 2023

Abstract
Recent studies have revealed that targeting amino acid metabolic enzymes is a promising strategy in cancer therapy. Acute myeloid leukemia (AML) downregulates the expression of argininosuccinate synthase (ASS1), a recognized rate-limiting enzyme for arginine synthesis, and yet displays a critical dependence on extracellular arginine for survival and proliferation. This dependence on extracellular arginine, also known as arginine auxotrophy, suggests that arginine deprivation would be a treatment strategy for AML. NEI-01, a novel arginine-depleting enzyme, is capable of binding to serum albumin to extend its circulating half-life, leading to a potent anticancer activity. Here we reported the preclinical activity of NEI-01 in arginine auxotrophic AMLs. NEI-01 efficiently depleted arginine both in vitro and in vivo. NEI-01-induced arginine deprivation was cytotoxic to arginine auxotrophic AML cells through induction of cell-cycle arrest and apoptosis. Furthermore, the potent anti-leukemia activities of NEI-01 were observed in three different types of mouse models including human cell line-derived xenograft, mouse cell line-derived homografts in syngeneic mice and patient-derived xenograft. This preclinical data provide strong evidence to support the potential use of NEI-01 as a therapeutic approach in AML treatment.
Data from PLK1 and NOTCH Positively Correlate in Melanoma and Their Combined Inhibition Results in Synergistic Modulations of Key Melanoma Pathways
Authors: Su Shengqin , Chhabra Gagan , Ndiaye Mary A. , Singh Chandra K. , Ye Ting , Huang Wei , Dewey Colin N. , Setaluri Vijayasaradhi , Ahmad Nihal
Published: April 2023

Abstract
Melanoma is one of the most serious forms of skin cancer, and its increasing incidence coupled with nonlasting therapeutic options for metastatic disease highlights the need for additional novel approaches for its management. In this study, we determined the potential interactions between polo-like kinase 1 (PLK1, a serine/threonine kinase involved in mitotic regulation) and NOTCH1 (a type I transmembrane protein deciding cell fate during development) in melanoma. Employing an in-house human melanoma tissue microarray (TMA) containing multiple cases of melanomas and benign nevi, coupled with high-throughput, multispectral quantitative fluorescence imaging analysis, we found a positive correlation between PLK1 and NOTCH1 in melanoma. Furthermore, The Cancer Genome Atlas database analysis of patients with melanoma showed an association of higher mRNA levels of PLK1 and NOTCH1 with poor overall, as well as disease-free, survival. Next, utilizing small-molecule inhibitors of PLK1 and NOTCH (BI 6727 and MK-0752, respectively), we found a synergistic antiproliferative response of combined treatment in multiple human melanoma cells. To determine the molecular targets of the overall and synergistic responses of combined PLK1 and NOTCH inhibition, we conducted RNA-sequencing analysis employing a unique regression model with interaction terms. We identified the modulations of several key genes relevant to melanoma progression/metastasis, including MAPK, PI3K, and RAS, as well as some new genes such as Apobec3G, BTK, and FCER1G, which have not been well studied in melanoma. In conclusion, our study demonstrated a synergistic antiproliferative response of concomitant targeting of PLK1 and NOTCH in melanoma, unraveling a potential novel therapeutic approach for detailed preclinical/clinical evaluation.
Data from Identification of ALDH1A3 as a Viable Therapeutic Target in Breast Cancer Metastasis–Initiating Cells
Authors: Yamashita Daisuke , Minata Mutsuko , Ibrahim Ahmed N. , Yamaguchi Shinobu , Coviello Vito , Bernstock Joshua D. , Harada Shuko , Cerione Richard A. , Tannous Bakhos A. , La Motta Concettina , Nakano Ichiro
Published: April 2023

Abstract
The development of efficacious therapies targeting metastatic spread of breast cancer to the brain represents an unmet clinical need. Accordingly, an improved understanding of the molecular underpinnings of central nervous system spread and progression of breast cancer brain metastases (BCBM) is required. In this study, the clinical burden of disease in BCBM was investigated, as well as the role of aldehyde dehydrogenase 1A3 (ALDH1A3) in the metastatic cascade leading to BCBM development. Initial analysis of clinical survival trends for breast cancer and BCBM determined improvement of breast cancer survival rates; however, this has failed to positively affect the prognostic milestones of triple-negative breast cancer (TNBC) brain metastases (BM). ALDH1A3 and a representative epithelial–mesenchymal transition (EMT) gene signature (mesenchymal markers, CD44 or Vimentin) were compared in tumors derived from BM, lung metastases (LM), or bone metastases (BoM) of patients as well as mice after injection of TNBC cells. Selective elevation of the EMT signature and ALDH1A3 were observed in BM, unlike LM and BoM, especially in the tumor edge. Furthermore, ALDH1A3 was determined to play a role in BCBM establishment via regulation of circulating tumor cell adhesion and migration phases in the BCBM cascade. Validation through genetic and pharmacologic inhibition of ALDH1A3 via lentiviral shRNA knockdown and a novel small-molecule inhibitor demonstrated selective inhibition of BCBM formation with prolonged survival of tumor-bearing mice. Given the survival benefits via targeting ALDH1A3, it may prove an effective therapeutic strategy for BCBM prevention and/or treatment.
Data from Epitope and Fc-Mediated Cross-linking, but Not High Affinity, Are Critical for Antitumor Activity of CD137 Agonist Antibody with Reduced Liver Toxicity
Authors: Ho Sun K. , Xu Zhenghai , Thakur Archana , Fox Melvin , Tan Siu Sze , DiGiammarino Enrico , Zhou Li , Sho Mien , Cairns Belinda , Zhao Vivian , Xiong Mengli , Samayoa Josue , Forsyth Charles M. , Powers David B. , Chao Debra T. , Hollenbaugh Diane , Alvarez Hamsell M. , Akamatsu Yoshiko
Published: April 2023

Abstract
CD137 (TNFRSF9, 4-1BB) agonist antibodies (mAb) have demonstrated potent antitumor activity with memory response while causing hepatotoxicity in mouse models. In clinical trials, the degrees of liver toxicity of anti-CD137 vary from grade 4 transaminitis (urelumab) to nonexistent (utomilumab). To exploit the antitumor potential of CD137 signaling, we identified a new class of CD137 agonist mAbs with strong antitumor potency without significant transaminitis in vivo compared with CD137 agonists previously reported. These mAbs are cross-reactive to mouse and cynomolgus monkey and showed cross-linking–dependent T-cell costimulation activity in vitro. Antitumor efficacy was maintained in Fc gamma receptor (FcγR) III–deficient mice but diminished in FcγRIIB-deficient mice, suggesting the critical role for FcγRIIB to provide cross-linking in vivo. Interestingly, a single dose of an affinity-reduced variant was sufficient to control tumor growth, but a higher affinity variant did not improve efficacy. These observations suggest that binding epitope and FcγR interaction, but not necessarily high affinity, are important for antitumor efficacy and reduced liver toxicity of CD137 mAb. Our study suggests the possibility of CD137 agonist therapy with improved safety profile in humans.
Data from An Anti–CD22-seco-CBI-Dimer Antibody–Drug Conjugate (ADC) for the Treatment of Non-Hodgkin Lymphoma That Provides a Longer Duration of Response than Auristatin-Based ADCs in Preclinical Models
Authors: Yu Shang-Fan , Lee Donna W. , Zheng Bing , del Rosario Geoffrey , Leipold Douglas , Booler Helen , Zhong Fiona , Carrasco-Triguero Montserrat , Hong Kyu , Yan Peter , Rowntree Rebecca K. , Schutten Melissa M. , Pillow Thomas , Sadowsky Jack D. , Dragovich Peter S. , Polson Andrew G.
Published: April 2023

Abstract
We are interested in developing a second generation of antibody–drug conjugates (ADCs) for the treatment of non-Hodgkin lymphoma (NHL) that could provide a longer duration of response and be more effective in indolent NHL than the microtubule-inhibiting ADCs pinatuzumab vedotin [anti–CD22-vc-monomethyl auristatin E (MMAE)] and polatuzumab vedotin (anti–CD79b-vc-MMAE). Pinatuzumab vedotin (anti–CD22-vc-MMAE) and polatuzumab vedotin (anti–CD79b-vc-MMAE) are ADCs that contain the microtubule inhibitor MMAE. Clinical trial data suggest that these ADCs have promising efficacy for the treatment of NHL; however, some patients do not respond or become resistant to the ADCs. We tested an anti-CD22 ADC with a seco-CBI-dimer payload, thio-Hu anti–CD22-(LC:K149C)-SN36248, and compared it with pinatuzumab vedotin for its efficacy and duration of response in xenograft models and its ability to deplete normal B cells in cynomolgus monkeys. We found that anti–CD22-(LC:K149C)-SN36248 was effective in xenograft models resistant to pinatuzumab vedotin, gave a longer duration of response, had a different mechanism of resistance, and was able to deplete normal B cells better than pinatuzumab vedotin. These studies provide evidence that anti–CD22-(LC:K149C)-SN36248 has the potential for longer duration of response and more efficacy in indolent NHL than MMAE ADCs and may provide the opportunity to improve outcomes for patients with NHL.
Data from Preclinical Testing of a Novel Niclosamide Stearate Prodrug Therapeutic (NSPT) Shows Efficacy Against Osteosarcoma
Authors: Reddy Gireesh B. , Kerr David L. , Spasojevic Ivan , Tovmasyan Artak , Hsu David S. , Brigman Brian E. , Somarelli Jason A. , Needham David , Eward William C.
Published: April 2023

Abstract
Therapeutic advances for osteosarcoma have stagnated over the past several decades, leading to an unmet clinical need for patients. The purpose of this study was to develop a novel therapy for osteosarcoma by reformulating and validating niclosamide, an established anthelminthic agent, as a niclosamide stearate prodrug therapeutic (NSPT). We sought to improve the low and inefficient clinical bioavailability of oral dosing, especially for the relatively hydrophobic classes of anticancer drugs. Nanoparticles were fabricated by rapid solvent shifting and verified using dynamic light scattering and UV-vis spectrophotometry. NSPT efficacy was then studied in vitro for cell viability, cell proliferation, and intracellular signaling by Western blot analysis; ex vivo pulmonary metastatic assay model; and in vivo pharmacokinetic and lung mouse metastatic model of osteosarcoma. NSPT formulation stabilizes niclosamide stearate against hydrolysis and delays enzymolysis; increases circulation in vivo with t1/2 approximately 5 hours; reduces cell viability and cell proliferation in human and canine osteosarcoma cells in vitro at 0.2–2 μmol/L IC50; inhibits recognized growth pathways and induces apoptosis at 20 μmol/L; eliminates metastatic lesions in the ex vivo lung metastatic model; and when injected intravenously at 50 mg/kg weekly, it prevents metastatic spread in the lungs in a mouse model of osteosarcoma over 30 days. In conclusion, niclosamide was optimized for preclinical drug delivery as a unique prodrug nanoparticle injected intravenously at 50 mg/kg (1.9 mmol/L). This increased bioavailability of niclosamide in the blood stream prevented metastatic disease in the mouse. This chemotherapeutic strategy is now ready for canine trials, and if successful, will be targeted for human trials in patients with osteosarcoma.
Data from Circulating Tumor Cells In Advanced Cervical Cancer: NRG Oncology—Gynecologic Oncology Group Study 240 (NCT 00803062)
Authors: Tewari Krishnansu S. , Sill Michael W. , Monk Bradley J. , Penson Richard T. , Moore David H. , Lankes Heather A. , Ramondetta Lois M. , Landrum Lisa M. , Randall Leslie M. , Oaknin Ana , Leitao Mario M. , Eisenhauer Eric L. , DiSilvestro Paul , Van Le Linda , Pearl Michael L. , Burke James J. , Salani Ritu , Richardson Debra L. , Michael Helen E. , Kindelberger David W. , Birrer Michael J.
Published: April 2023

Abstract
To isolate circulating tumor cells (CTC) from women with advanced cervical cancer and estimate the impact of CTCs and treatment on overall survival and progression-free survival (PFS). A total of 7.5 mL of whole blood was drawn pre-cycle 1 and 36 days post-cycle 1 from patients enrolled on Gynecologic Oncology Group 0240, the phase III randomized trial that led directly to regulatory approval of the antiangiogenesis drug, bevacizumab, in women with recurrent/metastatic cervical cancer. CTCs (defined as anti-cytokeratin+/anti-CD45− cells) were isolated from the buffy coat layer using an anti-EpCAM antibody-conjugated ferrofluid and rare earth magnet, and counted using a semiautomated fluorescence microscope. The median pre-cycle 1 CTC count was 7 CTCs/7.5 mL whole blood (range, 0–18) and, at 36 days posttreatment, was 4 (range, 0–17). The greater the declination in CTCs between time points studied, the lower the risk of death [HR, 0.87; 95% confidence interval (CI), 0.79–0.95)]. Among patients with high (≥ median) pretreatment CTCs, bevacizumab treatment was associated with a reduction in the hazard of death (HR, 0.57; 95% CI, 0.32–1.03) and PFS (HR, 0.59; 95% CI, 0.36–0.96). This effect was not observed with low (< median) CTCs. CTCs can be isolated from women with advanced cervical cancer and may have prognostic significance. A survival benefit conferred by bevacizumab among patients with high pretreatment CTCs may reflect increased tumor neovascularization and concomitant vulnerability to VEGF inhibition. These data support studying CTC capture as a potential predictive biomarker.
Data from Preclinical Activity of HER2-Selective Tyrosine Kinase Inhibitor Tucatinib as a Single Agent or in Combination with Trastuzumab or Docetaxel in Solid Tumor Models
Authors: Kulukian Anita , Lee Patrice , Taylor Janelle , Rosler Robert , de Vries Peter , Watson Daniel , Forero-Torres Andres , Peterson Scott
Published: April 2023

Abstract
HER2 is a transmembrane tyrosine kinase receptor that mediates cell growth, differentiation, and survival. HER2 is overexpressed in approximately 20% of breast cancers and in subsets of gastric, colorectal, and esophageal cancers. Both antibody and small-molecule drugs that target HER2 and block its tyrosine kinase activity are effective in treating HER2-driven cancers. In this article, we describe the preclinical properties of tucatinib, an orally available, reversible HER2-targeted small-molecule tyrosine kinase inhibitor. In both biochemical and cell signaling experiments, tucatinib inhibits HER2 kinase activity with single-digit nanomolar potency and provides exceptional selectivity for HER2 compared with the related receptor tyrosine kinase EGFR, with a >1,000-fold enhancement in potency for HER2 in cell signaling assays. Tucatinib potently inhibits signal transduction downstream of HER2 and HER3 through the MAPK and PI3K/AKT pathways and is selectively cytotoxic in HER2-amplified breast cancer cell lines in vitro. In vivo, tucatinib is active in multiple HER2+ tumor models as a single agent and shows enhanced antitumor activity in combination with trastuzumab or docetaxel, resulting in improved rates of partial and complete tumor regression. These preclinical data, taken together with the phase-I tucatinib clinical trial results demonstrating preliminary safety and activity, establish the unique pharmacologic properties of tucatinib and underscore the rationale for investigating its utility in HER2+ cancers.
Data from Repurposing of a Thromboxane Receptor Inhibitor Based on a Novel Role in Metastasis Identified by Phenome-Wide Association Study
Authors: Werfel Thomas A. , Hicks Donna J. , Rahman Bushra , Bendeman Wendy E. , Duvernay Matthew T. , Maeng Jae G. , Hamm Heidi , Lavieri Robert R. , Joly Meghan M. , Pulley Jill M. , Elion David L. , Brantley-Sieders Dana M. , Cook Rebecca S.
Published: April 2023

Abstract
Although new drug discoveries are revolutionizing cancer treatments, repurposing existing drugs would accelerate the timeline and lower the cost for bringing treatments to cancer patients. Our goal was to repurpose CPI211, a potent and selective antagonist of the thromboxane A2-prostanoid receptor (TPr), a G-protein–coupled receptor that regulates coagulation, blood pressure, and cardiovascular homeostasis. To identify potential new clinical indications for CPI211, we performed a phenome-wide association study (PheWAS) of the gene encoding TPr, TBXA2R, using robust deidentified health records and matched genomic data from more than 29,000 patients. Specifically, PheWAS was used to identify clinical manifestations correlating with a TBXA2R single-nucleotide polymorphism (rs200445019), which generates a T399A substitution within TPr that enhances TPr signaling. Previous studies have correlated 200445019 with chronic venous hypertension, which was recapitulated by this PheWAS analysis. Unexpectedly, PheWAS uncovered an rs200445019 correlation with cancer metastasis across several cancer types. When tested in several mouse models of metastasis, TPr inhibition using CPI211 potently blocked spontaneous metastasis from primary tumors, without affecting tumor cell proliferation, motility, or tumor growth. Further, metastasis following intravenous tumor cell delivery was blocked in mice treated with CPI211. Interestingly, TPr signaling in vascular endothelial cells induced VE-cadherin internalization, diminished endothelial barrier function, and enhanced transendothelial migration by tumor cells, phenotypes that were decreased by CPI211. These studies provide evidence that TPr signaling promotes cancer metastasis, supporting the study of TPr inhibitors as antimetastatic agents and highlighting the use of PheWAS as an approach to accelerate drug repurposing.
Data from Simultaneously Combined Cancer Cell- and CTLA4-Targeted NIR-PIT Causes a Synergistic Treatment Effect in Syngeneic Mouse Models
Authors: Kato Takuya , Okada Ryuhei , Furusawa Aki , Inagaki Fuyuki , Wakiyama Hiroaki , Furumoto Hideyuki , Okuyama Shuhei , Fukushima Hiroshi , Choyke Peter L. , Kobayashi Hisataka
Published: April 2023

Abstract
Near-infrared photoimmunotherapy (NIR-PIT) is a new cancer treatment that utilizes antibody–IRDye700DX (IR700) conjugates. The clinical use of NIR-PIT has recently been approved in Japan for patients with inoperable head and neck cancer targeting human epidermal growth factor receptor (hEGFR). Previously, cytotoxic T-lymphocyte antigen 4 (CTLA4)–targeted NIR-PIT has been shown to strongly inhibit tumor progression and prolonged survival was seen in different tumor models due to enhanced T-cell–mediated antitumor immunity. In this study, combined NIR-PIT targeting CTLA4 expressing cells and cancer cells was investigated in four tumor models including a newly established hEGFR-expressing murine oropharyngeal cancer cell (mEERL-hEGFR). While single molecule–targeted therapy (NIR-PIT targeting hEGFR or CTLA4) did not inhibit tumor progression in poorly immunogenic mEERL-hEGFR tumor, dual (CTLA4/hEGFR)-targeted NIR-PIT significantly suppressed tumor growth and prolonged survival resulting in a 38% complete response rate. After the dual-targeted NIR-PIT, depletion of CTLA4 expressing cells, which were mainly regulatory T cells (Tregs), and an increase in the CD8+/Treg ratio in the tumor bed were observed, suggesting enhanced host antitumor immunity. Furthermore, dual-targeted NIR-PIT showed antitumor immunity in distant untreated tumors of the same type. Thus, simultaneous cancer cell–targeted NIR-PIT and CTLA4-targeted NIR-PIT is a promising new cancer therapy strategy, especially in poorly immunogenic tumors where NIR-PIT monotherapy is suboptimal.
Data from Molecular Characterization of Appendiceal Goblet Cell Carcinoid
Authors: Arai Hiroyuki , Baca Yasmine , Battaglin Francesca , Kawanishi Natsuko , Wang Jingyuan , Soni Shivani , Zhang Wu , Millstein Joshua , Johnston Curtis , Goldberg Richard M. , Philip Philip A. , Seeber Andreas , Xiu Joanne , Hwang Jimmy J. , Shields Anthony F. , Marshall John L. , Korn W. Michael , Lenz Heinz-Josef
Published: April 2023

Abstract
Goblet cell carcinoid (GCC) is a distinct subtype of appendiceal neoplasm that exhibits unique clinical and pathologic features. We aimed to reveal the molecular profiles of GCC compared with other appendiceal tumors, such as adenocarcinomas and neuroendocrine tumors. A total of 495 appendiceal tumor samples (53 GCCs, 428 adenocarcinomas, and 14 neuroendocrine tumors) were tested with next-generation sequencing (NGS) on a 592-gene panel and IHC. Microsatellite instability (MSI)/mismatch repair (MMR) status was tested with a combination of NGS, IHC, and fragment analyses. Tumor mutational burden (TMB) was evaluated by NGS, and PD-L1 expression was tested by IHC (SP142). The most prevalent mutated genes within GCCs were TP53 (24.0%), ARID1A (15.4%), SMAD4 (9.4%), and KRAS (7.5%). Pathway-specific alterations were dominantly observed in cell cycle, MAPK, epigenetic, and TGFβ signaling pathways. GCCs as compared with adenocarcinomas exhibited significantly lower mutation rates in KRAS, GNAS, and APC, and significantly higher mutation rates in CDH1, CHEK2, CDC73, ERCC2, and FGFR2. GCCs as compared with neuroendocrine tumors showed significantly lower mutation rates in KRAS, APC, BRCA2, and FANCA. In GCCs, MSI high/MMR deficient, TMB high (≥17 mutations/Mb), and PD-L1 expression were seen in 0.0%, 0.0%, and 2.0% of tumors, respectively. No significant differences were observed in any immunotherapy-related markers examined when compared with adenocarcinomas and neuroendocrine tumors. In conclusion, GCCs had considerably distinct mutational profiles compared with appendiceal adenocarcinomas and neuroendocrine tumors. Understanding these molecular characteristics may be critical for the development of novel and more effective treatment strategies for GCC.
Data from Evaluation of Preclinical Activity of Isatuximab in Patients with Acute Lymphoblastic Leukemia
Authors: Wang Anlai , Song Zhili , Zheng Gang , Nicolazzi Celine , Fromm Jonathan R. , Shehu Elvis , Srinivasan Srimathi , Chen Xueyan , Zhu Chen , Blondel Marielle Chiron , Adrian Francisco J.
Published: April 2023

Abstract
This study reports the pharmacologic effects of isatuximab, a CD38 mAb, on T- and B-cell acute lymphoblastic leukemia (ALL). We analyzed CD38 expression in 50-T-ALL and 50 B-ALL clinical samples, and 16 T-ALL and 11 B-ALL cell lines. We primarily focused on in vitro assessments of isatuximab-mediated antibody-dependent cellular cytotoxicity (ADCC) and antibody-dependent cellular phagocytosis (ADCP). In vivo assessment of isatuximab activity was performed in several ALL xenograft models, including disseminated and subcutaneous tumor models in female C.B-17 severe combined immunodeficiency mice. Our study reveals that most patients (90%–100%) carried CD38+ blasts independent of disease burden. The median CD38 receptor density on abnormal lymphoblasts is 41,026 copies/cell on T-ALL and 28,137 copies/cell on B-ALL, respectively. In patients with T-ALL, there is a significant increase of CD38 expression in abnormal blasts compared with normal T cells. High-level CD38 receptor density (RD) is critical to trigger effective isatuximab-mediated ADCC against target ALL cells. In addition, a correlation between CD38 RD and isatuximab-mediated ADCP is demonstrated. In the disseminated CD38+, T-ALL, and B-ALL xenograft models, isatuximab is able to induce robust antitumor activity, even at low doses. This study shows that isatuximab has significant in vitro and in vivo activity against ALL cells with robust ADCC and ADCP effects that are associated with CD38 expression levels in both T-ALL and B-ALL.
Data from Preclinical Activity of PI3K Inhibitor Copanlisib in Gastrointestinal Stromal Tumor
Authors: García-Valverde Alfonso , Rosell Jordi , Serna Garazi , Valverde Claudia , Carles Joan , Nuciforo Paolo , Fletcher Jonathan A. , Arribas Joaquín , Politz Oliver , Serrano César
Published: April 2023

Abstract
KIT or PDGFRA gain-of-function mutations are the primary drivers of gastrointestinal stromal tumor (GIST) growth and progression throughout the disease course. The PI3K/mTOR pathway is critically involved in the transduction of KIT/PDGFRA oncogenic signaling regardless of the type of primary and secondary mutations, and therefore emerges as a relevant targetable node in GIST biology. We evaluated in GIST preclinical models the antitumor activity of copanlisib, a novel pan-class-I PI3K inhibitor with predominant activity against p110α and p110δ isoforms, as single-agent and in combination with first-line KIT inhibitor imatinib. In vitro studies undertaken in one imatinib-sensitive (GIST-T1) and two imatinib-resistant (GIST-T1/670 and GIST430/654) GIST cell models showed that single-agent copanlisib effectively suppressed PI3K pathway activation leading to decreased cell viability and proliferation in both imatinib-sensitive and -resistant cells irrespective of the type of primary or secondary KIT mutations. Simultaneous PI3K and KIT inhibition with copanlisib and imatinib resulted in enhanced impairment of cell viability in both imatinib-sensitive and -resistant GIST cell models, although apoptosis was mostly triggered in GIST-T1. Single-agent copanlisib inhibited GIST growth in vivo, and conjoined inhibition of PI3K and KIT was the most active therapeutic intervention in imatinib-sensitive GIST-T1 xenografts. IHC stain for cleaved-caspase 3 and phospho-S6 support a predominant antiproliferative effect of copanlisib in GIST. In conclusion, copanlisib has single-agent antitumor activity in GIST regardless KIT mutational status or sensitivity to imatinib. Effective KIT inhibition is necessary to achieve synergistic or additive effects with the combination of imatinib and any given PI3K/mTOR pathway inhibition.
Data from Aurora B Kinase Promotes CHIP-Dependent Degradation of HIF1α in Prostate Cancer Cells
Authors: Biswas Kuntal , Sarkar Sukumar , Said Neveen , Brautigan David L. , Larner James M.
Published: April 2023

Abstract
Hypoxia is a major factor in tumor progression and resistance to therapies, which involves elevated levels of the transcription factor HIF1α. Here, we report that prostate tumor xenografts express high levels of HIF1α and show greatly enhanced growth in response to knockdown of the E3 ligase CHIP (C-terminus of Hsp70-interacting protein). In multiple human prostate cancer cell lines under hypoxia, taxol treatment induces the degradation of HIF1α, and this response is abrogated by knockdown of CHIP, but not by E3 ligase VHL or RACK1. HIF1α degradation is accompanied by loss of function, evidenced by reduced expression of HIF1α-dependent genes. CHIP-dependent HIF1α degradation also occurs in cells arrested in mitosis by nocodazole instead of taxol. Mitotic kinase Aurora B activity is required for taxol-induced HIF1α degradation. Purified Aurora B directly phosphorylates HIF1α at multiple sites, and these modifications enhance its polyubiquitination by CHIP in a purified reconstituted system. Our results show how activation of Aurora B promotes CHIP-dependent degradation of HIF1α in prostate cancer cells. This new knowledge may affect the use of mitotic kinase inhibitors and open new approaches for treatment of hypoxic prostate tumors.
Data from The Dual Androgen Receptor and Glucocorticoid Receptor Antagonist CB-03-10 as Potential Treatment for Tumors that have Acquired GR-mediated Resistance to AR Blockade
Authors: Rosette Caridad , Agan Frances J. , Rosette Niccolette , Mazzetti Alessandro , Moro Luigi , Gerloni Mara
Published: April 2023

Abstract
CB-03–10 (cortexolone 17α-valerate-21-propionate) is a synthetic steroidal compound derived from cortexolone (11-deoxycortisone), an intermediate in cortisol biosynthesis. Characterization of the activity of CB-03-10 and its main related compound CB-03–05 (cortexolone 17α-valerate) included in vitro binding to the androgen and glucocorticoid receptors (AR and GR), antagonism of AR and GR transcriptional activities, and screening for antitumor activity across a selected panel of human prostate and in triple-negative breast cancer cell lines. CB-03-10 cytotoxic activity in these cancer cell lines was in the low micromolar range and was primarily associated with induction of the apoptotic cascade via activation of caspases. The compound's potential for antitumor activity was verified in a murine xenograft model utilizing the AR-positive LNCaP prostate cancer cell line as well as in an orthotopic model utilizing AR-negative/GR-positive MDA-MB-231 breast cancer cell line. Orally administered CB-03-10 inhibited prostate tumor growth and orthotopically implanted breast tumor growth in these mice and maintained body weight, as compared with vehicle-treated mice. On the basis of AR/GR binding affinities, antagonism of androgen and glucocorticoid-dependent transcriptional activities, and AR/GR mRNA and protein expression, the mechanism of tumor growth suppression is related to AR and GR antagonist activities. Importantly, these compounds lack biologically relevant AR/GR agonist activities. Overall, these preclinical findings support the selection of CB-03-10 for further development as an anticancer agent in cases where resistance to AR-targeted therapy or chemotherapy, via upregulation of GR activity, continues to limit the efficacy and duration of clinical benefit with these interventions.
Data from An Antibody–Drug Conjugate Targeting MUC1-Associated Carbohydrate CA6 Shows Promising Antitumor Activities
Authors: Nicolazzi Céline , Caron Anne , Tellier Alexia , Trombe Marc , Pinkas Jan , Payne Gillian , Carrez Chantal , Guérif Stéphane , Maguin Marie , Baffa Raffaele , Fassan Matteo , Adam Julien , Mangatal-Wade Lydie , Blanc Véronique
Published: April 2023

Abstract
Glycosylation is a complex multienzyme-related process that is frequently deregulated in cancer. Aberrant glycosylation can lead to the generation of novel tumor surface–specific glycotopes that can be targeted by antibodies. Murine DS6 mAb (muDS6) was generated from serous ovary adenocarcinoma immunization. It recognizes CA6, a Mucin-1 (MUC1)-associated sialoglycotope that is highly detected in breast, ovarian, lung, and bladder carcinomas. SAR566658 antibody–drug conjugate (ADC) is a humanized DS6 (huDS6) antibody conjugated through a cleavable linker to the cytotoxic maytansinoid derivative drug, DM4. SAR566658 binds to tumor cells with subnanomolar affinity, allowing good ADC internalization and intracellular delivery of DM4, resulting in tumor cell death (IC50 from 1 to 7.3 nmol/L). SAR566658 showed in vivo antitumor efficacy against CA6-positive human pancreas, cervix, bladder, and ovary tumor xenografts and against three breast patient-derived xenografts. Tumor regression was observed in all tumor models with minimal effective dose correlating with CA6 expression. SAR566658 displayed better efficacy than standard-of-care nontargeted tubulin binders. These data support the development of SAR566658 in patients with CA6-expressing tumors.
Data from Combined Targeting of the BRD4–NUT–p300 Axis in NUT Midline Carcinoma by Dual Selective Bromodomain Inhibitor, NEO2734
Authors: Morrison-Smith Chevaun D. , Knox Tatiana M. , Filic Ivona , Soroko Kara M. , Eschle Benjamin K. , Wilkens Margaret K. , Gokhale Prafulla C. , Giles Francis , Griffin Andrew , Brown Bill , Shapiro Geoffrey I. , Zucconi Beth E. , Cole Philip A. , Lemieux Madeleine E. , French Christopher A.
Published: April 2023

Abstract
NUT midline carcinoma (NMC) is a rare, aggressive subtype of squamous carcinoma that is driven by the BRD4-NUT fusion oncoprotein. BRD4, a BET protein, binds to chromatin through its two bromodomains, and NUT recruits the p300 histone acetyltransferse (HAT) to activate transcription of oncogenic target genes. BET-selective bromodomain inhibitors have demonstrated on-target activity in patients with NMC, but with limited efficacy. P300, like BRD4, contains a bromodomain. We show that combining selective p300/CBP and BET bromodomain inhibitors, GNE-781 and OTX015, respectively, induces cooperative depletion of MYC and synergistic inhibition of NMC growth. Treatment of NMC cells with the novel dual p300/CBP and BET bromodomain–selective inhibitor, NEO2734, potently inhibits growth and induces differentiation of NMC cells in vitro; findings that correspond with potentiated transcriptional effects from combined BET and p300 bromodomain inhibition. In three disseminated NMC xenograft models, NEO2734 provided greater growth inhibition, with tumor regression and significant survival benefit seen in two of three models, compared with a lead clinical BET inhibitor or “standard” chemotherapy. Our findings provide a strong rationale for clinical study of NEO2734 in patients with NMC. Moreover, the synergistic inhibition of NMC growth by CBP/p300 and BET bromodomain inhibition lays the groundwork for greater mechanistic understanding of the interplay between p300 and BRD4-NUT that drives this cancer.
Data from CHK1/2 Inhibitor Prexasertib Suppresses NOTCH Signaling and Enhances Cytotoxicity of Cisplatin and Radiation in Head and Neck Squamous Cell Carcinoma
Authors: Zeng Ling , Nikolaev Anatoly , Xing Chuan , Della Manna Deborah L. , Yang Eddy S.
Published: April 2023

Abstract
Platinum-based chemoradiotherapy is a mainstay of organ-preserving therapy for patients with head and neck squamous cell carcinoma cancer (HNSCC). However, the disease eventually becomes resistant to treatment necessitating new therapies. Checkpoint kinase 1 and 2 (CHK1/2) are serine/threonine kinases that activate cell-cycle checkpoints and serve a critical role in the DNA-damage response (DDR). As resistance to cisplatin and radiation may involve a heightened DDR, we hypothesized that prexasertib, an inhibitor of CHK1/2, may enhance the cytotoxicity induced by cisplatin and irradiation in HNSCC. In this study, we found that combining prexasertib with cisplatin and radiation significantly decreased the in vitro survival fraction in HNSCC cell lines both with and without radiotherapy. Reduced survival was accompanied by inhibition of DNA repair checkpoint activation, which resulted in persistent DNA damage and increased apoptosis. In addition, NanoString analysis with the PanCancer Pathways Panel revealed that prexasertib downregulated NOTCH signaling target genes (NOTCH1, NOTCH2, and NOTCH3) and their associated ligands (JAG1, JAG2, SKP2, MAML2, and DLL1). Prexasertib also reduced NOTCH1, NOTCH3 and HES1 protein expression. Importantly, a significant tumor growth delay was observed in vivo in both human papillomavirus (HPV)-positive UM-SCC47 and HPV-negative UM-SCC1 cell line xenografts treated with prexasertib, cisplatin, and radiotherapy without increased toxicity as measured by mouse body weight. Taken together, prexasertib reduced NOTCH signaling and enhanced the in vitro and in vivo response of HNSCCs to cisplatin and radiation, suggesting combination therapy may increase clinical benefit. A clinical trial has recently completed accrual (NCT02555644).
Data from Dual-Mechanism ERK1/2 Inhibitors Exploit a Distinct Binding Mode to Block Phosphorylation and Nuclear Accumulation of ERK1/2
Authors: Kidger Andrew M. , Munck Joanne M. , Saini Harpreet K. , Balmanno Kathryn , Minihane Emma , Courtin Aurelie , Graham Brent , O'Reilly Marc , Odle Richard , Cook Simon J.
Published: April 2023

Abstract
The RAS-regulated RAF-MEK1/2-ERK1/2 signaling pathway is frequently deregulated in cancer due to activating mutations of growth factor receptors, RAS or BRAF. Both RAF and MEK1/2 inhibitors are clinically approved and various ERK1/2 inhibitors (ERKi) are currently undergoing clinical trials. To date, ERKi display two distinct mechanisms of action (MoA): catalytic ERKi solely inhibit ERK1/2 catalytic activity, whereas dual mechanism ERKi additionally prevents the activating phosphorylation of ERK1/2 at its T-E-Y motif by MEK1/2. These differences may impart significant differences in biological activity because T-E-Y phosphorylation is the signal for nuclear entry of ERK1/2, allowing them to access many key transcription factor targets. Here, we characterized the MoA of five ERKi and examined their functional consequences in terms of ERK1/2 signaling, gene expression, and antiproliferative efficacy. We demonstrate that catalytic ERKi promote a striking nuclear accumulation of p-ERK1/2 in KRAS-mutant cell lines. In contrast, dual-mechanism ERKi exploits a distinct binding mode to block ERK1/2 phosphorylation by MEK1/2, exhibit superior potency, and prevent the nuclear accumulation of ERK1/2. Consequently, dual-mechanism ERKi exhibit more durable pathway inhibition and enhanced suppression of ERK1/2-dependent gene expression compared with catalytic ERKi, resulting in increased efficacy across BRAF- and RAS-mutant cell lines.
Data from PIM Kinase Inhibitors Block the Growth of Primary T-cell Acute Lymphoblastic Leukemia: Resistance Pathways Identified by Network Modeling Analysis
Authors: Lim James T. , Singh Neha , Leuvano Libia A. , Calvert Valerie S. , Petricoin Emanuel F. , Teachey David T. , Lock Richard B. , Padi Megha , Kraft Andrew S. , Padi Sathish K.R.
Published: April 2023

Abstract
Despite significant progress in understanding the genetic landscape of T-cell acute lymphoblastic leukemia (T-ALL), the discovery of novel therapeutic targets has been difficult. Our results demonstrate that the levels of PIM1 protein kinase is elevated in early T-cell precursor ALL (ETP-ALL) but not in mature T-ALL primary samples. Small-molecule PIM inhibitor (PIMi) treatment decreases leukemia burden in ETP-ALL. However, treatment of animals carrying ETP-ALL with PIMi was not curative. To model other pathways that could be targeted to complement PIMi activity, HSB-2 cells, previously characterized as a PIMi-sensitive T-ALL cell line, were grown in increasing doses of PIMi. Gene set enrichment analysis of RNA sequencing data and functional enrichment of network modules demonstrated that the HOXA9, mTOR, MYC, NFκB, and PI3K-AKT pathways were activated in HSB-2 cells after long-term PIM inhibition. Reverse phase protein array–based pathway activation mapping demonstrated alterations in the mTOR, PI3K-AKT, and NFκB pathways, as well. PIMi-tolerant HSB-2 cells contained phosphorylated RelA-S536 consistent with activation of the NFκB pathway. The combination of NFκB and PIMis markedly reduced the proliferation in PIMi-resistant leukemic cells showing that this pathway plays an important role in driving the growth of T-ALL. Together these results demonstrate key pathways that are activated when HSB-2 cell line develop resistance to PIMi and suggest pathways that can be rationally targeted in combination with PIM kinases to inhibit T-ALL growth.
Data from A Novel Approach to Safer Glucocorticoid Receptor–Targeted Anti-lymphoma Therapy via REDD1 (Regulated in Development and DNA Damage 1) Inhibition
Authors: Lesovaya Ekaterina A. , Savinkova Alena V. , Morozova Olga V. , Lylova Evgeniya S. , Zhidkova Ekaterina M. , Kulikov Evgeny P. , Kirsanov Kirill I. , Klopot Anna , Baida Gleb , Yakubovskaya Marianna G. , Gordon Leo I. , Readhead Ben , Dudley Joel T. , Budunova Irina
Published: April 2023

Abstract
Glucocorticoids are widely used for therapy of hematologic malignancies. Unfortunately, chronic treatment with glucocorticoids commonly leads to adverse effects including skin and muscle atrophy and osteoporosis. We found recently that REDD1 (regulated in development and DNA damage 1) plays central role in steroid atrophy. Here, we tested whether REDD1 suppression makes glucocorticoid-based therapy of blood cancer safer. Unexpectedly, approximately 50% of top putative REDD1 inhibitors selected by bioinformatics screening of Library of Integrated Network-Based Cellular Signatures database (LINCS) were PI3K/Akt/mTOR inhibitors. We selected Wortmannin, LY294002, and AZD8055 for our studies and showed that they blocked basal and glucocorticoid-induced REDD1 expression. Moreover, all PI3K/mTOR/Akt inhibitors modified glucocorticoid receptor function shifting it toward therapeutically important transrepression. PI3K/Akt/mTOR inhibitors enhanced anti-lymphoma effects of Dexamethasone in vitro and in vivo, in lymphoma xenograft model. The therapeutic effects of PI3K inhibitor+Dexamethasone combinations ranged from cooperative to synergistic, especially in case of LY294002 and Rapamycin, used as a previously characterized reference REDD1 inhibitor. We found that coadministration of LY294002 or Rapamycin with Dexamethasone protected skin against Dexamethasone-induced atrophy, and normalized RANKL/OPG ratio indicating a reduction of Dexamethasone-induced osteoporosis. Together, our results provide foundation for further development of safer and more effective glucocorticoid-based combination therapy of hematologic malignancies using PI3K/Akt/mTOR inhibitors.
Data from PAK4-NAMPT Dual Inhibition Sensitizes Pancreatic Neuroendocrine Tumors to Everolimus
Authors: Mpilla Gabriel B. , Uddin Md. Hafiz , Al-Hallak Mohammed N. , Aboukameel Amro , Li Yiwei , Kim Steve H. , Beydoun Rafic , Dyson Gregory , Baloglu Erkan , Senapedis William T. , Landesman Yosef , Wagner Kay-Uwe , Viola Nerissa T. , El-Rayes Bassel F. , Philip Philip A. , Mohammad Ramzi M. , Azmi Asfar S.
Published: April 2023

Abstract
Metastatic pancreatic neuroendocrine tumors (PNET) remain an unmet clinical problem. Chronologic treatment in PNETs includes observation (watchful protocol), surgery, targeted therapy, and chemotherapy. However, increasing evidence illustrates that the outcomes of targeted therapeutic options for the treatment of advanced PNETs show minimal response. The FDA-approved mTOR inhibitor everolimus does not shrink these tumors. It only delays disease progression in a subset of patients, while a significant fraction acquires resistance and shows disease progression. Thus, there is a need for more effective targeted approaches to sensitize PNETs to everolimus for better treatment outcomes. Previously, we showed that mTOR regulator p21 activated kinase 4 (PAK4) and nicotinamide adenine dinucleotide biosynthesis enzyme nicotinamide phosphoribosyl transferase (NAMPT) were aberrantly expressed in PNET tissue and promoted everolimus resistance. In this report, we demonstrate that PAK4-NAMPT dual inhibitor KPT-9274 can synergize with everolimus (growth inhibition, colony suppression, and glucose uptake assays). KPT-9274-everolimus disrupted spheroid formation in multiple PNET models. Molecular analysis showed alteration of mTORC2 through downregulation of RICTOR as a mechanism supporting synergy with everolimus in vitro. KPT-9274 suppressed β-catenin activity via inhibition of PAK4, highlighting the cross-talk between Rho GTPases and Wnt signaling in PNETs. KPT-9274, given at 150 mg/kg in combination with sub-MTD everolimus (2.5 mg/kg), significantly suppressed two PNET-derived xenografts. These studies bring forward a well-grounded strategy for advanced PNETs that fail to respond to single-agent everolimus.
Data from Reprogramming of Nucleotide Metabolism Mediates Synergy between Epigenetic Therapy and MAP Kinase Inhibition
Authors: Shorstova Tatiana , Su Jie , Zhao Tiejun , Dahabieh Michael , Leibovitch Matthew , De Sa Tavares Russo Mariana , Avizonis Daina , Rajkumar Shivshankari , Watson Ian R. , del Rincón Sonia V. , Miller Wilson H. , Foulkes William D. , Witcher Michael
Published: April 2023

Abstract
Small cell carcinoma of the ovary, hypercalcemic type (SCCOHT) is a rare but often lethal cancer that is diagnosed at a median age of 24 years. Optimal management of patients is not well defined, and current treatment remains challenging, necessitating the discovery of novel therapeutic approaches. The identification of SMARCA4-inactivating mutations invariably characterizing this type of cancer provided insights facilitating diagnostic and therapeutic measures against this disease. We show here that the BET inhibitor OTX015 acts in synergy with the MEK inhibitor cobimetinib to repress the proliferation of SCCOHT in vivo. Notably, this synergy is also observed in some SMARCA4-expressing ovarian adenocarcinoma models intrinsically resistant to BETi. Mass spectrometry, coupled with knockdown of newly found targets such as thymidylate synthase, revealed that the repression of a panel of proteins involved in nucleotide synthesis underlies this synergy both in vitro and in vivo, resulting in reduced pools of nucleotide metabolites and subsequent cell-cycle arrest. Overall, our data indicate that dual treatment with BETi and MEKi represents a rational combination therapy against SCCOHT and potentially additional ovarian cancer subtypes.
Data from Dual Epitope Targeting and Enhanced Hexamerization by DR5 Antibodies as a Novel Approach to Induce Potent Antitumor Activity Through DR5 Agonism
Authors: Overdijk Marije B. , Strumane Kristin , Beurskens Frank J. , Ortiz Buijsse Antonio , Vermot-Desroches Claudine , Vuillermoz Boris S. , Kroes Thessa , de Jong Bart , Hoevenaars Naomi , Hibbert Richard G. , Lingnau Andreas , Forssmann Ulf , Schuurman Janine , Parren Paul W.H.I. , de Jong Rob N. , Breij Esther C.W.
Published: April 2023

Abstract
Higher-order death receptor 5 (DR5) clustering can induce tumor cell death; however, therapeutic compounds targeting DR5 have achieved limited clinical efficacy. We describe HexaBody-DR5/DR5, an equimolar mixture of two DR5-specific IgG1 antibodies with an Fc-domain mutation that augments antibody hexamerization after cell surface target binding. The two antibodies do not compete for binding to DR5 as demonstrated using binding competition studies, and binding to distinct epitopes in the DR5 extracellular domain was confirmed by crystallography. The unique combination of dual epitope targeting and increased IgG hexamerization resulted in potent DR5 agonist activity by inducing efficient DR5 outside-in signaling and caspase-mediated cell death. Preclinical studies in vitro and in vivo demonstrated that maximal DR5 agonist activity could be achieved independent of Fc gamma receptor–mediated antibody crosslinking. Most optimal agonism was observed in the presence of complement complex C1, although without inducing complement-dependent cytotoxicity. It is hypothesized that C1 may stabilize IgG hexamers that are formed after binding of HexaBody-DR5/DR5 to DR5 on the plasma membrane, thereby strengthening DR5 clustering and subsequent outside-in signaling. We observed potent antitumor activity in vitro and in vivo in large panels of patient-derived xenograft models representing various solid cancers. The results of our preclinical studies provided the basis for an ongoing clinical trial exploring the activity of HexaBody-DR5/DR5 (GEN1029) in patients with malignant solid tumors.