bluuebunny/crossref_metadata_2025 huggingface dataset:
Data from Hyaluronidase-Expressing Salmonella Effectively Targets Tumor-Associated Hyaluronic Acid in Pancreatic Ductal Adenocarcinoma
Authors: Ebelt Nancy D. , Zuniga Edith , Passi Kevin B. , Sobocinski Lukas J. , Manuel Edwin R.
Published: March 2024

Abstract
In pancreatic ductal adenocarcinoma (PDAC), the extracellular matrix (ECM) surrounding cancer cells forms a barrier that often limits the ability of chemotherapeutic drugs and cytotoxic immune subsets to penetrate and eliminate tumors. The dense stromal matrix protecting cancer cells, also known as desmoplasia, results from the overproduction of major ECM components such as collagens and hyaluronic acid (HA). Although candidate drugs targeting ECM components have shown promise in increasing penetration of chemotherapeutic agents, severe adverse effects associated with systemic depletion of ECM in peripheral healthy tissues limits their use at higher, more effective doses. Currently, few strategies exist that preferentially degrade ECM in tumor tissue over healthy tissues. In light of this, we have developed an attenuated, tumor-targeting Salmonella typhimurium (ST) expressing functional bacterial hyaluronidase (bHs-ST), capable of degrading human HA deposited within PDAC tumors. Our data show that bHs-ST (i) targets and colonizes orthotopic human PDAC tumors following systemic administration and (ii) is efficiently induced in vivo to deplete tumor-derived HA, which in turn (iii) significantly increases diffusion of Salmonella typhimurium within desmoplastic tumors. BHs-ST represents a promising new tumor ECM–targeting strategy that may be instrumental in minimizing off-tumor toxicity while maximizing drug delivery into highly desmoplastic tumors.
Data from Novel Auristatins with High Bystander and Cytotoxic Activities in Drug Efflux–positive Tumor Models
Authors: Moquist Philip N. , Bovee Tim D. , Waight Andrew B. , Mitchell Jamie A. , Miyamoto Jamie B. , Mason Marsha L. , Emmerton Kim K. , Stevens Nicole , Balasubramanian Cindy , Simmons Jessica K. , Lyon Robert P. , Senter Peter D. , Doronina Svetlana O.
Published: March 2024

Abstract
Auristatins, a class of clinically validated anti-tubulin agents utilized as payloads in antibody–drug conjugates, are generally classified by their membrane permeability and the extent of cytotoxic bystander activity on neighboring cells after targeted delivery. The drugs typically fall within two categories: membrane permeable monomethyl auristatin E–type molecules with high bystander activities and susceptibility to efflux pumps, or charged and less permeable monomethyl auristatin F (MMAF) analogs with low bystander activities and resistance to efflux pumps. Herein, we report the development of novel auristatins that combine the attributes of each class by having both bystander activity and cytotoxicity on multidrug-resistant (MDR+) cell lines. Structure-based design focused on the hydrophobic functionalization of the N-terminal N-methylvaline of the MMAF scaffold to increase cell permeability. The resulting structure–activity relationships of the new auristatins demonstrate that optimization of hydrophobicity and structure can lead to highly active free drugs and antibody–drug conjugates with in vivo bystander activities.
Data from The Role of Specific ATP-Binding Cassette Transporters in the Acquired Resistance to Pyrrolobenzodiazepine Dimer–Containing Antibody–Drug Conjugates
Authors: Corbett Simon , Huang Shiran , Zammarchi Francesca , Howard Philip W. , van Berkel Patrick H. , Hartley John A.
Published: March 2024

Abstract
Antibody–drug conjugates (ADC) containing pyrrolobenzodiazepine (PBD) dimers are being evaluated clinically in both hematologic and solid tumors. These include ADCT-301 (camidanlumab tesirine) and ADCT-402 (loncastuximab tesirine) in pivotal phase II trials that contain the payload tesirine, which releases the PBD dimer warhead SG3199. An important consideration in future clinical development is acquired resistance. The aim was to generate and characterize PBD acquired resistant cell lines in both hematologic and solid tumor settings. Human Karpas-299 (ALCL) and NCI-N87 (gastric cancer) cells were incubated with increasing IC50 doses of ADC (targeting CD25 and HER2, respectively) or SG3199 in a pulsed manner until stable acquired resistance was established. The level of resistance achieved was approximately 3,000-fold for ADCT-301 and 3-fold for SG3199 in Karpas-299, and 8-fold for ADCT-502 and 4-fold for SG3199 in NCI-N87. Cross-resistance between ADC and SG3199, and with an alternative PBD-containing ADC or PBD dimer was observed. The acquired resistant lines produced fewer DNA interstrand cross-links, indicating an upstream mechanism of resistance. Loss of antibody binding or internalization was not observed. A human drug transporter PCR Array revealed several genes upregulated in all the resistant cell lines, including ABCG2 and ABCC2, but not ABCB1(MDR1). These findings were confirmed by RT-PCR and Western blot, and inhibitors and siRNA knockdown of ABCG2 and ABCC2 recovered drug sensitivity. These data show that acquired resistance to PBD-ADCs and SG3199 can involve specific ATP-binding cassette drug transporters. This has clinical implications as potential biomarkers of resistance and for the rational design of drug combinations.
Data from BRCA1/MAD2L1 Deficiency Disrupts the Spindle Assembly Checkpoint to Confer Vinorelbine Resistance in Mesothelioma
Authors: Busacca Sara , O'Regan Laura , Singh Anita , Sharkey Annabel J. , Dawson Alan G. , Dzialo Joanna , Parsons Aimee , Kumar Neelam , Schunselaar Laurel M. , Guppy Naomi , Nakas Apostolos , Sheaff Michael , Mansfield Aaron S. , Janes Sam M. , Baas Paul , Fry Andrew M. , Fennell Dean A.
Published: March 2024

Abstract
Mesothelioma is a universally lethal cancer lacking effective therapy. The spindle poison vinorelbine exhibits clinical activity in the relapsed setting, and in preclinical models requires BRCA1 to initiate apoptosis. However, the mechanisms underlying this regulation and the clinical implications have not been explored. Here, we show that BRCA1 silencing abrogated vinorelbine-induced cell-cycle arrest, recruitment of BUBR1 to kinetochores, and apoptosis. BRCA1 silencing led to codepletion of MAD2L1 at the mRNA and protein levels consistent with its status as a transcriptional target of BRCA1. Silencing of MAD2L1 phenocopied BRCA1 and was sufficient to confer resistance to vinorelbine. This was recapitulated in cell lines selected for resistance to vinorelbine, which acquired loss of both BRCA1 and MAD2L1 expression. Following ex vivo vinorelbine in 20 primary tumor explants, apoptotic response rate was 59% in BRCA1/MAD2L1-positive explants compared with 0% in BRCA1/MAD2L1-negative explants. In 48 patients, BRCA1 and/or MAD2L1 loss of expression was not prognostic; however, in a subset of patients treated with vinorelbine, survival was shorter for patients lacking BRCA1/MAD2L1 expression compared with double-positive patients (5.9 vs. 36.7 months, P = 0.03). Our data implicate BRCA1/MAD2L1 loss as a putative predictive marker of resistance to vinorelbine in mesothelioma and warrant prospective clinical evaluation.
Data from FGFR Inhibition Enhances Sensitivity to Radiation in Non–Small Cell Lung Cancer
Authors: SenthilKumar Gopika , Fisher Michael M. , Skiba Justin H. , Miller Margot C. , Brennan Sean R. , Kaushik Saakshi , Bradley Samantha T. , Longhurst Colin A. , Buehler Darya , Nickel Kwangok P. , Iyer Gopal , Kimple Randall J. , Baschnagel Andrew M.
Published: March 2024

Abstract
FGFRs are commonly altered in non–small cell lung cancer (NSCLC). FGFRs activate multiple pathways including RAS/RAF/MAPK, PI3K/AKT, and STAT, which may play a role in the cellular response to radiation. We investigated the effects of combining the selective FGFR 1-3 tyrosine kinase inhibitor AZD4547 with radiation in cell line and xenograft models of NSCLC. NSCLC cell lines were assessed with proliferation, clonogenic survival, apoptosis, autophagy, cell cycle, and DNA damage signaling and repair assays. In vivo xenografts and IHC were used to confirm in vitro results. NSCLC cell lines demonstrated varying degrees of FGFR protein and mRNA expression. In vitro clonogenic survival assays showed radiosensitization with AZD4547 in two NSCLC cell lines. In these two cell lines, an increase in apoptosis and autophagy was observed with combined radiation and AZD4547. The addition of AZD4547 to radiation did not significantly affect γH2AX foci formation. Enhanced xenograft tumor growth delay was observed with the combination of radiation and AZD4547 compared with radiation or drug alone. IHC results revealed inhibition of pMAPK and pS6 and demonstrated an increase in apoptosis in the radiation plus AZD4547 group. This study demonstrates that FGFR inhibition by AZD4547 enhances the response of radiation in FGFR-expressing NSCLC in vitro and in vivo model systems. These results support further investigation of combining FGFR inhibition with radiation as a clinical therapeutic strategy.
Data from Optimal Strategy and Benefit of Pulsed Therapy Depend On Tumor Heterogeneity and Aggressiveness at Time of Treatment Initiation
Authors: Mathur Deepti , Taylor Bradford P. , Chatila Walid K. , Scher Howard I. , Schultz Nikolaus , Razavi Pedram , Xavier Joao B.
Published: March 2024

Abstract
Therapeutic resistance is a fundamental obstacle in cancer treatment. Tumors that initially respond to treatment may have a preexisting resistant subclone or acquire resistance during treatment, making relapse theoretically inevitable. Here, we investigate treatment strategies that may delay relapse using mathematical modeling. We find that for a single-drug therapy, pulse treatment—short, elevated doses followed by a complete break from treatment—delays relapse compared with continuous treatment with the same total dose over a length of time. For tumors treated with more than one drug, continuous combination treatment is only sometimes better than sequential treatment, while pulsed combination treatment or simply alternating between the two therapies at defined intervals delays relapse the longest. These results are independent of the fitness cost or benefit of resistance, and are robust to noise. Machine-learning analysis of simulations shows that the initial tumor response and heterogeneity at the start of treatment suffice to determine the benefit of pulsed or alternating treatment strategies over continuous treatment. Analysis of eight tumor burden trajectories of breast cancer patients treated at Memorial Sloan Kettering Cancer Center shows the model can predict time to resistance using initial responses to treatment and estimated preexisting resistant populations. The model calculated that pulse treatment would delay relapse in all eight cases. Overall, our results support that pulsed treatments optimized by mathematical models could delay therapeutic resistance.
Data from Gene Expression Signature Correlates with Outcomes in Metastatic Renal Cell Carcinoma Patients Treated with Everolimus Alone or with a Vascular Disrupting Agent
Authors: Yang Eddy S. , Nassar Amin H. , Adib Elio , Jegede Opeyemi A. , Alaiwi Sarah Abou , Manna Deborah L. Della , Braun David A. , Zarei Mahsa , Du Heng , Pal Sumanta K. , Naik Gurudatta , Sonpavde Guru P.
Published: March 2024

Abstract
Everolimus monotherapy use for metastatic renal cell carcinoma (mRCC) has diminished due to recent approvals of immune checkpoint and VEGF inhibitors. We hypothesized that gene expression associated with everolimus benefit may provide rationale to select appropriate patients. To address this hypothesis, tumors from a phase I/II trial that compared everolimus alone or with BNC105P, a vascular disrupting agent, were profiled using Nanostring as a discovery cohort. A phase III trial (CheckMate 025) was used for validation. Clinical benefit (CB) was defined as response or stable disease for ≥6 months. A propensity score covariate adjustment was used, and model discrimination performance was assessed using the area under the ROC curve (AUC). In a discovery cohort of 82 patients, 35 (43%) were treated with everolimus alone and 47 (57%) received everolimus + BNC105P. Median PFS (mPFS) was 4.9 (95% CI, 2.8–6.2) months. A four-gene signature (ASXL1, DUSP6, ERCC2, and HSPA6) correlated with CB with everolimus ± BNC105P [AUC, 86.9% (95% CI, 79.2–94.7)]. This was validated in 130 patients from CheckMate 025 treated with everolimus [AUC, 60.2% (95% CI, 49.7–70.7)]. Among 43 patients (52.4%) with low expression of an 18-gene signature, everolimus + BNC105P was associated with significantly longer mPFS compared with everolimus alone (10.4 vs. 6.9 months; HR, 0.49; 95% CI, 0.24–1.002; P = 0.047). These signatures warrant further validation to select patients who may benefit from everolimus alone or with a vascular disrupting agent.
Data from ASN004, A 5T4-targeting scFv-Fc Antibody–Drug Conjugate with High Drug-to-Antibody Ratio, Induces Complete and Durable Tumor Regressions in Preclinical Models
Authors: Smith Roger A. , Zammit David J. , Damle Nitin K. , Usansky Helen , Reddy Sanjeeva P. , Lin Jun-Hsiang , Mistry Mahesh , Rao Niranjan S. , Denis Louis J. , Gupta Sandeep
Published: March 2024

Abstract
The 5T4 oncofetal antigen (trophoblast glycoprotein) is expressed in a wide range of malignant tumors but shows very limited expression in normal adult tissues. ASN004 is a 5T4-targeted antibody–drug conjugate (ADC) that incorporates a novel single-chain Fv-Fc antibody and Dolaflexin drug-linker technology, with an Auristatin F hydroxypropylamide payload drug-to-antibody ratio of approximately 10–12. The pharmacology, toxicology, and pharmacokinetic properties of ASN004 and its components were investigated in vitro and in vivo. ASN004 showed high affinity for the 5T4 antigen and was selectively bound to and internalized into 5T4-expressing tumor cells, and potent cytotoxicity was demonstrated for a diverse panel of solid tumor cell lines. ASN004 induced complete and durable tumor regression in multiple tumor xenograft models, derived from human lung, breast, cervical, and gastric tumor cell lines having a wide range of 5T4 expression levels. A single dose of ASN004, as low as 1 mg/kg i.v., achieved complete tumor regression leading to tumor-free survivors in the A431 cervical cancer model. In head-to-head studies, superior activity of ASN004 was demonstrated against trastuzumab-DM1, in a low-5T4/high-HER2 expressing gastric tumor model, and 10-fold greater potency was found for ASN004 against the 5T4-targeted ADC PF-06263507 in a lung tumor model. In marmoset monkeys, ASN004 was well tolerated at doses up to 1.5 mg/kg Q3W i.v., and showed dose-dependent exposure, linear pharmacokinetics, and markedly low exposure of free payload drug. Taken together, these findings identify ASN004 as a promising new ADC therapeutic for clinical evaluation in a broad range of solid tumor types.
Data from Critical Role for Cap-Independent c-MYC Translation in Progression of Multiple Myeloma
Authors: Shi Yijiang , Sun Fumou , Cheng Yan , Holmes Brent , Dhakal Binod , Gera Joseph F. , Janz Siegfried , Lichtenstein Alan
Published: March 2024

Abstract
Dysregulated c-myc is a determinant of multiple myeloma progression. Translation of c-myc can be achieved by an mTOR-mediated, cap-dependent mechanism or a cap-independent mechanism where a sequence in the 5′UTR of mRNA, termed the internal ribosome entry site (IRES), recruits the 40S ribosomal subunit. This mechanism requires the RNA-binding factor hnRNP A1 (A1) and becomes critical when cap-dependent translation is inhibited during endoplasmic reticulum (ER) stress. Thus, we studied the role of A1 and the myc IRES in myeloma biology. A1 expression correlated with enhanced c-myc expression in patient samples. Expression of A1 in multiple myeloma lines was mediated by c-myc itself, suggesting a positive feedback circuit where myc induces A1 and A1 enhances myc translation. We then deleted the A1 gene in a myc-driven murine myeloma model. A1-deleted multiple myeloma cells demonstrated downregulated myc expression and were inhibited in their growth in vivo. Decreased myc expression was due to reduced translational efficiency and depressed IRES activity. We also studied the J007 inhibitor, which prevents A1's interaction with the myc IRES. J007 inhibited myc translation and IRES activity and diminished myc expression in murine and human multiple myeloma lines as well as primary samples. J007 also inhibited tumor outgrowth in mice after subcutaneous or intravenous challenge and prevented osteolytic bone disease. When c-myc was ectopically reexpressed in A1-deleted multiple myeloma cells, tumor growth was reestablished. These results support the critical role of A1-dependent myc IRES translation in myeloma.
Data from Development of Highly Effective Anti-Mesothelin hYP218 Chimeric Antigen Receptor T Cells With Increased Tumor Infiltration and Persistence for Treating Solid Tumors
Authors: Tomar Sakshi , Zhang Jingli , Khanal Manakamana , Hong Jessica , Venugopalan Abhilash , Jiang Qun , Sengupta Manjistha , Miettinen Markku , Li Nan , Pastan Ira , Ho Mitchell , Hassan Raffit
Published: March 2024

Abstract
Mesothelin targeting CAR T cells have limited activity in patients. In this study, we sought to determine if efficacy of anti-mesothelin CAR T cells is dependent on the mesothelin epitopes that are recognized by them. To do so, we developed hYP218 (against membrane-proximal epitope) and SS1 (against membrane-distal epitope) CAR T cells. Their efficacy was assessed in vitro using mesothelin-positive tumor cell lines and in vivo in NSG mice with mesothelin-expressing ovarian cancer (OVCAR-8), pancreatic cancer (KLM-1), and mesothelioma patient-derived (NCI-Meso63) tumor xenografts. Persistence and tumor infiltration of CAR T cells was determined using flow cytometry. hYP218 CAR T cells killed cancer cells more efficiently than SS1 CAR T cells, with a two- to fourfold lower ET50 value (effector-to-target ratio for 50% killing of tumor cells). In mice with established tumors, single intravenous administration of hYP218 CAR T cells lead to improved tumor response and survival compared with SS1 CAR T cells, with complete regression of OVCAR-8 and NCI-Meso63 tumors. Compared with SS1 CAR T cells, there was increased peripheral blood expansion, persistence, and tumor infiltration of hYP218 CAR T cells in the KLM-1 tumor model. Persistence of hYP218 CAR T cells in treated mice led to antitumor immunity when rechallenged with KLM-1 tumor cells. Our results show that hYP218 CAR T cells, targeting mesothelin epitope close to cell membrane, are very effective against mesothelin-positive tumors and are associated with increased persistence and tumor infiltration. These results support its clinical development to treat patients with mesothelin-expressing cancers.
Data from NAP1051, a Lipoxin A4 Biomimetic Analogue, Demonstrates Antitumor Activity Against the Tumor Microenvironment
Authors: Dong Tiange , Dave Priyal , Yoo EunJeong , Ebright Brandon , Ahluwalia Kabir , Zhou Eugene , Asante Isaac , Salimova Malika , Pei Hua , Lin Tracey , Mead Andrew , Li Zeyang , Humayun Mark , Petasis Nicos A. , Epstein Alan L. , Louie Stan G.
Published: March 2024

Abstract
Resolving tumor-associated inflammation in the tumor microenvironment (TME) may promote antitumor effects. Lipoxin A4 (LXA4) is a short-lived endogenous bioactive lipid with potent anti-inflammatory and pro-resolving properties. Here, a biomimetic of LXA4, NAP1051, was shown to have LXA4-like in vitro properties and antitumor activity in colorectal cancer xenograft models. NAP1051 inhibited neutrophil chemotaxis toward fMLP and dose-dependently promoted dTHP-1 efferocytosis which was equipotent to aspirin-triggered lipoxin A4 (ATLA). In dTHP-1 cells, NAP1051 induced strong phosphorylation on ERK1/2 and AKT similar to formyl peptide receptor 2 (FPR2/ALX) agonists. In two mouse xenograft colorectal cancer models, NAP1051 significantly inhibited tumor growth when given orally at 4.8 to 5 mg/kg/day. Flow cytometric analyses showed that NAP1051 reduced splenic and intratumoral neutrophil and myeloid-derived suppressor cell populations, which correlated to the antitumor effect. In addition, NAP1051 reduced NETosis in the TME while stimulating T-cell recruitment. Overall, these results show that NAP1051 possesses key lipoxin-like properties and has antitumor activity against colorectal cancer via modulation of neutrophils and NETosis in the TME.
Data from Anti-Glypican-1 Antibody–drug Conjugate as Potential Therapy Against Tumor Cells and Tumor Vasculature for Glypican-1–Positive Cholangiocarcinoma
Authors: Yokota Keiichiro , Serada Satoshi , Tsujii Shigehiro , Toya Keisuke , Takahashi Tsuyoshi , Matsunaga Takashi , Fujimoto Minoru , Uemura Sunao , Namikawa Tsutomu , Murakami Ichiro , Kobayashi Shogo , Eguchi Hidetoshi , Doki Yuichiro , Hanazaki Kazuhiro , Naka Tetsuji
Published: March 2024

Abstract
Cholangiocarcinoma is a highly malignant cancer. Many patients need systemic chemotherapy to prevent tumor development and recurrence; however, their prognosis is poor due to the lack of effective therapy. Therefore, a new treatment option is urgently required. We recently identified glypican-1 (GPC1) as a novel cancer antigen of esophageal squamous cell carcinoma. We also demonstrated the efficacy and safety of GPC1-targeted ADC (GPC1–ADC) conjugating anti-GPC1 mAb possessing high internalization activity with monomethyl auristatin F (MMAF), which is a potent tubulin polymerizing inhibitor. In this study, we confirmed that GPC1 was highly expressed in cholangiocarcinoma cells and tissues. IHC analysis of 49 extrahepatic cholangiocarcinoma patient tumor specimens revealed high expression of GPC1 in 47% of patients. These patients demonstrated significantly poorer prognosis compared with the low-expression group in terms of disease-free survival and overall survival (P < 0.05). GPC1 was also expressed in tumor vessels of cholangiocarcinoma, but not on the vessels of nontumor tissues. MMAF-conjugated GPC1–ADC showed potent tumor growth inhibition against GPC1-positive cholangiocarcinoma cells in vitro and in vivo. In a GPC1 knockout xenograft model, GPC1–ADC partially inhibited tumor growth. Vascular endothelial cells in tumor tissues of GPC1-negative xenograft mice expressed GPC1 and were arrested in the G2–M phase of cell cycle by GPC1–ADC. GPC1–ADC exhibits direct as well as indirect antitumor effects via inhibition of tumor angiogenesis. Our preclinical data highlight GPC1–ADC as a promising therapy for GPC1-positive cholangiocarcinoma.
Data from Preclinical Studies of OBI-999: A Novel Globo H–Targeting Antibody–Drug Conjugate
Authors: Yang Ming-Chen , Shia Chi-Sheng , Li Wan-Fen , Wang Chun-Chung , Chen I-Ju , Huang Teng-Yi , Chen Yu-Jung , Chang Hui-Wen , Lu Chi-Huan , Wu Yueh-Chin , Wang Nan-Hsuan , Lai Jiann-Shiun , Yu Cheng-Der , Lai Ming-Tain
Published: March 2024

Abstract
Globo H (GH), a hexasaccharide, is expressed at low levels in normal tissues but is highly expressed in multiple cancer types, rendering it a promising target for cancer immunotherapy. OBI-999, a novel antibody–drug conjugate, is derived from a conjugation of a GH-specific mAb with a monomethyl auristatin E (MMAE) payload through a site-specific ThioBridge and a cleavable linker. OBI-999 high homogeneity with a drug-to-antibody ratio of 4 (>95%) was achieved using ThioBridge. OBI-999 displayed GH-dependent cellular internalization and trafficked to endosome and lysosome within 1 and 5 hours, respectively. Furthermore, OBI-999 showed low nanomolar cytotoxicity in the assay with high GH expression on tumor cells and exhibited a bystander killing effect on tumor cells with minimal GH expression. Tissue distribution indicated that OBI-999 and free MMAE gradually accumulated in the tumor, reaching maximum level at 168 hours after treatment, whereas OBI-999 and free MMAE decreased quickly at 4 hours after treatment in normal organs. Maximum MMAE level in the tumor was 16-fold higher than in serum, suggesting that OBI-999 is stable during circulation and MMAE is selectively released in the tumor. Excellent tumor growth inhibition of OBI-999 was demonstrated in breast, gastric, and pancreatic cancer xenograft or lung patient–derived xenograft models in a dose-dependent manner. The highest nonseverely toxic dose in cynomolgus monkeys is 10 mg/kg determined by a 3-week repeated-dose toxicology study demonstrating an acceptable safety margin. Taken together, these results support further clinical development of OBI-999, which is currently in a phase I/II clinical study in multiple solid tumors (NCT04084366). OBI-999, the first GH-targeting ADC, displayed excellent tumor inhibition in animal models across multiple cancer types, including breast, gastric, pancreatic, and lung cancers, warranting further investigation in the treatment of solid tumors.
Data from Cellular-Resolution Imaging of Bystander Payload Tissue Penetration from Antibody-Drug Conjugates
Authors: Khera Eshita , Dong Shujun , Huang Haolong , de Bever Laureen , van Delft Floris L. , Thurber Greg M.
Published: March 2024

Abstract
After several notable clinical failures in early generations, antibody–drug conjugates (ADC) have made significant gains with seven new FDA approvals within the last 3 years. These successes have been driven by a shift towards mechanistically informed ADC design, where the payload, linker, drug-to-antibody ratio, and conjugation are increasingly tailored to a specific target and clinical indication. However, fundamental aspects needed for design, such as payload distribution, remain incompletely understood. Payloads are often classified as “bystander” or “nonbystander” depending on their ability to diffuse out of targeted cells into adjacent cells that may be antigen-negative or more distant from tumor vessels, helping to overcome heterogeneous distribution. Seven of the 11 FDA-approved ADCs employ these bystander payloads, but the depth of penetration and cytotoxic effects as a function of physicochemical properties and mechanism of action have not been fully characterized. Here, we utilized tumor spheroids and pharmacodynamic marker staining to quantify tissue penetration of the three major classes of agents: microtubule inhibitors, DNA-damaging agents, and topoisomerase inhibitors. PAMPA data and coculture assays were performed to compare with the 3D tissue culture data. The results demonstrate a spectrum in bystander potential and tissue penetration depending on the physicochemical properties and potency of the payload. Generally, directly targeted cells show a greater response even with bystander payloads, consistent with the benefit of deeper ADC tissue penetration. These results are compared with computational simulations to help scale the data from in vitro and preclinical animal models to the clinic.
Data from Neoadjuvant Radiation Therapy and Surgery Improves Metastasis-Free Survival over Surgery Alone in a Primary Mouse Model of Soft Tissue Sarcoma
Authors: Patel Rutulkumar , Mowery Yvonne M. , Qi Yi , Bassil Alex M. , Holbrook Matt , Xu Eric S. , Hong Cierra S. , Himes Jonathon E. , Williams Nerissa T. , Everitt Jeffrey , Ma Yan , Luo Lixia , Selitsky Sara R. , Modliszewski Jennifer L. , Gao Junheng , Jung Sin-Ho , Kirsch David G. , Badea Cristian T.
Published: March 2024

Abstract
This study aims to investigate whether adding neoadjuvant radiotherapy (RT), anti-programmed cell death protein-1 (PD-1) antibody (anti–PD-1), or RT + anti-PD-1 to surgical resection improves disease-free survival for mice with soft tissue sarcomas (STS). We generated a high mutational load primary mouse model of STS by intramuscular injection of adenovirus expressing Cas9 and guide RNA targeting Trp53 and intramuscular injection of 3-methylcholanthrene (MCA) into the gastrocnemius muscle of wild-type mice (p53/MCA model). We randomized tumor-bearing mice to receive isotype control or anti–PD-1 antibody with or without radiotherapy (20 Gy), followed by hind limb amputation. We used micro-CT to detect lung metastases with high spatial resolution, which was confirmed by histology. We investigated whether sarcoma metastasis was regulated by immunosurveillance by lymphocytes or tumor cell-intrinsic mechanisms. Compared with surgery with isotype control antibody, the combination of anti–PD-1, radiotherapy, and surgery improved local recurrence-free survival (P = 0.035) and disease-free survival (P = 0.005), but not metastasis-free survival. Mice treated with radiotherapy, but not anti–PD-1, showed significantly improved local recurrence-free survival and metastasis-free survival over surgery alone (P = 0.043 and P = 0.007, respectively). The overall metastasis rate was low (∼12%) in the p53/MCA sarcoma model, which limited the power to detect further improvement in metastasis-free survival with addition of anti–PD-1 therapy. Tail vein injections of sarcoma cells into immunocompetent mice suggested that impaired metastasis was due to inability of sarcoma cells to grow in the lungs rather than a consequence of immunosurveillance. In conclusion, neoadjuvant radiotherapy improves metastasis-free survival after surgery in a primary model of STS.
Data from Antibody–Drug Conjugate Sacituzumab Govitecan Drives Efficient Tissue Penetration and Rapid Intracellular Drug Release
Authors: Kopp Anna , Hofsess Scott , Cardillo Thomas M. , Govindan Serengulam V. , Donnell Jennifer , Thurber Greg M.
Published: March 2024

Abstract
Antibody–drug conjugates (ADC) are a rapidly growing class of targeted cancer treatments, but the field has experienced significant challenges from their complex design. This study examined the multiscale distribution of sacituzumab govitecan (SG; Trodelvy), a recently clinically approved ADC, to clarify the mechanism(s) of efficacy given its unique design strategy. We employed a multiscale quantitative pharmacokinetic approach, including near-infrared fluorescence imaging, single-cell flow cytometry measurements, payload distribution via γH2AX pharmacodynamic staining, and a novel dual-labeled fluorescent technique to track the ADC and payload in a high trophoblast cell-surface antigen 2 expression xenograft model of gastric cancer (NCI-N87). We found that rapid release of the SN-38 payload from the hydrolysable linker inside cells imparts more DNA damage in vitro and in vivo than an ADC with a more stable enzyme cleavable linker. With SG, little to no extracellular payload release in the tumor was observed using a dual-labeled fluorescence technique, although bystander effects were detected. The high dosing regimen allowed the clinical dose to reach the majority of cancer cells, which has been linked to improved efficacy. In addition, the impact of multiple doses (day 1 and day 8) of a 21-day cycle was found to further improve tissue penetration despite not changing tumor uptake [percent injected dose per gram (%ID/g)] of the ADC. These results show increased ADC efficacy with SG can be attributed to efficient tumor penetration and intracellular linker cleavage after ADC internalization. This quantitative approach to study multiscale delivery can be used to inform the design of next-generation ADCs and prodrugs for other targets.
Data from Combination of Ribociclib and Gemcitabine for the Treatment of Medulloblastoma
Authors: Pribnow Allison , Jonchere Barbara , Liu Jingjing , Smith Kyle S. , Campagne Olivia , Xu Ke , Robinson Sarah , Patel Yogesh , Onar-Thomas Arzu , Wu Gang , Stewart Clinton F. , Northcott Paul A. , Yu Jiyang , Robinson Giles W. , Roussel Martine F.
Published: March 2024

Abstract
Group3 (G3) medulloblastoma (MB) is one of the deadliest forms of the disease for which novel treatment is desperately needed. Here we evaluate ribociclib, a highly selective CDK4/6 inhibitor, with gemcitabine in mouse and human G3MBs. Ribociclib central nervous system (CNS) penetration was assessed by in vivo microdialysis and by IHC and gene expression studies and found to be CNS-penetrant. Tumors from mice treated with short term oral ribociclib displayed inhibited RB phosphorylation, downregulated E2F target genes, and decreased proliferation. Survival studies to determine the efficacy of ribociclib and gemcitabine combination were performed on mice intracranially implanted with luciferase-labeled mouse and human G3MBs. Treatment of mice with the combination of ribociclib and gemcitabine was well tolerated, slowed tumor progression and metastatic spread, and increased survival. Expression-based gene activity and cell state analysis investigated the effects of the combination after short- and long-term treatments. Molecular analysis of treated versus untreated tumors showed a significant decrease in the activity and expression of genes involved in cell-cycle progression and DNA damage response, and an increase in the activity and expression of genes implicated in neuronal identity and neuronal differentiation. Our findings in both mouse and human patient-derived orthotopic xenograft models suggest that ribociclib and gemcitabine combination therapy warrants further investigation as a treatment strategy for children with G3MB.
Data from Inhibition of Integrin αVβ3 Signaling Improves the Antineoplastic Effect of Bexarotene in Cutaneous T-Cell Lymphoma
Authors: Cayrol Florencia , Revuelta Maria V. , Debernardi Mercedes , Paulazo Alejandra , Phillip Jude M. , Zamponi Nahuel , Sterle Helena , Díaz Flaqué María C. , Magro Cynthia , Marullo Rossella , Mulvey Erin , Ruan Jia , Cremaschi Graciela A. , Cerchietti Leandro
Published: March 2024

Abstract
Bexarotene is a specific retinoid X receptor agonist that has been used for the treatment of cutaneous T-cell lymphoma (CTCL). Because bexarotene causes hypothyroidism, it requires the administration of levothyroxine. However, levothyroxine, in addition to its ubiquitous nuclear receptors, can activate the αVβ3 integrin that is overexpressed in CTCL, potentially interfering the antineoplastic effect of bexarotene. We thus investigated the biological effect of levothyroxine in relation to bexarotene treatment. Although in isolated CTCL cells levothyroxine decreased, in an αVβ3-dependent manner, the antineoplastic effect of bexarotene, levothyroxine supplementation in preclinical models was necessary to avoid suppression of lymphoma immunity. Accordingly, selective genetic and pharmacologic inhibition of integrin αVβ3 improved the antineoplastic effect of bexarotene plus levothyroxine replacement while maintaining lymphoma immunity. Our results provide a mechanistic rationale for clinical testing of integrin αVβ3 inhibitors as part of CTCL regimens based on bexarotene administration.
Teaser:
Inhibiting αVβ3 integrin improves the antineoplastic effect of bexarotene while maintaining lymphoma immunity.
Data from Discovery and Characterization of a Novel Aryl Hydrocarbon Receptor Inhibitor, IK-175, and Its Inhibitory Activity on Tumor Immune Suppression
Authors: McGovern Karen , Castro Alfredo C. , Cavanaugh Jill , Coma Silvia , Walsh Meghan , Tchaicha Jeremy , Syed Sakeena , Natarajan Prabitha , Manfredi Mark , Zhang Xiaoyan M. , Ecsedy Jeffrey
Published: March 2024

Abstract
Aryl hydrocarbon receptor (AHR) is a transcription factor that regulates the activity of multiple innate and adaptive immune cells subsequent to binding to numerous endogenous and exogenous ligands. For example, AHR is activated by the metabolite kynurenine, which is secreted into the tumor microenvironment by cancer cells leading to broad immunosuppression. Therefore, AHR inhibition provides a novel and ideal approach to stimulate immune-mediated recognition and subsequent eradication of tumor cells. We report here the discovery and characterization of IK-175, a novel, potent and selective AHR antagonist with favorable ADME and pharmacokinetic profiles in preclinical species. IK-175 inhibits AHR activity in experimental systems derived from multiple species including mouse, rat, monkey, and humans. In human primary immune cells, IK-175 decreased AHR target gene expression and anti-inflammatory cytokine release and increased proinflammatory cytokine release. Moreover, IK-175 led to a decrease in suppressive IL17A–, IL-22+ expressing T cells in a Th17 differentiation assay. IK-175 dose dependently blocks ligand-stimulated AHR activation of Cyp1a1 transcription in mouse liver and spleen, demonstrating on-target in vivo activity. IK-175 increases proinflammatory phenotype of the tumor microenvironment in mouse syngeneic tumors and in adjacent tumor-draining lymph nodes. As a monotherapy and combined with an anti-PD-1 antibody, IK-175 demonstrates antitumor activity in syngeneic mouse models of colorectal cancer and melanoma. IK-175 also demonstrates antitumor activity combined with liposomal doxorubicin in syngeneic mouse tumors. These studies provide rationale for targeting AHR in patients with cancer. IK-175 is being evaluated in a phase I clinical trial in patients with advanced solid tumors.
Data from Luxeptinib (CG-806) Targets FLT3 and Clusters of Kinases Operative in Acute Myeloid Leukemia
Authors: Rice William G. , Howell Stephen B. , Zhang Hongying , Rastgoo Nasrin , Local Andrea , Kurtz Stephen E. , Lo Pierrette , Bottomly Daniel , Wilmot Beth , McWeeney Shannon K. , Druker Brian J. , Tyner Jeffrey W.
Published: March 2024

Abstract
Luxeptinib (CG-806) simultaneously targets FLT3 and select other kinase pathways operative in myeloid malignancies. We investigated the range of kinases it inhibits, its cytotoxicity landscape ex vivo with acute myeloid leukemia (AML) patient samples, and its efficacy in xenograft models. Luxeptinib inhibits wild-type (WT) and many of the clinically relevant mutant forms of FLT3 at low nanomolar concentrations. It is a more potent inhibitor of the activity of FLT3—internal tandem duplication, FLT3 kinase domain and gatekeeper mutants than against WT FLT3. Broad kinase screens disclosed that it also inhibits other kinases that can drive oncogenic signaling and rescue pathways, but spares kinases known to be associated with clinical toxicity. In vitro profiling of luxeptinib against 186 AML fresh patient samples demonstrated greater potency relative to other FLT3 inhibitors, including cases with mutations in FLT3, isocitrate dehydrogenase-1/2, ASXL1, NPM1, SRSF2, TP53, or RAS, and activity was documented in a xenograft AML model. Luxeptinib administered continuously orally every 12 hours at a dose that yielded a mean Cmin plasma concentration of 1.0 ± 0.3 μmol/L (SEM) demonstrated strong antitumor activity but no myelosuppression or evidence of tissue damage in mice or dogs in acute toxicology studies. On the basis of these studies, luxeptinib was advanced into a phase I trial for patients with AML and myelodysplastic/myeloproliferative neoplasms.
Data from External Validation of Association of Baseline Circulating Tumor Cell Counts with Survival Outcomes in Men with Metastatic Castration-Sensitive Prostate Cancer
Authors: Swami Umang , Sayegh Nicolas , Jo Yeonjung , Haaland Benjamin , McFarland Taylor Ryan , Nussenzveig Roberto H. , Goel Divyam , Sirohi Deepika , Hahn Andrew W. , Maughan Benjamin L. , Goldkorn Amir , Agarwal Neeraj
Published: March 2024

Abstract
Approximately 20% of men with metastatic castration-sensitive prostate cancer (mCSPC) progress within 1 year of treatment, and biomarkers to identify them up front are lacking. In a randomized phase III trial in men with mCSPC (SWOG S1216), higher baseline circulating tumor cells (CTCs) were prognostic of inferior outcomes. We aimed to validate these findings and interrogate corresponding tumor genomic profiles. Consecutively seen men with newly diagnosed mCSPC undergoing systemic therapy and baseline CTC enumeration by CellSearch assay were included. Gene alterations were determined by comprehensive genomic profiling of tumor tissue by Clinical Laboratory Improvement Amendments—certified lab. The relationship between categorized CTC counts and both progression-free survival (PFS) and overall survival (OS) was assessed in the context of Cox proportional hazards models, both unadjusted and adjusted for age, Gleason score, PSA at androgen-deprivation therapy initiation, disease volume, de novo status, treatment intensification, and number of altered genes. Overall, 103 patients were included in the analysis. On multivariate analysis high CTCs (≥ 5 vs. 0) were associated with poorer PFS [HR, 4.52; 95% confidence interval (CI), 1.84–11.11; P = 0.001) and OS (HR, 3.59; 95% CI, 0.95–13.57; P = 0.060). Patients with higher CTC counts had a greater number of altered genes and total number of alterations (all P < 0.02). In this article, for the first time, we externally validate the association of higher CTC counts with inferior survival outcomes in men with mCSPC and show a distinct associated tumor genomic landscape. These findings may improve prognostication, patient counseling, and treatment selection in men with mCSPC.
Data from IDO1 Is a Therapeutic Target for Pancreatic Cancer–Associated Depression
Authors: Hue Jonathan J. , Graor Hallie J. , Zarei Mehrdad , Katayama Erryk S. , Ji Karen , Hajihassani Omid , Loftus Alexander W. , Vaziri-Gohar Ali , Winter Jordan M.
Published: March 2024

Abstract
Metabolites of tryptophan degradation are known to alter mood. Their effects have only been superficially examined in the context of pancreatic cancer. Herein, we study the role of indoleamine 2,3-dioxygenase 1 (IDO1), an enzyme important in the conversion of tryptophan to kynurenine, in a murine model of pancreatic cancer–associated depression. Behavioral tests (open field, forced swim, tail suspension, and elevated plus maze) and biochemical assays (LC-MS metabolomics) were used to characterize a depressive-phenotype in tumor-bearing mice (relative to non–tumor-bearing mice). In addition, we determine whether pharmacologic blockade of IDO1 affects mood in tumor-bearing mice. Immunocompetent mice bearing orthotopic pancreatic tumors exhibit depressive-like behavior relative to non–tumor-bearing mice. Pancreatic tumors strongly express IDO1. Consequently, serum kynurenine levels in tumor-bearing mice are elevated relative to non–tumor-bearing mice. Tumor-bearing mice treated with epacadostat, an IDO1 inhibitor, exhibited improved mood relative to mice receiving vehicle. There was a 95% reduction in serum kynurenine levels in mice receiving epacadostat relative to mice treated with vehicle. As confirmatory evidence of on-target activity, tumors of mice treated with epacadostat exhibited a compensatory increase in IDO1 protein levels. Escitalopram, an approved antidepressant, was ineffective at improving mood in tumor-bearing mice as measured by behavioral assays and did not affect kynurenine levels. Neither epacadostat, nor escitalopram, affected overall survival relative to vehicle. Mice with pancreatic cancer exhibit depressive-like behavior. Epacadostat was effective as an antidepressant for pancreatic cancer–associated depression in mice. These data offer a rationale to consider IDO1 inhibition as a therapeutic strategy to mitigate depressive symptoms in patients with pancreatic cancer.
Data from IGF1R Inhibition Enhances the Therapeutic Effects of Gq/11 Inhibition in Metastatic Uveal Melanoma Progression
Authors: Lapadula Dominic , Lam Bao , Terai Mizue , Sugase Takahito , Tanaka Ryota , Farias Eduardo , Kadamb Rama , Lopez-Anton Melisa , Heine Christian C. , Modasia Bhavik , Aguirre-Ghiso Julio A. , Aplin Andrew E. , Sato Takami , Benovic Jeffrey L.
Published: March 2024

Abstract
Uveal melanoma (UM) is the most common intraocular tumor in adults, and up to 50% of patients develop metastatic disease, which remains uncurable. Because patients with metastatic UM have an average survival of less than 1 year after diagnosis, there is an urgent need to develop new treatment strategies. Although activating mutations in Gαq or Gα11 proteins are major drivers of pathogenesis, the therapeutic intervention of downstream Gαq/11 targets has been unsuccessful in treating UM, possibly due to alternative signaling pathways and/or resistance mechanisms. Activation of the insulin-like growth factor 1 (IGF1) signaling pathway promotes cell growth, metastasis, and drug resistance in many types of cancers, including UM, where expression of the IGF1 receptor (IGF1R) correlates with a poor prognosis. In this article, we show that direct inhibition of Gαq/11 by the cyclic depsipeptide YM-254890 in combination with inhibition of IGF1R by linsitinib cooperatively inhibits downstream signaling and proliferation of UM cells. We further demonstrate that a 2-week combination treatment of 0.3 to 0.4 mg/kg of YM-254890 administered by intraperitoneal injection and 25 to 40 mg/kg linsitinib administered by oral gavage effectively inhibits the growth of metastatic UM tumors in immunodeficient NOD scid gamma (NSG) mice and identifies the IGF1 pathway as a potential resistance mechanism in response to Gαq/11 inhibition in UM. These data suggest that the combination of Gαq/11 and IGF1R inhibition provides a promising therapeutic strategy to treat metastatic UM.
Data from DNA Damage-Inducible Gene, Reprimo Functions as a Tumor Suppressor and Is Suppressed by Promoter Methylation in Gastric Cancer
Authors: Ooki Akira , Yamashita Keishi , Yamaguchi Kensei , Mondal Anupom , Nishimiya Hiroshi , Watanabe Masahiko
Published: March 2024

Abstract
In several types of human cancer, the gene expression of Reprimo, a highly glycosylated protein, is frequently silenced via methylation of its promoter. The aim of this study was to characterize the epigenetic inactivation of Reprimo and its biologic function and clinical relevance in gastric cancer. The correlation between Reprimo methylation and clinical relevance was assessed in 83 primary human gastric cancer tissues. The effects of Reprimo expression were also examined using in vitro and in vivo assays. Reprimo methylation was cancer specific and frequently observed. In two gastric cancer cell lines without Reprimo methylation, we observed faint or weak Reprimo expression under normal conditions and high expression under DNA-damaging conditions. In four gastric cancer cell lines with Reprimo methylation, however, Reprimo expression remained faint even under DNA-damaging conditions, with expression being restored in combination with agents that induce demethylation. Enforced Reprimo expression robustly inhibited cell proliferation and anchorage-independent colony formation and enhanced DNA damage-induced apoptosis. Inverse effects were observed via siRNA-mediated knockdown of endogenous Reprimo. Reprimo expression inhibited tumorigenesis in vivo. Reprimo methylation was also associated with a poor response in patients with gastric cancer treated with chemotherapy (P¼ 0.028), and a poor prognosis in patients with advanced gastric cancer (P¼ 0.03). In conclusion, Reprimo expression is normally induced in response to DNA damage, acting as a novel tumor suppressor in gastric cancer. However, Reprimo methylation abrogates its expression and effects. The clinical assessment of Reprimo promoter methylation may serve not only as a predictive marker for chemotherapy, but also as a marker for tumor aggressiveness. Mol Cancer Res; 11(11); 1362–74. ©2013 AACR.