bluuebunny/crossref_metadata_2025 huggingface dataset:
Data from Enzalutamide-Induced Feed-Forward Signaling Loop Promotes Therapy-Resistant Prostate Cancer Growth Providing an Exploitable Molecular Target for Jak2 Inhibitors
Authors: Udhane Vindhya , Maranto Cristina , Hoang David T. , Gu Lei , Erickson Andrew , Devi Savita , Talati Pooja G. , Banerjee Anjishnu , Iczkowski Kenneth A. , Jacobsohn Kenneth , See William A. , Mirtti Tuomas , Kilari Deepak , Nevalainen Marja T.
Published: April 2023

Abstract
The second-generation antiandrogen, enzalutamide, is approved for castrate-resistant prostate cancer (CRPC) and targets androgen receptor (AR) activity in CRPC. Despite initial clinical activity, acquired resistance to enzalutamide arises rapidly and most patients develop terminal disease. Previous work has established Stat5 as a potent inducer of prostate cancer growth. Here, we investigated the significance of Jak2–Stat5 signaling in resistance of prostate cancer to enzalutamide. The levels of Jak2 and Stat5 mRNA, proteins and activation were evaluated in prostate cancer cells, xenograft tumors, and clinical prostate cancers before and after enzalutamide therapy. Jak2 and Stat5 were suppressed by genetic knockdown using lentiviral shRNA or pharmacologic inhibitors. Responsiveness of primary and enzalutamide-resistant prostate cancer to pharmacologic inhibitors of Jak2–Stat5 signaling was assessed in vivo in mice bearing prostate cancer xenograft tumors. Patient-derived prostate cancers were tested for responsiveness to Stat5 blockade as second-line treatment after enzalutamide ex vivo in tumor explant cultures. Enzalutamide-liganded AR induces sustained Jak2–Stat5 phosphorylation in prostate cancer leading to the formation of a positive feed-forward loop, where activated Stat5, in turn, induces Jak2 mRNA and protein levels contributing to further Jak2 activation. Mechanistically, enzalutamide-liganded AR induced Jak2 phosphorylation through a process involving Jak2-specific phosphatases. Stat5 promoted prostate cancer growth during enzalutamide treatment. Jak2–Stat5 inhibition induced death of prostate cancer cells and patient-derived prostate cancers surviving enzalutamide treatment and blocked enzalutamide-resistant tumor growth in mice. This work introduces a novel concept of a pivotal role of hyperactivated Jak2–Stat5 signaling in enzalutamide-resistant prostate cancer, which is readily targetable by Jak2 inhibitors in clinical development.
Data from WNK1 Enhances Migration and Invasion in Breast Cancer Models
Authors: Jaykumar Ankita B. , Jung Ji-Ung , Parida Pravat Kumar , Dang Tuyen T. , Wichaidit Chonlarat , Kannangara Ashari Rashmi , Earnest Svetlana , Goldsmith Elizabeth J. , Pearson Gray W. , Malladi Srinivas , Cobb Melanie H.
Published: April 2023

Abstract
Metastasis is the major cause of mortality in patients with breast cancer. Many signaling pathways have been linked to cancer invasiveness, but blockade of few protein components has succeeded in reducing metastasis. Thus, identification of proteins contributing to invasion that are manipulable by small molecules may be valuable in inhibiting spread of the disease. The protein kinase with no lysine (K) 1 (WNK1) has been suggested to induce migration of cells representing a range of cancer types. Analyses of mouse models and patient data have implicated WNK1 as one of a handful of genes uniquely linked to invasive breast cancer. Here, we present evidence that inhibition of WNK1 slows breast cancer metastasis. We show that depletion or inhibition of WNK1 reduces migration of several breast cancer cell lines in wound healing assays and decreases invasion in collagen matrices. Furthermore, WNK1 depletion suppresses expression of AXL, a tyrosine kinase implicated in metastasis. Finally, we demonstrate that WNK inhibition in mice attenuates tumor progression and metastatic burden. These data showing reduced migration, invasion, and metastasis upon WNK1 depletion in multiple breast cancer models suggest that WNK1 contributes to the metastatic phenotype, and that WNK1 inhibition may offer a therapeutic avenue for attenuating progression of invasive breast cancers.
Data from Identification of Genes Required for Enzalutamide Resistance in Castration-Resistant Prostate Cancer Cells In Vitro
Authors: Kohrt Sarah E. , Awadallah Wisam N. , Phillips Robert A. , Case Thomas C. , Jin Renjie , Nanda Jagpreet S. , Yu Xiuping , Clark Peter E. , Yi Yajun , Matusik Robert J. , Anderson Philip D. , Grabowska Magdalena M.
Published: April 2023

Abstract
Castration-resistant prostate cancer can be treated with the antiandrogen enzalutamide, but responses and duration of response are variable. To identify genes that support enzalutamide resistance, we performed a short hairpin RNA (shRNA) screen in the bone-homing, castration-resistant prostate cancer cell line, C4-2B. We identified 11 genes (TFAP2C, CAD, SPDEF, EIF6, GABRG2, CDC37, PSMD12, COL5A2, AR, MAP3K11, and ACAT1) whose loss resulted in decreased cell survival in response to enzalutamide. To validate our screen, we performed transient knockdowns in C4-2B and 22Rv1 cells and evaluated cell survival in response to enzalutamide. Through these studies, we validated three genes (ACAT1, MAP3K11, and PSMD12) as supporters of enzalutamide resistance in vitro. Although ACAT1 expression is lower in metastatic castration-resistant prostate cancer samples versus primary prostate cancer samples, knockdown of ACAT1 was sufficient to reduce cell survival in C4-2B and 22Rv1 cells. MAP3K11 expression increases with Gleason grade, and the highest expression is observed in metastatic castration-resistant disease. Knockdown of MAP3K11 reduced cell survival, and pharmacologic inhibition of MAP3K11 with CEP-1347 in combination with enzalutamide resulted in a dramatic increase in cell death. This was associated with decreased phosphorylation of AR-Serine650, which is required for maximal AR activation. Finally, although PSMD12 expression did not change during disease progression, knockdown of PSMD12 resulted in decreased AR and AR splice variant expression, likely contributing to the C4-2B and 22Rv1 decrease in cell survival. Our study has therefore identified at least three new supporters of enzalutamide resistance in castration-resistant prostate cancer cells in vitro.
Data from Targeting CDK4/6 Represents a Therapeutic Vulnerability in Acquired BRAF/MEK Inhibitor–Resistant Melanoma
Authors: Nassar Kelsey W. , Hintzsche Jennifer D. , Bagby Stacey M. , Espinoza Veronica , Langouët-Astrié Christophe , Amato Carol M. , Chimed Tugs-Saikhan , Fujita Mayumi , Robinson William , Tan Aik Choon , Schweppe Rebecca E.
Published: April 2023

Abstract
There is a clear need to identify targetable drivers of resistance and potential biomarkers for salvage therapy for patients with melanoma refractory to the combination of BRAF and MEK inhibition. In this study, we performed whole-exome sequencing on BRAF-V600E–mutant melanoma patient tumors refractory to the combination of BRAF/MEK inhibition and identified acquired oncogenic mutations in NRAS and loss of the tumor suppressor gene CDKN2A. We hypothesized the acquired resistance mechanisms to BRAF/MEK inhibition were reactivation of the MAPK pathway and activation of the cell-cycle pathway, which can both be targeted pharmacologically with the combination of a MEK inhibitor (trametinib) and a CDK4/6 inhibitor (palbociclib). In vivo, we found that combination of CDK4/6 and MEK inhibition significantly decreased tumor growth in two BRAF/MEK inhibitor–resistant patient-derived xenograft models. In vitro, we observed that the combination of CDK4/6 and MEK inhibition resulted in synergy and significantly reduced cellular growth, promoted cell-cycle arrest, and effectively inhibited downstream signaling of MAPK and cell-cycle pathways in BRAF inhibitor–resistant cell lines. Knockdown of CDKN2A in BRAF inhibitor–resistant cells increased sensitivity to CDK4/6 inhibition alone and in combination with MEK inhibition. A key implication of our study is that the combination of CDK4/6 and MEK inhibitors overcomes acquired resistance to BRAF/MEK inhibitors, and loss of CDKN2A may represent a biomarker of response to the combination. Inhibition of the cell-cycle and MAPK pathway represents a promising strategy for patients with metastatic melanoma who are refractory to BRAF/MEK inhibitor therapy.
Data from A Novel Peptidylarginine Deiminase 4 (PAD4) Inhibitor BMS-P5 Blocks Formation of Neutrophil Extracellular Traps and Delays Progression of Multiple Myeloma
Authors: Li Marina , Lin Cindy , Deng Hui , Strnad Joann , Bernabei Luca , Vogl Dan T. , Burke James J. , Nefedova Yulia
Published: April 2023

Abstract
Multiple myeloma is a plasma cell malignancy, which grows in the bone marrow (BM). The major population of cells in the BM is represented by neutrophils and they can form neutrophil extracellular traps (NET). Here, we investigated whether multiple myeloma cells induce NET formation and whether targeting this process would delay multiple myeloma progression. We demonstrated that murine and human multiple myeloma cells stimulate citrullination of histone H3 and NET formation by neutrophils and that this process is abrogated by pharmacological targeting of peptidylarginine deiminase 4 (PAD4) with a novel-specific small molecule inhibitor BMS-P5. Administration of BMS-P5 to multiple myeloma-bearing mice delays appearance of symptoms and disease progression. Taken together, our data demonstrate that targeting PAD4 may be beneficial for treatment of multiple myeloma.
Data from Individualized Prediction of Drug Response and Rational Combination Therapy in NSCLC Using Artificial Intelligence–Enabled Studies of Acute Phosphoproteomic Changes
Authors: Coker Elizabeth A. , Stewart Adam , Ozer Bugra , Minchom Anna , Pickard Lisa , Ruddle Ruth , Carreira Suzanne , Popat Sanjay , O'Brien Mary , Raynaud Florence , de Bono Johann , Al-Lazikani Bissan , Banerji Udai
Published: April 2023

Abstract
We hypothesize that the study of acute protein perturbation in signal transduction by targeted anticancer drugs can predict drug sensitivity of these agents used as single agents and rational combination therapy. We assayed dynamic changes in 52 phosphoproteins caused by an acute exposure (1 hour) to clinically relevant concentrations of seven targeted anticancer drugs in 35 non–small cell lung cancer (NSCLC) cell lines and 16 samples of NSCLC cells isolated from pleural effusions. We studied drug sensitivities across 35 cell lines and synergy of combinations of all drugs in six cell lines (252 combinations). We developed orthogonal machine-learning approaches to predict drug response and rational combination therapy. Our methods predicted the most and least sensitive quartiles of drug sensitivity with an AUC of 0.79 and 0.78, respectively, whereas predictions based on mutations in three genes commonly known to predict response to the drug studied, for example, EGFR, PIK3CA, and KRAS, did not predict sensitivity (AUC of 0.5 across all quartiles). The machine-learning predictions of combinations that were compared with experimentally generated data showed a bias to the highest quartile of Bliss synergy scores (P = 0.0243). We confirmed feasibility of running such assays on 16 patient samples of freshly isolated NSCLC cells from pleural effusions. We have provided proof of concept for novel methods of using acute ex vivo exposure of cancer cells to targeted anticancer drugs to predict response as single agents or combinations. These approaches could complement current approaches using gene mutations/amplifications/rearrangements as biomarkers and demonstrate the utility of proteomics data to inform treatment selection in the clinic.
Data from Datopotamab Deruxtecan, a Novel TROP2-directed Antibody–drug Conjugate, Demonstrates Potent Antitumor Activity by Efficient Drug Delivery to Tumor Cells
Authors: Okajima Daisuke , Yasuda Satoru , Maejima Takanori , Karibe Tsuyoshi , Sakurai Ken , Aida Tetsuo , Toki Tadashi , Yamaguchi Junko , Kitamura Michiko , Kamei Reiko , Fujitani Tomomichi , Honda Tomoyo , Shibutani Tomoko , Muramatsu Sumie , Nakada Takashi , Goto Riki , Takahashi Shu , Yamaguchi Miki , Hamada Hirofumi , Noguchi Yutaka , Murakami Masato , Abe Yuki , Agatsuma Toshinori
Published: April 2023

Abstract
Trophoblast cell surface antigen 2 (TROP2) is highly expressed on various epithelial tumors and correlates with poor prognosis. We developed the novel TROP2-directed antibody–drug conjugate (ADC), datopotamab deruxtecan (Dato-DXd, DS-1062a), with a potent DNA topoisomerase I inhibitor (DXd), and evaluated its antitumor activity and safety profiles in preclinical models.

The pharmacologic activity and mechanism of action of Dato-DXd were investigated in several human cancer cell lines and xenograft mouse models including patient-derived xenograft (PDX) models. Safety profiles were also assessed in rats and cynomolgus monkeys.

Dato-DXd bound specifically to TROP2 and was internalized into tumor cells followed by intracellular trafficking to lysosome and DXd release, which induced DNA damage and apoptosis in TROP2-expressing tumor cells in vitro. Dato-DXd exhibited in vivo antitumor activity with DNA damage induced by the accumulated DXd in TROP2-expressing xenograft tumors, but neither isotype control IgG-ADC nor anti-TROP2 antibody had this effect. Dato-DXd also showed potent antitumor activity with tumor regression in several TROP2-expressing xenograft tumors including NSCLC PDX models. Safety profiles of Dato-DXd in rats and cynomolgus monkeys were acceptable.

Dato-DXd demonstrated potent antitumor activity against TROP2-expressing tumors by efficient payload delivery into tumors and acceptable safety profiles in preclinical models. These results suggest Dato-DXd could be a valuable treatment option for patients with TROP2-expressing tumors in the clinical setting.
Data from Targeted Therapy to β3 Integrin Reduces Chemoresistance in Breast Cancer Bone Metastases
Authors: Fox Gregory C. , Su Xinming , Davis Jennifer L. , Xu Yalin , Kwakwa Kristin A. , Ross Michael H. , Fontana Francesca , Xiang Jingyu , Esser Alison K. , Cordell Elizabeth , Pagliai Kristen , Dang Ha X. , Sivapackiam Jothilingam , Stewart Sheila A. , Maher Christopher A. , Bakewell Suzanne J. , Fitzpatrick James A.J. , Sharma Vijay , Achilefu Samuel , Veis Deborah J. , Lanza Gregory M. , Weilbaecher Katherine N.
Published: April 2023

Abstract
Breast cancer bone metastases are common and incurable. Tumoral integrin β3 (β3) expression is induced through interaction with the bone microenvironment. Although β3 is known to promote bone colonization, its functional role during therapy of established bone metastases is not known. We found increased numbers of β3+ tumor cells in murine bone metastases after docetaxel chemotherapy. β3+ tumor cells were present in 97% of post-neoadjuvant chemotherapy triple-negative breast cancer patient samples (n = 38). High tumoral β3 expression was associated with worse outcomes in both pre- and postchemotherapy triple-negative breast cancer groups. Genetic deletion of tumoral β3 had minimal effect in vitro, but significantly enhanced in vivo docetaxel activity, particularly in the bone. Rescue experiments confirmed that this effect required intact β3 signaling. Ultrastructural, transcriptomic, and functional analyses revealed an alternative metabolic response to chemotherapy in β3-expressing cells characterized by enhanced oxygen consumption, reactive oxygen species generation, and protein production. We identified mTORC1 as a candidate for therapeutic targeting of this β3-mediated, chemotherapy-induced metabolic response. mTORC1 inhibition in combination with docetaxel synergistically attenuated murine bone metastases. Furthermore, micelle nanoparticle delivery of mTORC1 inhibitor to cells expressing activated αvβ3 integrins enhanced docetaxel efficacy in bone metastases. Taken together, we show that β3 integrin induction by the bone microenvironment promotes resistance to chemotherapy through an altered metabolic response that can be defused by combination with αvβ3-targeted mTORC1 inhibitor nanotherapy. Our work demonstrates the importance of the metastatic microenvironment when designing treatments and presents new, bone-specific strategies for enhancing chemotherapeutic efficacy.
Data from Tissue-Agnostic Activity of BRAF plus MEK Inhibitor in BRAF V600–Mutant Tumors
Authors: Adashek Jacob J. , Menta Arjun K. , Reddy Neha K. , Desai Aakash P. , Roszik Jason , Subbiah Vivek
Published: April 2023

Abstract
BRAF plus MEK inhibitor combinations are currently FDA-approved for melanoma, non–small cell lung cancer, and anaplastic thyroid cancer. The lack of clinical benefit with BRAF inhibition in BRAF V600–mutated colorectal cancer has prevented its tissue-agnostic drug development. We reviewed the AACR GENIE database for the prevalence of BRAF V600 mutations across tumor types. We reviewed the literature for case reports of clinical responses, outcomes in patients with BRAF V600 mutation—positive nonmelanoma malignancies who received BRAF inhibitor therapy, and data from published adult and pediatric trials. BRAF V600 mutations are prevalent across multiple nonmelanoma malignancies (>40 different tumor types), lead to oncogene addiction, and are clinically actionable in a broad range of adult and pediatric nonmelanoma rare malignancies. Continued tissue-agnostic drug development is warranted beyond the current BRAF plus MEK approved cancers.
Data from Cigarette Smoke Containing Acrolein Contributes to Cisplatin Resistance in Human Bladder Cancers through the Regulation of HER2 Pathway or FGFR3 Pathway
Authors: Hong Jian-Hua , Tong Zhen-Jie , Wei Tung-En , Lu Yu-Chuan , Huang Cheng-Yu , Huang Chao-Yuan , Chiang Chih-Hung , Jaw Fu-Shan , Cheng Hsiao-Wei , Wang Hsiang-Tsui
Published: April 2023

Abstract
Cisplatin-based chemotherapy is the first-line therapy for bladder cancer. However, cisplatin resistance has been associated with the recurrence of bladder cancer. Previous studies have shown that activation of FGFR and HER2 signaling are involved in bladder cancer cell proliferation and drug resistance. Smoking is the most common etiologic risk factor for bladder cancer, and there is emerging evidence that smoking is associated with cisplatin resistance. However, the underlying mechanism remains elusive. Acrolein, a highly reactive aldehyde, is abundant in tobacco smoke, cooking fumes, and automobile exhaust fumes. Our previous studies have shown that acrolein contributes to bladder carcinogenesis through the induction of DNA damage and inhibition of DNA repair. In this study, we found that acrolein induced cisplatin resistance and tumor progression in both non–muscle invasive bladder cancer (NMIBC) and muscle invasive bladder cancer (MIBC) cell lines RT4 and T24, respectively. Activation of HER2 and FGFR3 signaling contributes to acrolein-induced cisplatin resistance in RT4 and T24 cells, respectively. Furthermore, trastuzumab, an anti-HER2 antibody, and PD173074, an FGFR inhibitor, reversed cisplatin resistance in RT4 and T24 cells, respectively. Using a xenograft mouse model with acrolein-induced cisplatin-resistant T24 clones, we found that cisplatin combined with PD173074 significantly reduced tumor size compared with cisplatin alone. These results indicate that differential molecular alterations behind cisplatin resistance in NMIBC and MIBC significantly alter the effectiveness of targeted therapy combined with chemotherapy. This study provides valuable insights into therapeutic strategies for cisplatin-resistant bladder cancer.
Data from SLFN11 is Widely Expressed in Pediatric Sarcoma and Induces Variable Sensitization to Replicative Stress Caused By DNA-Damaging Agents
Authors: Gartrell Jessica , Mellado-Largarde Marcia , Clay Michael R. , Bahrami Armita , Sahr Natasha A. , Sykes April , Blankenship Kaley , Hoffmann Lauren , Xie Jia , Cho Hyekyung P. , Twarog Nathaniel , Connelly Michele , Yan Koon-Kiu , Yu Jiyang , Porter Shaina N. , Pruett-Miller Shondra M. , Neale Geoffrey , Tinkle Christopher L. , Federico Sara M. , Stewart Elizabeth A. , Shelat Anang A.
Published: April 2023

Abstract
Pediatric sarcomas represent a heterogeneous group of malignancies that exhibit variable response to DNA-damaging chemotherapy. Schlafen family member 11 protein (SLFN11) increases sensitivity to replicative stress and has been implicated as a potential biomarker to predict sensitivity to DNA-damaging agents (DDA). SLFN11 expression was quantified in 220 children with solid tumors using IHC. Sensitivity to the PARP inhibitor talazoparib (TAL) and the topoisomerase I inhibitor irinotecan (IRN) was assessed in sarcoma cell lines, including SLFN11 knock-out (KO) and overexpression models, and a patient-derived orthotopic xenograft model (PDOX). SLFN11 was expressed in 69% of pediatric sarcoma sampled, including 90% and 100% of Ewing sarcoma and desmoplastic small round-cell tumors, respectively, although the magnitude of expression varied widely. In sarcoma cell lines, protein expression strongly correlated with response to TAL and IRN, with SLFN11 KO resulting in significant loss of sensitivity in vitro and in vivo. Surprisingly, retrospective analysis of children with sarcoma found no association between SLFN11 levels and favorable outcome. Subsequently, high SLFN11 expression was confirmed in a PDOX model derived from a patient with recurrent Ewing sarcoma who failed to respond to treatment with TAL + IRN. Selective inhibition of BCL-xL increased sensitivity to TAL + IRN in SLFN11-positive resistant tumor cells. Although SLFN11 appears to drive sensitivity to replicative stress in pediatric sarcomas, its potential to act as a biomarker may be limited to certain tumor backgrounds or contexts. Impaired apoptotic response may be one mechanism of resistance to DDA-induced replicative stress.
Data from Inhibition of Oxidized Nucleotide Sanitation By TH1579 and Conventional Chemotherapy Cooperatively Enhance Oxidative DNA Damage and Survival in AML
Authors: Centio Anders , Estruch Montserrat , Reckzeh Kristian , Sanjiv Kumar , Vittori Camilla , Engelhard Sophia , Warpman Berglund Ulrika , Helleday Thomas , Theilgaard-Mönch Kim
Published: April 2023

Abstract
Currently, the majority of patients with acute myeloid leukemia (AML) still die of their disease due to primary resistance or relapse toward conventional reactive oxygen species (ROS)- and DNA damage–inducing chemotherapy regimens. Herein, we explored the therapeutic potential to enhance chemotherapy response in AML, by targeting the ROS scavenger enzyme MutT homolog 1 (MTH1, NUDT1), which protects cellular integrity through prevention of fatal chemotherapy-induced oxidative DNA damage. We demonstrate that MTH1 is a potential druggable target expressed by the majority of patients with AML and the inv(16)/KITD816Y AML mouse model mimicking the genetics of patients with AML exhibiting poor response to standard chemotherapy (i.e., anthracycline & cytarabine). Strikingly, combinatorial treatment of inv(16)/KITD816Y AML cells with the MTH1 inhibitor TH1579 and ROS- and DNA damage-inducing standard chemotherapy induced growth arrest and incorporated oxidized nucleotides into DNA leading to significantly increased DNA damage. Consistently, TH1579 and chemotherapy synergistically inhibited growth of clonogenic inv(16)/KITD816Y AML cells without substantially inhibiting normal clonogenic bone marrow cells. In addition, combinatorial treatment of inv(16)/KITD816Y AML mice with TH1579 and chemotherapy significantly reduced AML burden and prolonged survival compared with untreated or single treated mice. In conclusion, our study provides a rationale for future clinical studies combining standard AML chemotherapy with TH1579 to boost standard chemotherapy response in patients with AML. Moreover, other cancer entities treated with ROS- and DNA damage–inducing chemo- or radiotherapies might benefit therapeutically from complementary treatment with TH1579.
Data from Loss of Claudin-4 Reduces DNA Damage Repair and Increases Sensitivity to PARP Inhibitors
Authors: Yamamoto Tomomi M. , Webb Patricia G. , Davis Dana M. , Baumgartner Heidi K. , Woodruff Elizabeth R. , Guntupalli Saketh R. , Neville Margaret , Behbakht Kian , Bitler Benjamin G.
Published: April 2023

Abstract
High-grade serous ovarian cancer is the deadliest gynecologic malignancy due to progression to resistant disease. Claudin-4 is classically defined as a tight junction protein and is often associated with epithelial cancers. Claudin-4 is aberrantly expressed in nearly 70% of all ovarian cancer tumors and conveys a worse overall prognosis. Elevated claudin-4 expression correlates to increased DNA repair activity and resistance to DNA damaging agents. PARP inhibitors are emerging as an effective therapeutic option for patients with ovarian cancer and function by promoting DNA damage. The study examines the relationship between claudin-4 expression and the response to PARP inhibitors using both genetic and pharmacologic inhibition of claudin-4 in in vitro and ex vivo models of ovarian cancer to examine DNA repair markers and functional activity. Genetic inhibition of claudin-4 results in the downregulation of several DNA damage repair effectors, including 53BP1 and XRCC1. Claudin-4 knockdown did not change homology-directed repair but inhibited nonhomologous end-joining and reduced 53BP1 foci formation. In 15 primary ovarian cancer tumors, higher claudin-4 expression significantly correlated to a dampened PARP inhibitor-mediated antiproliferation response. Further, claudin-4 inhibition in high claudin-4 tumors sensitized tumor sections to PARP inhibition. These data highlight that claudin-4 expression in ovarian cancer tumors could serve as both a marker of PARP inhibitor response and a therapeutic target to improve PARP inhibitor response.
Data from Calicheamicin Antibody–Drug Conjugates with Improved Properties
Authors: Vollmar Breanna S. , Frantz Chris , Schutten Melissa M. , Zhong Fiona , del Rosario Geoffrey , Go Mary Ann T. , Yu Shang-Fan , Leipold Douglas D. , Kamath Amrita V. , Ng Carl , Xu Keyang , dela Cruz-Chuh Josefa , Kozak Katherine R. , Chen Jinhua , Xu Zijin , Wai John , Adhikari Pragya , Erickson Hans K. , Dragovich Peter S. , Polson Andrew G. , Pillow Thomas H.
Published: April 2023

Abstract
Calicheamicin antibody–drug conjugates (ADCs) are effective therapeutics for leukemias with two recently approved in the United States: Mylotarg (gemtuzumab ozogamicin) targeting CD33 for acute myeloid leukemia and Besponsa (inotuzumab ozogamicin) targeting CD22 for acute lymphocytic leukemia. Both of these calicheamicin ADCs are heterogeneous, aggregation-prone, and have a shortened half-life due to the instability of the acid-sensitive hydrazone linker in circulation. We hypothesized that we could improve upon the heterogeneity, aggregation, and circulation stability of calicheamicin ADCs by directly attaching the thiol of a reduced calicheamicin to an engineered cysteine on the antibody via a disulfide bond to generate a linkerless and traceless conjugate. We report herein that the resulting homogeneous conjugates possess minimal aggregation and display high in vivo stability with 50% of the drug remaining conjugated to the antibody after 21 days. Furthermore, these calicheamicin ADCs are highly efficacious in mouse models of both solid tumor (HER2+ breast cancer) and hematologic malignancies (CD22+ non-Hodgkin lymphoma). Safety studies in rats with this novel calicheamicin ADC revealed an increased tolerability compared with that reported for Mylotarg. Overall, we demonstrate that applying novel linker chemistry with site-specific conjugation affords an improved, next-generation calicheamicin ADC.
Data from Activity of Combined Androgen Receptor Antagonism and Cell Cycle Inhibition in Androgen Receptor Positive Triple Negative Breast Cancer
Authors: Christenson Jessica L. , O'Neill Kathleen I. , Williams Michelle M. , Spoelstra Nicole S. , Jones Kenneth L. , Trahan G. Devon , Reese Jordan , Van Patten Elaina T. , Elias Anthony , Eisner Joel R. , Richer Jennifer K.
Published: April 2023

Abstract
Triple-negative breast cancer (TNBC) is an aggressive subtype, with a peak recurrence rate within the first few years after diagnosis. Few targeted therapies are available to treat this breast cancer subtype, defined by the lack of estrogen receptor (ER) and progesterone receptor and without amplification of human epidermal growth factor receptor 2 (HER2). Although cell cycle cyclin-dependent kinase (CDK) 4/6 inhibitors are approved for treatment of ER-positive (ER+) breast cancer, they have not proven effective as monotherapy in patients with TNBC. The androgen receptor (AR) has emerged as a therapeutic target in a subset of TNBCs and with significant clinical benefit observed in multiple trials. The purpose of this study was to investigate the preclinical activity of the CDK4/6 inhibitor, abemaciclib, in combination with an agent that targets both androgen biosynthesis and AR activity, seviteronel, using TNBC cell lines expressing high AR, cell line xenografts, and an AR-positive (AR+), androgen-responsive TNBC patient-derived xenograft (PDX). Single-cell RNA sequencing demonstrated heterogeneity in AR levels, even in a highly AR+ cell line, and identified cell cycle pathway activation in ARHigh- versus ARLow-expressing cells. Combination treatment with the cell cycle CDK4/6 inhibitor, abemaciclib, and seviteronel showed synergy in an AR+ TNBC model compared with each drug alone. Although cell cycle inhibitors are FDA approved for use in ER+ breast cancer, our studies suggest that they may also be effective in AR+ TNBC, perhaps combined with AR-targeted agents.
Data from Concurrent Inhibition of ERK and Farnesyltransferase Suppresses the Growth of HRAS Mutant Head and Neck Squamous Cell Carcinoma
Authors: Javaid Sehrish , Schaefer Antje , Goodwin Craig M. , Nguyen Victoria V. , Massey Frances L. , Pierobon Mariaelena , Gambrell-Sanders Da'Jhnae , Waters Andrew M. , Lambert Kathryn N. , Diehl J. Nathaniel , Hobbs G. Aaron , Wood Kris C. , Petricoin Emanuel F. , Der Channing J. , Cox Adrienne D.
Published: April 2023

Abstract
Human papilloma virus (HPV)-negative head and neck squamous cell carcinoma (HNSCC) is a common cancer worldwide with an unmet need for more effective, less toxic treatments. Currently, both the disease and the treatment of HNSCC cause significant mortality and morbidity. Targeted therapies hold new promise for patients with HPV-negative status whose tumors harbor oncogenic HRAS mutations. Recent promising clinical results have renewed interest in the development of farnesyltransferase inhibitors (FTIs) as a therapeutic strategy for HRAS-mutant cancers. With the advent of clinical evaluation of the FTI tipifarnib for the treatment of HRAS-mutant HNSCC, we investigated the activity of tipifarnib and inhibitors of HRAS effector signaling in HRAS-mutant HNSCC cell lines. First, we validated that HRAS is a cancer driver in HRAS-mutant HNSCC lines. Second, we showed that treatment with the FTI tipifarnib largely phenocopied HRAS silencing, supporting HRAS as a key target of FTI antitumor activity. Third, we performed reverse-phase protein array analyses to profile FTI treatment-induced changes in global signaling, and conducted CRISPR/Cas9 genetic loss-of-function screens to identify previously unreported genes and pathways that modulate sensitivity to tipifarnib. Fourth, we determined that concurrent inhibition of HRAS effector signaling (ERK, PI3K, mTORC1) increased sensitivity to tipifarnib treatment, in part by overcoming tipifarnib-induced compensatory signaling. We also determined that ERK inhibition could block tipifarnib-induced epithelial-to-mesenchymal transition, providing a potential basis for the effectiveness of this combination. Our results support future investigations of these and other combination treatments for HRAS mutant HNSCC.
Data from Targeting CD73 with AB680 (Quemliclustat), a Novel and Potent Small-Molecule CD73 Inhibitor, Restores Immune Functionality and Facilitates Antitumor Immunity
Authors: Piovesan Dana , Tan Joanne B.L. , Becker Annette , Banuelos Jesus , Narasappa Nell , DiRenzo Daniel , Zhang Kristen , Chen Ada , Ginn Elaine , Udyavar Akshata R. , Yin Fangfang , Paprcka Susan L. , Purandare Bhamini , Park Timothy W. , Kimura Nikki , Kalisiak Jaroslaw , Young Stephen W. , Powers Jay P. , Schindler Uli , Sivick Kelsey E. , Walters Matthew J.
Published: April 2023

Abstract
T cells play a critical role in the control of cancer. The development of immune checkpoint blockers (ICB) aimed at enhancing antitumor T-cell responses has revolutionized cancer treatment. However, durable clinical benefit is observed in only a subset of patients, prompting research efforts to focus on strategies that target multiple inhibitory signals within the tumor microenvironment (TME) to limit tumor evasion and improve patient outcomes. Adenosine has emerged as a potent immune suppressant within the TME, and CD73 is the major enzyme responsible for its extracellular production. CD73 can be co-opted within the TME to impair T-cell–mediated antitumor immunity and promote tumor growth. To target this pathway and block the formation of adenosine, we designed a novel, selective, and potent class of small-molecule inhibitors of CD73, including AB680 (quemliclustat), which is currently being tested in patients with cancer. AB680 effectively restored T-cell proliferation, cytokine secretion, and cytotoxicity that were dampened by the formation of immunosuppressive adenosine by CD73. Furthermore, in an allogeneic mixed lymphocyte reaction where CD73-derived adenosine had a dominant suppressive effect in the presence of PD-1 blockade, AB680 restored T-cell activation and function. Finally, in a preclinical mouse model of melanoma, AB680 inhibited CD73 in the TME and increased the antitumor activity of PD-1 blockade. Collectively, these data provide a rationale for the inhibition of CD73 with AB680 in combination with ICB, such as anti–PD-1, to improve cancer patient outcomes.
Data from The Mitochondrial Disruptor Devimistat (CPI-613) Synergizes with Genotoxic Anticancer Drugs in Colorectal Cancer Therapy in a Bim-Dependent Manner
Authors: Arnold Carina , Demuth Philipp , Seiwert Nina , Wittmann Simon , Boengler Kerstin , Rasenberger Birgit , Christmann Markus , Huber Magdalena , Brunner Thomas , Linnebacher Michael , Fahrer Jörg
Published: April 2023

Abstract
Colorectal cancer is one of the most frequent tumor entities, with an increasing incidence and mortality in younger adults in Europe and the United States. Five-year survival rates for advanced colorectal cancer are still low, highlighting the need for novel targets in colorectal cancer therapy. Here, we investigated the therapeutic potential of the compound devimistat (CPI-613) that targets altered mitochondrial cancer cell metabolism and its synergism with the antineoplastic drugs 5-fluorouracil (5-FU) and irinotecan (IT) in colorectal cancer. Devimistat exerted a comparable cytotoxicity in a panel of established colorectal cancer cell lines and patient-derived short-term cultures independent of their genetic and epigenetic status, whereas human colonic epithelial cells were more resistant, indicating tumor selectivity. These findings were corroborated in intestinal organoid and tumoroid models. Mechanistically, devimistat disrupted mitochondrial membrane potential and severely impaired mitochondrial respiration, resulting in colorectal cancer cell death induction independent of p53. Combination treatment of devimistat with 5-FU or IT demonstrated synergistic cell killing in colorectal cancer cells as shown by Combenefit modeling and Chou–Talalay analysis. Increased cell death induction was revealed as a major mechanism involving downregulation of antiapoptotic genes and accumulation of proapoptotic Bim, which was confirmed by its genetic knockdown. In human colorectal cancer xenograft mouse models, devimistat showed antitumor activity and synergized with IT, resulting in prolonged survival and enhanced therapeutic efficacy. In human tumor xenografts, devimistat prevented IT-triggered p53 stabilization and caused synergistic Bim induction. Taken together, our study revealed devimistat as a promising candidate in colorectal cancer therapy by synergizing with established antineoplastic drugs in vitro and in vivo.
Data from In Vivo Efficacy of Tesevatinib in EGFR-Amplified Patient-Derived Xenograft Glioblastoma Models May Be Limited by Tissue Binding and Compensatory Signaling
Authors: Kizilbash Sani H. , Gupta Shiv K. , Parrish Karen E. , Laramy Janice K. , Kim Minjee , Gampa Gautham , Carlson Brett L. , Bakken Katrina K. , Mladek Ann C. , Schroeder Mark A. , Decker Paul A. , Elmquist William F. , Sarkaria Jann N.
Published: April 2023

Abstract
Tesevatinib is a potent oral brain penetrant EGFR inhibitor currently being evaluated for glioblastoma therapy. Tesevatinib distribution was assessed in wild-type (WT) and Mdr1a/b(-/-)Bcrp(-/-) triple knockout (TKO) FVB mice after dosing orally or via osmotic minipump; drug–tissue binding was assessed by rapid equilibrium dialysis. Two hours after tesevatinib dosing, brain concentrations in WT and TKO mice were 0.72 and 10.03 μg/g, respectively. Brain-to-plasma ratios (Kp) were 0.53 and 5.73, respectively. With intraperitoneal infusion, brain concentrations were 1.46 and 30.6 μg/g (Kp 1.16 and 25.10), respectively. The brain-to-plasma unbound drug concentration ratios were substantially lower (WT mice, 0.03–0.08; TKO mice, 0.40–1.75). Unbound drug concentrations in brains of WT mice were 0.78 to 1.59 ng/g. In vitro cytotoxicity and EGFR pathway signaling were evaluated using EGFR-amplified patient-derived glioblastoma xenograft models (GBM12, GBM6). In vivo pharmacodynamics and efficacy were assessed using athymic nude mice bearing either intracranial or flank tumors treated by oral gavage. Tesevatinib potently reduced cell viability [IC50 GBM12 = 11 nmol/L (5.5 ng/mL), GBM6 = 102 nmol/L] and suppressed EGFR signaling in vitro. However, tesevatinib efficacy compared with vehicle in intracranial (GBM12, median survival: 23 vs. 18 days, P = 0.003) and flank models (GBM12, median time to outcome: 41 vs. 33 days, P = 0.007; GBM6, 44 vs. 33 days, P = 0.007) was modest and associated with partial inhibition of EGFR signaling. Overall, tesevatinib efficacy in EGFR-amplified PDX GBM models is robust in vitro but relatively modest in vivo, despite a high brain-to-plasma ratio. This discrepancy may be explained by drug-tissue binding and compensatory signaling.
Data from Enhancing Proteotoxic Stress in Leiomyosarcoma Cells Triggers Mitochondrial Dysfunctions, Cell Death, and Antitumor Activity in vivo
Authors: Iuliano Luca , Drioli Sara , Pignochino Ymera , Cafiero Claudia Maria , Minisini Martina , D'Este Francesca , Picco Raffaella , Dalla Emiliano , Giordano Giorgia , Grignani Giovanni , Di Giorgio Eros , Benedetti Fabio , Felluga Fulvia , Brancolini Claudio
Published: April 2023

Abstract
Leiomyosarcomas are rare and aggressive tumors characterized by a complex karyotype. Surgical resection with or without radiotherapy and chemotherapy is the standard curative treatment. Unfortunately, a high percentage of leiomyosarcomas recurs and metastasizes. In these cases, doxorubicin and ifosfamide represent the standard treatment but with low response rates. Here, we evaluated the induction of proteotoxic stress as a possible strategy to kill leiomyosarcoma cells in a therapeutic perspective. We show that aggressive leiomyosarcomas coexist with high levels of proteotoxic stress. As a consequence, we hypothesized that leiomyosarcoma cells are vulnerable to further increases of proteotoxic stress. The small compound 2c is a strong inducer of proteotoxic stress. In leiomyosarcoma cells, it triggers cell death coupled to a profound reorganization of the mitochondrial network. By using stimulated emission depletion microscopy, we have unveiled the existence of DIABLO/SMAC clusters that are modulated by 2c. Finally, we have engineered a new version of 2c linked to polyethylene glycol though a short peptide, named 2cPP. This new prodrug is specifically activated by proteases present in the tumor microenvironment. 2cPP shows a strong antitumor activity in vivo against leiomyosarcomas and no toxicity against normal cells.
Data from Dual G9A/EZH2 Inhibition Stimulates Antitumor Immune Response in Ovarian High-Grade Serous Carcinoma
Authors: Spiliopoulou Pavlina , Spear Sarah , Mirza Hasan , Garner Ian , McGarry Lynn , Grundland-Freile Fabio , Cheng Zhao , Ennis Darren P. , Iyer Nayana , McNamara Sophie , Natoli Marina , Mason Susan , Blyth Karen , Adams Peter D. , Roxburgh Patricia , Fuchter Matthew J. , Brown Bob , McNeish Iain A.
Published: April 2023

Abstract
Ovarian high-grade serous carcinoma (HGSC) prognosis correlates directly with presence of intratumoral lymphocytes. However, cancer immunotherapy has yet to achieve meaningful survival benefit in patients with HGSC. Epigenetic silencing of immunostimulatory genes is implicated in immune evasion in HGSC and re-expression of these genes could promote tumor immune clearance. We discovered that simultaneous inhibition of the histone methyltransferases G9A and EZH2 activates the CXCL10–CXCR3 axis and increases homing of intratumoral effector lymphocytes and natural killer cells while suppressing tumor-promoting FoxP3+ CD4 T cells. The dual G9A/EZH2 inhibitor HKMTI-1–005 induced chromatin changes that resulted in the transcriptional activation of immunostimulatory gene networks, including the re-expression of elements of the ERV-K endogenous retroviral family. Importantly, treatment with HKMTI-1–005 improved the survival of mice bearing Trp53−/− null ID8 ovarian tumors and resulted in tumor burden reduction. These results indicate that inhibiting G9A and EZH2 in ovarian cancer alters the immune microenvironment and reduces tumor growth and therefore positions dual inhibition of G9A/EZH2 as a strategy for clinical development.
Data from Durvalumab Combined with Immunomodulatory Drugs (IMiD) Overcomes Suppression of Antitumor Responses due to IMiD-induced PD-L1 Upregulation on Myeloma Cells
Authors: Ishibashi Mariko , Yamamoto Junichi , Ito Takumi , Handa Hiroshi , Sunakawa-Kii Mika , Inokuchi Koiti , Morita Rimpei , Tamura Hideto
Published: April 2023

Abstract
We previously showed that the interaction of programmed death-ligand 1 (PD-L1) on multiple myeloma (MM) cells with PD-1 not only inhibits tumor-specific cytotoxic T-lymphocyte activity via the PD-1 signaling pathway but also induces drug resistance via PD-L1–mediated reverse signals. We here examined the regulation of PD-L1 expression by immunomodulatory drugs (IMiDs) and antimyeloma effects of the anti–PD-L1 antibody durvalumab in combination with IMiDs. IMiDs induced PD-L1 expression on IMiD-insensitive MM cells and plasma cells from patients newly diagnosed with MM. Gene-expression profiling analysis demonstrated that not only PD-L1, but also a proliferation-inducing ligand (APRIL), was enhanced by IMiDs. PD-L1 induction by IMiDs was suppressed by using the APRIL inhibitor recombinant B-cell maturation antigen (BCMA)-Ig, the antibody against BCMA, or an MEK/ERK inhibitor in in vitro and in vivo assays. In addition, its induction was abrogated in cereblon (CRBN)-knockdown MM cells, whereas PD-L1 expression was increased and strongly induced by IMiDs in Ikaros-knockdown cells. These results demonstrated that PD-L1 upregulation by IMiDs on IMiD-insensitive MM cells was induced by (i) the BCMA–APRIL pathway via IMiD-mediated induction of APRIL and (ii) Ikaros degradation mediated by CRBN, which plays a role in inhibiting PD-L1 expression. Furthermore, T-cell inhibition induced by PD-L1–upregulated cells was effectively recovered after combination treatment with durvalumab and IMiDs. PD-L1 upregulation by IMiDs on MM cells might promote aggressive myeloma behaviors and immune escape in the bone marrow microenvironment.
Data from Novel TGFβ Inhibitors Ameliorate Oral Squamous Cell Carcinoma Progression and Improve the Antitumor Immune Response of Anti–PD-L1 Immunotherapy
Authors: Ludwig Nils , Wieteska Łukasz , Hinck Cynthia S. , Yerneni Saigopalakrishna S. , Azambuja Juliana H. , Bauer Richard J. , Reichert Torsten E. , Hinck Andrew P. , Whiteside Theresa L.
Published: April 2023

Abstract
TGFβ is a key regulator of oral squamous cell carcinoma (OSCC) progression, and its potential role as a therapeutic target has been investigated with a limited success. This study evaluates two novel TGFβ inhibitors as mono or combinatorial therapy with anti–PD-L1 antibodies (α-PD-L1 Ab) in a murine OSCC model. Immunocompetent C57BL/6 mice bearing malignant oral lesions induced by 4-nitroquinoline 1-oxide (4-NQO) were treated for 4 weeks with TGFβ inhibitors mRER (i.p., 50 μg/d) or mmTGFβ2-7m (10 μg/d delivered by osmotic pumps) alone or in combination with α-PD-L1 Abs (7× i.p. of 100 μg/72 h). Tumor progression and body weight were monitored. Levels of bioactive TGFβ in serum were quantified using a TGFβ bioassay. Tissues were analyzed by immunohistology and flow cytometry. Therapy with mRER or mmTGFβ2-7m reduced tumor burden (P < 0.05) and decreased body weight loss compared with controls. In inhibitor-treated mice, levels of TGFβ in tumor tissue and serum were reduced (P < 0.05), whereas they increased with tumor progression in controls. Both inhibitors enhanced CD8+ T-cell infiltration into tumors and mRER reduced levels of myeloid-derived suppressor cells (P < 0.001). In combination with α-PD-L1 Abs, tumor burden was not further reduced; however, mmTGFβ2-7m further reduced weight loss (P < 0.05). The collagen-rich stroma was reduced by using combinatorial TGFβ/PD-L1 therapies (P < 0.05), enabling an accelerated lymphocyte infiltration into tumor tissues. The blockade of TGFβ signaling by mRER or mmTGFβ2-7m ameliorated in vivo progression of established murine OSCC. The inhibitors promoted antitumor immune responses, alone and in combination with α-PD-L1 Abs.
Data from Phase I/II Trial of Vemurafenib in Dogs with Naturally Occurring, BRAF-mutated Urothelial Carcinoma
Authors: Rossman Paul , Zabka Tanja S. , Ruple Audrey , Tuerck Dietrich , Ramos-Vara José A. , Liu Liling , Mohallem Rodrigo , Merchant Mark , Franco Jackeline , Fulkerson Christopher M. , Bhide Ketaki P. , Breen Matthew , Aryal Uma K. , Murray Elaine , Dybdal Noel , Utturkar Sagar M. , Fourez Lindsey M. , Enstrom Alexander W. , Dhawan Deepika , Knapp Deborah W.
Published: April 2023

Abstract
BRAF-targeted therapies including vemurafenib (Zelboraf) induce dramatic cancer remission; however, drug resistance commonly emerges. The purpose was to characterize a naturally occurring canine cancer model harboring complex features of human cancer, to complement experimental models to improve BRAF-targeted therapy. A phase I/II clinical trial of vemurafenib was performed in pet dogs with naturally occurring invasive urothelial carcinoma (InvUC) harboring the canine homologue of human BRAFV600E. The safety, MTD, pharmacokinetics, and antitumor activity were determined. Changes in signaling and immune gene expression were assessed by RNA sequencing and phosphoproteomic analyses of cystoscopic biopsies obtained before and during treatment, and at progression. The vemurafenib MTD was 37.5 mg/kg twice daily. Anorexia was the most common adverse event. At the MTD, partial remission occurred in 9 of 24 dogs (38%), with a median progression-free interval of 181 days (range, 53–608 days). In 18% of the dogs, new cutaneous squamous cell carcinoma and papillomas occurred, a known pharmacodynamic effect of vemurafenib in humans. Upregulation of genes in the classical and alternative MAPK-related pathways occurred in subsets of dogs at cancer progression. The most consistent transcriptomic changes were the increase in patterns of T lymphocyte infiltration during the first month of vemurafenib, and of immune failure accompanying cancer progression. In conclusion, the safety, antitumor activity, and cutaneous pharmacodynamic effects of vemurafenib, and the development of drug resistance in dogs closely mimic those reported in humans. This suggests BRAF-mutated canine InvUC offers an important complementary animal model to improve BRAF-targeted therapies in humans.