bluuebunny/crossref_metadata_2025 huggingface dataset:
Supplementary Figure S1 from Nonclinical Development of Next-generation Site-specific HER2-targeting Antibody–drug Conjugate (ARX788) for Breast Cancer Treatment
Authors: Nagaraja Shastri Prathap , Zhu Jingjing , Skidmore Lillian , Liang Xuejun , Ji Yanping , Gu Yi , Tian Feng , Yao Sulan , Xia Gang
Published: April 2023

Supplementary Figure S1 illustrates ARX788 metabolism study in HCC1954 (HER2 expressing) cells. The metabolite pAF-AS269 was more abundantly detected in the cell sample than in the culture medium (A). Briefly, after ARX788 incubation for various time points, supernatant was collected from the wells and centrifuged to pellet floating cells. The resulting supernatant was processed as the medium sample, while the cell pellet was combined with the remaining adherent cells and processed as the cell sample, for LC-MS analysis. Additionally, the cell sample was counted in a hemacytometer to determine cell number and protein content was measured by BCA analysis. When compared to untreated cells, ARX788 inhibited HCC1954 cellular proliferation with fewer cells (B) and lower protein content (C) observed after 24-hours. Cell number (D) and protein content (E) remained unchanged in ARX788-treated and untreated MDA-MB-468 cells, a non-HER2 expressing cell line.
Suppl. Table S5 from A Cell-Based MAPK Reporter Assay Reveals Synergistic MAPK Pathway Activity Suppression by MAPK Inhibitor Combination in BRAF-Driven Pediatric Low-Grade Glioma Cells
Authors: Usta Diren , Sigaud Romain , Buhl Juliane L. , Selt Florian , Marquardt Viktoria , Pauck David , Jansen Jennifer , Pusch Stefan , Ecker Jonas , Hielscher Thomas , Vollmer Johanna , Sommerkamp Alexander C. , Rubner Tobias , Hargrave Darren , van Tilburg Cornelis M. , Pfister Stefan M. , Jones David T.W. , Remke Marc , Brummer Tilman , Witt Olaf , Milde Till
Published: April 2023

Suppl. Table S5 shows "Validated IC50 values and cmax and ctrough levels of chosen MAPKi".
Suppl. Table S4 from A Cell-Based MAPK Reporter Assay Reveals Synergistic MAPK Pathway Activity Suppression by MAPK Inhibitor Combination in BRAF-Driven Pediatric Low-Grade Glioma Cells
Authors: Usta Diren , Sigaud Romain , Buhl Juliane L. , Selt Florian , Marquardt Viktoria , Pauck David , Jansen Jennifer , Pusch Stefan , Ecker Jonas , Hielscher Thomas , Vollmer Johanna , Sommerkamp Alexander C. , Rubner Tobias , Hargrave Darren , van Tilburg Cornelis M. , Pfister Stefan M. , Jones David T.W. , Remke Marc , Brummer Tilman , Witt Olaf , Milde Till
Published: April 2023

Suppl. Table S4 shows "MAPKi combinations chosen based on low estimated IC50s and pharmaceutical company in consideration of future clinical studies".
Supplemental File 1 from The Landscape of Glycogen Synthase Kinase-3 Beta Genomic Alterations in Cancer
Authors: Borden Brittany A. , Baca Yasmine , Xiu Joanne , Tavora Fabio , Winer Ira , Weinberg Benjamin A. , Vanderwalde Ari M. , Darabi Sourat , Korn W. Michael , Mazar Andrew P. , Giles Francis J. , Crawford Lorin , Safran Howard , El-Deiry Wafik S. , Carneiro Benedito A.
Published: April 2023

References analyzed as part of cBioPortal cohort
Supplementary Table 1 from Targeting Casein Kinase 1 Delta Sensitizes Pancreatic and Bladder Cancer Cells to Gemcitabine Treatment by Upregulating Deoxycytidine Kinase
Authors: Vena Francesca , Bayle Simon , Nieto Ainhoa , Quereda Victor , Aceti Massimiliano , Frydman Sylvia M. , Sansil Samer S. , Grant Wayne , Monastyrskyi Andrii , McDonald Patricia , Roush William R. , Teng Mingxiang , Duckett Derek
Published: April 2023

Supplementary Table 1
Figure S7 from Blockade of Cell Volume Regulatory Protein NKCC1 Increases TMZ-Induced Glioma Apoptosis and Reduces Astrogliosis
Authors: Luo Lanxin , Guan Xiudong , Begum Gulnaz , Ding Dawei , Gayden Jenesis , Hasan Md Nabiul , Fiesler Victoria M. , Dodelson Jacob , Kohanbash Gary , Hu Baoli , Amankulor Nduka M. , Jia Wang , Castro Maria G. , Sun Baoshan , Sun Dandan
Published: April 2023

Fig. S7. Schematic illustration of NKCC1 activation in glioma and glioma-associated reactive astrocytes.
Supplementary Data from NRG1/ERBB3 Pathway Activation Induces Acquired Resistance to XPO1 Inhibitors
Authors: Miyake Takahito M. , Pradeep Sunila , Bayraktar Emine , Stur Elaine , Handley Katelyn F. , Wu Sherry Y. , Rodriguez-Aguayo Cristian , Lee Ju-Seog , Lopez-Berestein Gabriel , Coleman Robert L. , Sood Anil K.
Published: April 2023

Supplementary Data Word Document
Figure S2 from Blockade of Cell Volume Regulatory Protein NKCC1 Increases TMZ-Induced Glioma Apoptosis and Reduces Astrogliosis
Authors: Luo Lanxin , Guan Xiudong , Begum Gulnaz , Ding Dawei , Gayden Jenesis , Hasan Md Nabiul , Fiesler Victoria M. , Dodelson Jacob , Kohanbash Gary , Hu Baoli , Amankulor Nduka M. , Jia Wang , Castro Maria G. , Sun Baoshan , Sun Dandan
Published: April 2023

Fig. S2. TMZ stimulates NKCC1 expression in glioma tissue.
Supplementary Tables S1-S2 from EWS-FLI1–regulated Serine Synthesis and Exogenous Serine are Necessary for Ewing Sarcoma Cellular Proliferation and Tumor Growth
Authors: Issaq Sameer H. , Mendoza Arnulfo , Kidner Ria , Rosales Tracy I. , Duveau Damien Y. , Heske Christine M. , Rohde Jason M. , Boxer Matthew B. , Thomas Craig J. , DeBerardinis Ralph J. , Helman Lee J.
Published: April 2023

Supplementary Table S1. Evaluation of drug combinations in TC32 and TC71 Ewing sarcoma cells; Supplementary Table S2. Evaluation of drug combinations in EW8 Ewing sarcoma cells.
Supplementary Figure 3 from Novel Miniaturized Drug Conjugate Leverages HSP90-driven Tumor Accumulation to Overcome PI3K Inhibitor Delivery Challenges to Solid Tumors
Authors: Perino Samantha , Moreau Benoit , Freda Jessica , Cirello Amanda , White Brian H. , Quinn James M. , Kriksciukaite Kristina , Someshwar Ashwajith , Romagnoli Janel , Robinson Megan , Movassaghian Sara , Cipriani Tyler , Wooster Richard , Bilodeau Mark T. , Whalen Kerry A.
Published: April 2023

Supplementary Figure 3 shows the body weights from all efficacy studies discussed in the article.
Supplementary Figure S1 from A Phase I Study of LY3009120, a Pan-RAF Inhibitor, in Patients with Advanced or Metastatic Cancer
Authors: Sullivan Ryan J. , Hollebecque Antoine , Flaherty Keith T. , Shapiro Geoffrey I. , Rodon Ahnert Jordi , Millward Michael J. , Zhang Wei , Gao Ling , Sykes Amanda , Willard Melinda D. , Yu Danni , Schade Andrew E. , Crowe KrisAnne , Flynn Daniel L. , Kaufman Michael D. , Henry James R. , Peng Sheng-Bin , Benhadji Karim A. , Conti Ilaria , Gordon Michael S. , Tiu Ramon V. , Hong David S.
Published: April 2023

Supplementary Figure S1 shows the study design. The study consisted of Part A (dose escalation) and Part B (dose confirmation). In Part A, 8 dosing levels starting with 50 mg twice daily (BID) were originally planned. Estimation of the maximum tolerated dose was conducted using a Continual Reassessment design. In Part B, 3 confirmation cohorts were included: Cohort A: advanced unresectable and metastatic melanoma with BRAF V600X mutations after treatment with BRAFi, MEKi or the combination of BRAFi/ MEKi; Cohort B, NRAS-mutant melanoma; and Cohort C, BRAF- or KRAS-mutant non-small cell lung cancer (NSCLC).
Table S3 from EMP2 Is a Novel Regulator of Stemness in Breast Cancer Cells
Authors: Dillard Christen , Kiyohara Meagan , Mah Vei , McDermott Sean P. , Bazzoun Dana , Tsui Jessica , Chan Ann M. , Haddad Ghassan , Pellegrini Matteo , Chang Yu-Ling , Elshimali Yahya , Wu Yanyuan , Vadgama Jaydutt V. , Kim Sara R. , Goodglick Lee , Law Samuel M. , Patel Deven D. , Dhawan Puneet , O'Brien Neil A. , Gordon Lynn K. , Braun Jonathan , Lazar Gary , Wicha Max S. , Wadehra Madhuri
Published: April 2023

Selected functional pathways affected by changes in EMP2 expression.
Supplementary Methods from ASTX029, a Novel Dual-mechanism ERK Inhibitor, Modulates Both the Phosphorylation and Catalytic Activity of ERK
Authors: Munck Joanne M. , Berdini Valerio , Bevan Luke , Brothwood Jessica L. , Castro Juan , Courtin Aurélie , East Charlotte , Ferraldeschi Roberta , Heightman Tom D. , Hindley Christopher J. , Kucia-Tran Justyna , Lyons John F. , Martins Vanessa , Muench Sandra , Murray Christopher W. , Norton David , O'Reilly Marc , Reader Michael , Rees David C. , Rich Sharna J. , Richardson Caroline J. , Shah Alpesh D. , Stanczuk Lukas , Thompson Neil T. , Wilsher Nicola E. , Woolford Alison J.-A. , Wallis Nicola G.
Published: April 2023

Table listing details of antibodies used
Supplemental Data from EMP2 Is a Novel Regulator of Stemness in Breast Cancer Cells
Authors: Dillard Christen , Kiyohara Meagan , Mah Vei , McDermott Sean P. , Bazzoun Dana , Tsui Jessica , Chan Ann M. , Haddad Ghassan , Pellegrini Matteo , Chang Yu-Ling , Elshimali Yahya , Wu Yanyuan , Vadgama Jaydutt V. , Kim Sara R. , Goodglick Lee , Law Samuel M. , Patel Deven D. , Dhawan Puneet , O'Brien Neil A. , Gordon Lynn K. , Braun Jonathan , Lazar Gary , Wicha Max S. , Wadehra Madhuri
Published: April 2023

Supplemental Methods
Supplementary Data from Anti-Extra Domain B Splice Variant of Fibronectin Antibody–Drug Conjugate Eliminates Tumors with Enhanced Efficacy When Combined with Checkpoint Blockade
Authors: Hooper Andrea T. , Marquette Kimberly , Chang Chao-Pei Betty , Golas Jonathon , Jain Sadhana , Lam My-Hanh , Guffroy Magali , Leal Mauricio , Falahatpisheh Hadi , Mathur Divya , Chen Ting , Kelleher Kerry , Khandke Kiran , Muszynska Elwira , Loganzo Frank , Rosfjord Edward , Lucas Judy , Kan Zhengyan , Subramanyam Chakrapani , O'Donnell Christopher , Neri Dario , Gerber Hans-Peter , May Chad , Sapra Puja
Published: April 2023

Supplementary Data from Anti-Extra Domain B Splice Variant of Fibronectin Antibody–Drug Conjugate Eliminates Tumors with Enhanced Efficacy When Combined with Checkpoint Blockade
Supplementary Data from An Enzymatically Cleavable Tripeptide Linker for Maximizing the Therapeutic Index of Antibody–Drug Conjugates
Authors: Ha Summer Y.Y. , Anami Yasuaki , Yamazaki Chisato M. , Xiong Wei , Haase Candice M. , Olson Scott D. , Lee Jangsoon , Ueno Naoto T. , Zhang Ningyan , An Zhiqiang , Tsuchikama Kyoji
Published: April 2023

Supplementary Data from An Enzymatically Cleavable Tripeptide Linker for Maximizing the Therapeutic Index of Antibody–Drug Conjugates
Supplementary Tables from High-Throughput Functional Evaluation of MAP2K1 Variants in Cancer
Authors: Mizuno Sho , Ikegami Masachika , Koyama Takafumi , Sunami Kuniko , Ogata Dai , Kage Hidenori , Yanagaki Mitsuru , Ikeuchi Hiroshi , Ueno Toshihide , Tanikawa Michihiro , Oda Katsutoshi , Osuga Yutaka , Mano Hiroyuki , Kohsaka Shinji
Published: April 2023

Table S1 to S9
Supplementary Figure from CDK7 Inhibition Synergizes with Topoisomerase I Inhibition in Small Cell Lung Cancer Cells by Inducing Ubiquitin-Mediated Proteolysis of RNA Polymerase II
Authors: Sun Yilun , Zhang Yang , Schultz Christopher W. , Pommier Yves , Thomas Anish
Published: April 2023

Supplementary Figure from CDK7 Inhibition Synergizes with Topoisomerase I Inhibition in Small Cell Lung Cancer Cells by Inducing Ubiquitin-Mediated Proteolysis of RNA Polymerase II
Supplementary Table S1 from Efficacious Combination Drug Treatment for Colorectal Cancer That Overcomes Resistance to KRAS G12C Inhibitors
Authors: Matsubara Hiroyuki , Miyoshi Hiroyuki , Kakizaki Fumihiko , Morimoto Tomonori , Kawada Kenji , Yamamoto Takehito , Obama Kazutaka , Sakai Yoshiharu , Taketo Makoto Mark
Published: April 2023

Suppl. Table S1.
Figure S7 from Inhibition of LILRB2 by a Novel Blocking Antibody Designed to Reprogram Immunosuppressive Macrophages to Drive T-Cell Activation in Tumors
Authors: Umiker Ben , Hashambhoy-Ramsay Yasmin , Smith Jeff , Rahman Tanzila , Mueller Amy , Davidson Rachel , Meyer Christina , Patankar Gayatri , Alam Mohammad Murshid , Jaffe Sarah , Krukenberg Kristin , Goodman Allison , Spaulding Vikki , Priess Michelle , Dhaneshwar Abha , Wong Masie , Diiorio Alexa , O'Malley Kristin , McGrath Lara , Willer Margaret , Pepper Lauren , Gostissa Monica , Kis-Toth Katalin , Wiederschain Dmitri , Cohen Heather , Shaffer Donald R.
Published: April 2023

A set of 23 anti-LILRB2 antibodies were tested for their ability to block HLA-G tetramer binding to LILRB2 and induce TNF alpha in a macrophage activation assay.
Data from Efficacious Combination Drug Treatment for Colorectal Cancer That Overcomes Resistance to KRAS G12C Inhibitors
Authors: Matsubara Hiroyuki , Miyoshi Hiroyuki , Kakizaki Fumihiko , Morimoto Tomonori , Kawada Kenji , Yamamoto Takehito , Obama Kazutaka , Sakai Yoshiharu , Taketo Makoto Mark
Published: April 2023

Abstract
Recent advances in combinatorial chemistry led to the discovery of inhibitors targeting the KRAS G12C-mutant protein. However, efficacy of its monotherapy on colorectal cancer is limited. Thus, effective combination drugs should be explored for applicable patients with colorectal cancer to fully benefit from the KRAS G12C inhibitor treatment. Here we used a patient-derived colorectal cancer stem cell (PD–CRC-SC) spheroid culture and showed that three-drug combination of inhibitors against KRAS G12C, EGFR, and FGFR synergistically suppressed the growth of colorectal cancer cells carrying the KRAS G12C mutation. Likewise, a combination of KRAS G12C and SHP2 inhibitors was also effective. Importantly, activation of the PI3K/AKT pathway in heregulin-responsive colorectal cancer cells canceled out the effect of KRAS G12C inhibition, which was largely overcome by PI3K inhibitors. These results reveal that evaluating efficacy of combination therapies with PD–CRC-SC spheroids can be a promising strategy to find the best regimen for patients with colorectal cancer.
Data from miR-20a Regulates FAS Expression in Osteosarcoma Cells by Modulating FAS Promoter Activity and Can be Therapeutically Targeted to Inhibit Lung Metastases
Authors: Yang Yuanzheng , Huang Gangxiong , Zhou Zhichao , Fewell Jason G. , Kleinerman Eugenie S.
Published: April 2023

Abstract
The metastatic potential of osteosarcoma cells is inversely correlated to cell surface FAS expression. Downregulation of FAS allows osteosarcoma cells to escape FAS ligand–mediated apoptosis when they enter a FAS ligand–positive microenvironment such as the lung. We have previously demonstrated that miR-20a, encoded by the miR-17-92 cluster, downregulates FAS expression in osteosarcoma. We further demonstrated an inverse correlation between FAS expression and miR-20a expression. However, the mechanism of FAS regulation by miR-20a was still unclear. The purpose of the current study was to evaluate the mechanism of FAS regulation by miR-20a in vitro and test the effect of targeting miR-20a in vivo. We investigated whether miR-20a's downregulation of FAS was mediated by binding to the 3′-untranslated region (3′-UTR) of FAS mRNA with the consequent induction of mRNA degradation or translational suppression. We identified and mutated two miR-20a binding sites on the FAS mRNA 3′-UTR. Using luciferase reporter assays, we demonstrated that miR-20a did not bind to either the wild-type or mutated FAS 3′-UTR. In contrast, overexpression of miR-20a resulted in downregulation of FAS promoter activity. Similarly, the inhibition of miR-20a increased FAS promoter activity. The critical region identified on the FAS promoter was between −240 bp and −150 bp. Delivery of anti-miR-20a in vivo using nanoparticles in mice with established osteosarcoma lung metastases resulted in upregulation of FAS and tumor growth inhibition. Taken together, our data suggest that miR-20a regulates FAS expression through the modulation of the FAS promoter and that targeting miR-20a using anti-miR-20a has therapeutic potential. Mol Cancer Ther; 17(1); 130–9. ©2017 AACR.
Data from Noninvasive Detection of ctDNA Reveals Intratumor Heterogeneity and Is Associated with Tumor Burden in Gastrointestinal Stromal Tumor
Authors: Namløs Heidi M. , Boye Kjetil , Mishkin Skyler J. , Barøy Tale , Lorenz Susanne , Bjerkehagen Bodil , Stratford Eva W. , Munthe Else , Kudlow Brian A. , Myklebost Ola , Meza-Zepeda Leonardo A.
Published: April 2023

Abstract
Molecular analysis of circulating tumor DNA (ctDNA) has a large potential for clinical application by capturing tumor-specific aberrations through noninvasive sampling. In gastrointestinal stromal tumor (GIST), analysis of KIT and PDGFRA mutations is important for therapeutic decisions, but the invasiveness of traditional biopsies limits the possibilities for repeated sampling. Using targeted next-generation sequencing, we have analyzed circulating cell-free DNA from 50 GIST patients. Tumor-specific mutations were detected in 16 of 44 plasma samples (36%) from treatment-naïve patients and in three of six (50%) patients treated with tyrosine kinase inhibitors. A significant association between detection of ctDNA and the modified National Institutes of Health risk classification was found. All patients with metastatic disease had detectable ctDNA, and tumor burden was the most important detection determinant. Median tumor size was 13.4 cm for patients with detectable mutation in plasma compared with 4.4 cm in patients without detectable mutation (P = 0.006). ctDNA analysis of a patient with disease progression on imatinib revealed that multiple resistance mutations were synchronously present, and detailed analysis of tumor tissue showed that these were spatially distributed in the primary tumor. Plasma samples taken throughout the course of treatment demonstrated that clonal evolution can be monitored over time. In conclusion, we have shown that detection of GIST-specific mutations in plasma is particularly feasible for patients with high tumor burden. In such cases, we have demonstrated that mutational analysis by use of liquid biopsies can capture the molecular heterogeneity of the whole tumor, and may guide treatment decisions during progression. Mol Cancer Ther; 17(11); 2473–80. ©2018 AACR.
Data from A Raf-Competitive K-Ras Binder Can Fail to Functionally Antagonize Signaling
Authors: Kauke Monique J. , Tisdale Alison W. , Kelly Ryan L. , Braun Christian J. , Hemann Michael T. , Wittrup K. Dane
Published: April 2023

Abstract
Mutated in approximately 30% of human cancers, Ras GTPases are the most common drivers of oncogenesis and render tumors unresponsive to many standard therapies. Despite decades of research, no drugs directly targeting Ras are currently available. We have previously characterized a small protein antagonist of K-Ras, R11.1.6, and demonstrated its direct competition with Raf for Ras binding. Here we evaluate the effects of R11.1.6 on Ras signaling and cellular proliferation in a panel of human cancer cell lines. Through lentiviral transduction, we generated cell lines that constitutively or through induction with doxycycline express R11.1.6 or a control protein YW1 and show specific binding by R11.1.6 to endogenous Ras through microscopy and co-immunoprecipitation experiments. Genetically encoded intracellular expression of this high-affinity Ras antagonist, however, fails to measurably disrupt signaling through either the MAPK or PI3K pathway. Consistently, cellular proliferation was unaffected as well. To understand this lack of signaling inhibition, we quantified the number of molecules of R11.1.6 expressed by the inducible cell lines and developed a simple mathematical model describing the competitive binding of Ras by R11.1.6 and Raf. This model supports a potential mechanism for the lack of biological effects that we observed, suggesting stoichiometric and thermodynamic barriers that should be overcome in pharmacologic efforts to directly compete with downstream effector proteins localized to membranes at very high effective concentrations. Mol Cancer Ther; 17(8); 1773–80. ©2018 AACR.
Data from Treating Tissue Factor–Positive Cancers with Antibody–Drug Conjugates That Do Not Affect Blood Clotting
Authors: Theunissen Jan-Willem , Cai Allen G. , Bhatti Maryam M. , Cooper Anthony B. , Avery Andrew D. , Dorfman Ryan , Guelman Sebastian , Levashova Zoia , Migone Thi-Sau
Published: April 2023

Abstract
The primary function of tissue factor (TF) resides in the vasculature as a cofactor of blood clotting; however, multiple solid tumors aberrantly express this transmembrane receptor on the cell surface. Here, we developed anti-TF antibody–drug conjugates (ADC) that did not interfere with the coagulation cascade and benchmarked them against previously developed anti-TF ADCs. After screening an affinity-matured antibody panel of diverse paratopes and affinities, we identified one primary paratope family that did not inhibit conversion of Factor X (FX) to activated Factor X (FXa) and did not affect conversion of prothrombin to thrombin. The rest of the antibody panel and previously developed anti-TF antibodies were found to perturb coagulation to varying degrees. To compare the anticancer activity of coagulation-inert and -inhibitory antibodies as ADCs, a selection of antibodies was conjugated to the prototypic cytotoxic agent monomethyl auristatin E (MMAE) through a protease-cleavable linker. The coagulation-inert and -inhibitory anti-TF ADCs both killed cancer cells effectively. Importantly, the coagulation-inert ADCs were as efficacious as tisotumab vedotin, a clinical stage ADC that affected blood clotting, including in patient-derived xenografts from three solid tumor indications with a need for new therapeutic treatments—squamous cell carcinoma of the head and neck (SCCHN), ovarian, and gastric adenocarcinoma. Furthermore, a subset of the anti-TF antibodies could also be considered for the treatment of other diseases associated with upregulation of membranous TF expression, such as macular degeneration. Mol Cancer Ther; 17(11); 2412–26. ©2018 AACR.
Data from KRAS and EGFR Amplifications Mediate Resistance to Rociletinib and Osimertinib in Acquired Afatinib-Resistant NSCLC Harboring Exon 19 Deletion/T790M in EGFR
Authors: Nakatani Kaori , Yamaoka Toshimitsu , Ohba Motoi , Fujita Ken-Ichi , Arata Satoru , Kusumoto Sojiro , Taki-Takemoto Iori , Kamei Daisuke , Iwai Shinichi , Tsurutani Junji , Ohmori Tohru
Published: April 2023

Abstract
The critical T790M mutation in EGFR, which mediates resistance to first- and second-generation EGFR tyrosine kinase inhibitors (TKI; gefitinib, erlotinib, and afatinib), has facilitated the development of third-generation mutation-selective EGFR TKIs (rociletinib and osimertinib). We previously reported heterogeneous afatinib-resistant mechanisms, including emergence of T790M-EGFR, and responses to third-generation EGFR TKIs. Here, we used afatinib-resistant lung adenocarcinoma cells [AfaR (formerly AFR3) cells], carrying exon 19 deletion/T790M in EGFR. To identify the novel resistance mechanisms in post-afatinib treatment, RocR1/RocR2 and OsiR1/OsiR2 cells were established using increasing concentrations of rociletinib and osimertinib, respectively. Attenuation of exon 19 deletion and T790M was confirmed in both rociletinib-resistant cells; in addition, EGFR and KRAS amplification was observed in RocR1 and RocR2, respectively. Significant KRAS amplification was observed in the osimertinib-resistant cell lines, indicating a linear and reversible increase with increased osimertinib concentrations in OsiR1 and OsiR2 cells. OsiR1 cells maintained osimertinib resistance with KRAS amplification after osimertinib withdrawal for 2 months. OsiR2 cells exhibited KRAS attenuation, and osimertinib sensitivity was entirely recovered. Phospho-EGFR (Y1068) and growth factor receptor–bound protein 2 (GRB2)/son of sevenless homolog 1 (SOS1) complex was found to mediate osimertinib resistance in OsiR1 cells with sustained KRAS activation. After 2 months of osimertinib withdrawal, this complex was dissociated, and the EGFR signal, but not the GRB2/SOS1 signal, was activated. Concomitant inhibition of MAPK kinase and EGFR could overcome osimertinib resistance. Thus, we identified a heterogeneous acquired resistance mechanism for third-generation EGFR TKIs, providing insights into the development of novel treatment strategies.
Data from The Protein Tyrosine Phosphatase Activity of Eyes Absent Contributes to Tumor Angiogenesis and Tumor Growth
Authors: Wang Yuhua , Pandey Ram Naresh , Riffle Stephen , Chintala Hemabindu , Wikenheiser-Brokamp Kathryn A. , Hegde Rashmi S.
Published: April 2023

Abstract
DNA damage repair capacity is required for cells to survive catastrophic DNA damage and proliferate under conditions of intratumoral stress. The ability of the minor histone protein H2AX to serve as a hub for the assembly of a productive DNA damage repair complex is a necessary step in preventing DNA damage–induced cell death. The Eyes Absent (EYA) proteins dephosphorylate the terminal tyrosine residue of H2AX, thus permitting assembly of a productive DNA repair complex. Here, we use genetic and chemical biology approaches to separately query the roles of host vascular endothelial cell and tumor cell EYA in tumor growth. Deletion of Eya3 in host endothelial cells significantly reduced tumor angiogenesis and limited tumor growth in xenografts. Deletion of Eya3 in tumor cells reduced tumor cell proliferation and tumor growth without affecting tumor angiogenesis. A chemical inhibitor of the EYA tyrosine phosphatase activity inhibited both tumor angiogenesis and tumor growth. Simultaneously targeting the tumor vasculature and tumor cells is an attractive therapeutic strategy because it could counter the development of the more aggressive phenotype known to emerge from conventional antiangiogenic agents. Mol Cancer Ther; 17(8); 1659–69. ©2018 AACR.
Data from Exploiting Radiation-Induced Signaling to Increase the Susceptibility of Resistant Cancer Cells to Targeted Drugs: AKT and mTOR Inhibitors as an Example
Authors: Eke Iris , Makinde Adeola Y. , Aryankalayil Molykutty J. , Sandfort Veit , Palayoor Sanjeewani T. , Rath Barbara H. , Liotta Lance , Pierobon Mariaelena , Petricoin Emanuel F. , Brown Matthew F. , Stommel Jayne M. , Ahmed Mansoor M. , Coleman C. Norman
Published: April 2023

Abstract
Implementing targeted drug therapy in radio-oncologic treatment regimens has greatly improved the outcome of cancer patients. However, the efficacy of molecular targeted drugs such as inhibitory antibodies or small molecule inhibitors essentially depends on target expression and activity, which both can change during the course of treatment. Radiotherapy has previously been shown to activate prosurvival pathways, which can help tumor cells to adapt and thereby survive treatment. Therefore, we aimed to identify changes in signaling induced by radiation and evaluate the potential of targeting these changes with small molecules to increase the therapeutic efficacy on cancer cell survival. Analysis of “The Cancer Genome Atlas” database disclosed a significant overexpression of AKT1, AKT2, and MTOR genes in human prostate cancer samples compared with normal prostate gland tissue. Multifractionated radiation of three-dimensional–cultured prostate cancer cell lines with a dose of 2 Gy/day as a clinically relevant schedule resulted in an increased protein phosphorylation and enhanced protein–protein interaction between AKT and mTOR, whereas gene expression of AKT, MTOR, and related kinases was not altered by radiation. Similar results were found in a xenograft model of prostate cancer. Pharmacologic inhibition of mTOR/AKT signaling after activation by multifractionated radiation was more effective than treatment prior to radiotherapy. Taken together, our findings provide a proof-of-concept that targeting signaling molecules after activation by radiotherapy may be a novel and promising treatment strategy for cancers treated with multifractionated radiation regimens such as prostate cancer to increase the sensitivity of tumor cells to molecular targeted drugs. Mol Cancer Ther; 17(2); 355–67. ©2017 AACR.

See all articles in this MCT Focus section, “Developmental Therapeutics in Radiation Oncology.”
Data from “Wnt/β-Catenin in GIST”—Letter
Authors: Bertucci François , Finetti Pascal , De Nonneville Alexandre , Birnbaum Daniel
Published: April 2023

Abstract
In an article published on September, 2017, in Molecular Cancer Therapeutics, Zeng and colleagues showed that the WNT/β-catenin oncogenic pathway was activated in a subset of human gastrointestinal stromal tumors (GIST) and that inhibiting its signaling alone or in combination with imatinib has antitumor efficacy in vitro and in vivo in imatinib-sensitive and resistant preclinical models. However, they concluded that “more investigation is needed to correlate β-catenin activation with clinicopathologic features in GIST clinical samples.” Here, we examined the activation score of the β-catenin pathway in 160 clinically annotated clinical samples of operated primary GISTs. We showed that the β-catenin activation score, assessed as continuous variable, was heterogeneous across samples. Higher score was associated with certain prognostic clinicopathologic characteristics, including mutational status, tumor size, and AFIP classification, and even more importantly, with more postoperative relapses in uni- and multivariate analyses. Such unfavorable independent prognostic value of β-catenin activation reinforces the potential therapeutic value of this new target in GIST and nicely complements Zeng's study. Mol Cancer Ther; 17(1); 327–8. ©2018 AACR.
Data from Pharmacological and Structural Characterizations of Naquotinib, a Novel Third-Generation EGFR Tyrosine Kinase Inhibitor, in EGFR-Mutated Non–Small Cell Lung Cancer
Authors: Hirano Toshiyuki , Yasuda Hiroyuki , Hamamoto Junko , Nukaga Shigenari , Masuzawa Keita , Kawada Ichiro , Naoki Katsuhiko , Niimi Tatsuya , Mimasu Shinya , Sakagami Hideki , Soejima Kenzo , Betsuyaku Tomoko
Published: April 2023

Abstract
Multiple epidermal growth factor receptor (EGFR) tyrosine kinase inhibitors (EGFR-TKI) have been developed to effectively inhibit EGFR-derived signals in non–small cell lung cancer (NSCLC). In this study, we assessed the efficacy of EGFR-TKIs, including a novel third-generation inhibitor naquotinib (ASP8273), in clinically relevant EGFR mutations, including L858R, exon 19 deletion, L858R+T790M, exon 19 deletion+T790M with or without a C797S mutation, and several exon 20 insertion mutations. Using structural analyses, we also elucidated the mechanism of activation and sensitivity/resistance to EGFR-TKIs in EGFR exon 20 insertion mutations. The efficacy of naquotinib in cells with L858R, exon 19 deletion and exon 19 deletion+T790M was comparable with that of osimertinib. Interestingly, naquotinib was more potent than osimertinib for L858R+T790M. Additionally, naquotinib and osimertinib had comparable efficacy and a wide therapeutic window for cells with EGFR exon 20 insertions. Structural modeling partly elucidated the mechanism of activation and sensitivity/resistance to EGFR-TKIs in two EGFR exon 20 insertion mutants, A767_V769dupASV and Y764_V765insHH. In summary, we have characterized the efficacy of EGFR-TKIs for NSCLC using in vitro and structural analyses and suggested the mechanism of activation and resistance to EGFR-TKIs of EGFR exon 20 insertion mutations. Our findings should guide the selection of appropriate EGFR-TKIs for the treatment of NSCLC with EGFR mutations and help clarify the biology of EGFR exon 20 insertion mutations. Mol Cancer Ther; 17(4); 740–50. ©2018 AACR.
Data from Leelamine Is a Novel Lipogenesis Inhibitor in Prostate Cancer Cells In Vitro and In Vivo
Authors: Singh Krishna B. , Hahm Eun-Ryeong , Pore Subrata K. , Singh Shivendra V.
Published: April 2023

Abstract
Increased de novo synthesis of fatty acids is implicated in the pathogenesis of human prostate cancer, but a safe and effective clinical inhibitor of this metabolic pathway is still lacking. We have shown previously that leelamine (LLM) suppresses transcriptional activity of androgen receptor, which is known to regulate fatty acid synthesis. Therefore, the current study was designed to investigate the effect of LLM on fatty acid synthesis. Exposure of 22Rv1, LNCaP, and PC-3 prostate cancer cells, but not RWPE-1 normal prostate epithelial cell line, to LLM resulted in a decrease in intracellular levels of neutral lipids or total free fatty acids. LLM was superior to another fatty acid synthesis inhibitor (cerulenin) for suppression of total free fatty acid levels. LLM treatment downregulated protein and/or mRNA expression of key fatty acid synthesis enzymes, including ATP citrate lyase, acetyl-CoA carboxylase 1, fatty acid synthase, and sterol regulatory element-binding protein 1 (SREBP1) in each cell line. Consistent with these in vitro findings, we also observed a significant decrease in ATP citrate lyase and SREBP1 protein expression as well as number of neutral lipid droplets in vivo in 22Rv1 tumor sections of LLM-treated mice when compared with that of controls. LLM-mediated suppression of intracellular levels of total free fatty acids and neutral lipids was partly attenuated by overexpression of SREBP1. In conclusion, these results indicate that LLM is a novel inhibitor of SREBP1-regulated fatty acid/lipid synthesis in prostate cancer cells that is not affected by androgen receptor status.
Data from Recombinant MDA-7/IL24 Suppresses Prostate Cancer Bone Metastasis through Downregulation of the Akt/Mcl-1 Pathway
Authors: Pradhan Anjan K. , Bhoopathi Praveen , Talukdar Sarmistha , Shen Xue-Ning , Emdad Luni , Das Swadesh K. , Sarkar Devanand , Fisher Paul B.
Published: April 2023

Abstract
Prostate cancer is a principal cause of cancer-associated morbidity in men. Although 5-year survival of patients with localized prostate cancer approaches 100%, survival decreases precipitously after metastasis. Bone is the preferred site for disseminated prostate cancer cell colonization, altering the equilibrium of bone homeostasis resulting in weak and fragile bones. Currently, no curative options are available for prostate cancer bone metastasis. Melanoma differentiation associated gene-7 (MDA-7)/IL24 is a well-studied cytokine established as a therapeutic in a wide array of cancers upon delivery as a gene therapy. In this study, we explored the potential anticancer properties of MDA-7/IL24 delivered as a recombinant protein. Using bone metastasis experimental models, animals treated with recombinant MDA-7/IL24 had significantly less metastatic lesions in their femurs as compared with controls. The inhibitory effects of MDA-7/IL24 on bone metastasis resulted from prostate cancer–selective killing and inhibition of osteoclast differentiation, which is necessary for bone resorption. Gain- and loss-of-function genetic approaches document that prosurvival Akt and Mcl-1 pathways are critically important in the antibone metastatic activity of MDA-7/IL24. Our previous findings showed that MDA-7/IL24 gene therapy plus Mcl-1 inhibitors cooperate synergistically. Similarly, an Mcl-1 small-molecule inhibitor synergized with MDA-7/IL24 and induced robust antibone metastatic activity. These results expand the potential applications of MDA-7/IL24 as an anticancer molecule and demonstrate that purified recombinant protein is nontoxic in preclinical animal models and has profound inhibitory effects on bone metastasis, which can be enhanced further when combined with an Mcl-1 inhibitory small molecule. Mol Cancer Ther; 17(9); 1951–60. ©2018 AACR.
Data from Mechanistic Investigations of Diarrhea Toxicity Induced by Anti-HER2/3 Combination Therapy
Authors: Moisan Annie , Michielin Francesca , Jacob Wolfgang , Kronenberg Sven , Wilson Sabine , Avignon Blandine , Gérard Régine , Benmansour Fethallah , McIntyre Christine , Meneses-Lorente Georgina , Hasmann Max , Schneeweiss Andreas , Weisser Martin , Adessi Céline
Published: April 2023

Abstract
Combination of targeted therapies is expected to provide superior efficacy in the treatment of cancer either by enhanced antitumor activity or by preventing or delaying the development of resistance. Common challenges in developing combination therapies include the potential of additive and aggravated toxicities associated with pharmacologically related adverse effects. We have recently reported that combination of anti-HER2 and anti-HER3 antibodies, pertuzumab and lumretuzumab, along with paclitaxel chemotherapy in metastatic breast cancer, resulted in a high incidence of diarrhea that ultimately limited further clinical development of this combination. Here, we further dissected the diarrhea profile of the various patient dose cohorts and carried out in vitro investigations in human colon cell lines and explants to decipher the contribution and the mechanism of anti-HER2/3 therapeutic antibodies to intestinal epithelium malfunction. Our clinical investigations in patients revealed that while dose reduction of lumretuzumab, omission of pertuzumab loading dose, and introduction of a prophylactic antidiarrheal treatment reduced most severe adverse events, patients still suffered from persistent diarrhea during the treatment. Our in vitro investigations showed that pertuzumab and lumretuzumab combination treatment resulted in upregulation of chloride channel activity without indication of intestinal barrier disruption. Overall, our findings provide a mechanistic rationale to explore alternative of conventional antigut motility using medication targeting chloride channel activity to mitigate diarrhea of HER combination therapies. Mol Cancer Ther; 17(7); 1464–74. ©2018 AACR.
Data from Preclinical Development of a Bispecific Antibody that Safely and Effectively Targets CD19 and CD47 for the Treatment of B-Cell Lymphoma and Leukemia
Authors: Buatois Vanessa , Johnson Zoë , Salgado-Pires Susana , Papaioannou Anne , Hatterer Eric , Chauchet Xavier , Richard Françoise , Barba Leticia , Daubeuf Bruno , Cons Laura , Broyer Lucile , D'Asaro Matilde , Matthes Thomas , LeGallou Simon , Fest Thierry , Tarte Karin , Clarke Hinojosa Robert K. , Genescà Ferrer Eulàlia , Ribera José María , Dey Aditi , Bailey Katharine , Fielding Adele K. , Eissenberg Linda , Ritchey Julie , Rettig Michael , DiPersio John F. , Kosco-Vilbois Marie H. , Masternak Krzysztof , Fischer Nicolas , Shang Limin , Ferlin Walter G.
Published: April 2023

Abstract
CD47, an ubiquitously expressed innate immune checkpoint receptor that serves as a universal “don't eat me” signal of phagocytosis, is often upregulated by hematologic and solid cancers to evade immune surveillance. Development of CD47-targeted modalities is hindered by the ubiquitous expression of the target, often leading to rapid drug elimination and hemotoxicity including anemia. To overcome such liabilities, we have developed a fully human bispecific antibody, NI-1701, designed to coengage CD47 and CD19 selectively on B cells. NI-1701 demonstrates favorable elimination kinetics with no deleterious effects seen on hematologic parameters following single or multiple administrations to nonhuman primates. Potent in vitro and in vivo activity is induced by NI-1701 to kill cancer cells across a plethora of B-cell malignancies and control tumor growth in xenograft mouse models. The mechanism affording maximal tumor growth inhibition by NI-1701 is dependent on the coengagement of CD47/CD19 on B cells inducing potent antibody-dependent cellular phagocytosis of the targeted cells. NI-1701–induced control of tumor growth in immunodeficient NOD/SCID mice was more effective than that achieved with the anti-CD20 targeted antibody, rituximab. Interestingly, a synergistic effect was seen when tumor-implanted mice were coadministered NI-1701 and rituximab leading to significantly improved tumor growth inhibition and regression in some animals. We describe herein, a novel bispecific antibody approach aimed at sensitizing B cells to become more readily phagocytosed and eliminated thus offering an alternative or adjunct therapeutic option to patients with B-cell malignancies refractory/resistant to anti-CD20–targeted therapy. Mol Cancer Ther; 17(8); 1739–51. ©2018 AACR.
Data from Analysis of Tissue and Circulating Tumor DNA by Next-Generation Sequencing of Hepatocellular Carcinoma: Implications for Targeted Therapeutics
Authors: Ikeda Sadakatsu , Lim Jordan S. , Kurzrock Razelle
Published: April 2023

Abstract
Hepatocellular carcinoma (HCC) has limited treatment options. Molecular analysis of its mutational landscape may enable the identification of novel therapies. However, biopsy is not routinely performed in HCC. The utility of analyzing cell-free circulating tumor DNA (ctDNA) by next-generation sequencing (NGS) is not established. We performed 32 ctDNA NGS analyses on 26 patients; 10 of these patients had tissue NGS (236 to 626 genes). ctDNA was evaluated using an assay that detects single nucleotide variants, amplifications, fusions, and specific insertion/deletion alterations in 54 to 70 genes. The ctDNA demonstrated that 23 of 26 patients (88.5%) had ≥1 characterized alteration, and all these individuals had ≥1 potentially actionable alteration. The most frequently mutated gene was TP53 (16 of 26 patients, 61.5%). There were 47 unique characterized molecular alterations among 18 total gene alterations [variants of unknown significance (VUS) excluded)]. ctDNA and tissue NGS frequently showed different profiles, perhaps due to length of time between tissue and blood samples [median = 370 days (range, 29 to 876 days)]. Serial ctDNA evaluation in an illustrative patient treated with capecitabine demonstrated emergence of a new TP53 alteration after progression. In conclusion, ctDNA profiling is feasible in advanced HCC, and serial assessment using ctDNA NGS can reveal genomic changes with time. NGS of ctDNA provides a minimally invasive alternative for identifying potentially actionable gene alterations and potential molecular targeted therapies. Dynamic changes in molecular portfolio associated with therapeutic pressure in difficult-to-biopsy patients can be observed. Mol Cancer Ther; 17(5); 1114–22. ©2018 AACR.
Data from Theranostic Radiolabeled Anti-CD20 sdAb for Targeted Radionuclide Therapy of Non-Hodgkin Lymphoma
Authors: Krasniqi Ahmet , D'Huyvetter Matthias , Xavier Catarina , Van der Jeught Kevin , Muyldermans Serge , Van Der Heyden José , Lahoutte Tony , Tavernier Jan , Devoogdt Nick
Published: April 2023

Abstract
Anti-CD20 radioimmunotherapy is an effective approach for therapy of relapsed or refractory CD20pos lymphomas, but faces limitations due to poor tumor penetration and undesirable pharmacokinetics of full antibodies. Camelid single-domain Ab fragments (sdAb) might circumvent some of the limitations of radiolabeled full antibodies. In this study, a set of hCD20-targeting sdAbs was generated, and their capacity to bind hCD20 was evaluated in vitro and in vivo. A lead sdAb, sdAb 9079, was selected on the basis of its specific tumor targeting and significant lower kidney accumulation compared with other sdAbs. SdAb 9079 was then radiolabeled with 68Ga and 177Lu for PET imaging and targeted therapy. The therapeutic potential of 177Lu-DTPA-sdAb was compared with that of 177Lu-DTPA-rituximab and unlabeled rituximab in mice bearing hCD20pos tumors. Radiolabeled with 68Ga, sdAb 9079 showed specific tumor uptake, with very low accumulation in nontarget organs, except kidneys. The tumor uptake of 177Lu-DTPA-sdAb 9079 after 1.5 h was 3.4 ± 1.3% ID/g, with T/B and T/M ratios of 13.3 ± 4.6 and 32.9 ± 15.6. Peak tumor accumulation of 177Lu-DTPA-rituximab was about 9 times higher, but concomitantly with high accumulation in nontarget organs and very low T/B and T/M ratios (0.8 ± 0.1 and 7.1 ± 2.4). Treatment of mice with 177Lu-DTPA-sdAb 9079 significantly prolonged median survival compared with control groups and was as effective as treatment with rituximab or its 177Lu-labeled variant. Taken together, sdAb 9079 displays promising features as a theranostic drug to treat CD20pos lymphomas. Mol Cancer Ther; 16(12); 2828–39. ©2017 AACR.
Data from Afatinib Is a New Therapeutic Approach in Chordoma with a Unique Ability to Target EGFR and Brachyury
Authors: Magnaghi Paola , Salom Barbara , Cozzi Liviana , Amboldi Nadia , Ballinari Dario , Tamborini Elena , Gasparri Fabio , Montagnoli Alessia , Raddrizzani Laura , Somaschini Alessio , Bosotti Roberta , Orrenius Christian , Bozzi Fabio , Pilotti Silvana , Galvani Arturo , Sommer Josh , Stacchiotti Silvia , Isacchi Antonella
Published: April 2023

Abstract
Chordomas are rare bone tumors with no approved therapy. These tumors express several activated tyrosine kinase receptors, which prompted attempts to treat patients with tyrosine kinase inhibitors. Although clinical benefit was observed in phase II clinical trials with imatinib and sorafenib, and sporadically also with EGFR inhibitors, therapies evaluated to date have shown modest activity. With the goal of identifying new drugs with immediate therapeutic potential for chordoma patients, we collected clinically approved drugs and other advanced inhibitors of MET, PDGFRβ, and EGFR tyrosine kinases, and assessed their antiproliferative activity against a panel of chordoma cell lines. Chordoma cell lines were not responsive to MET and PDGFRβ inhibitors. U-CH1 and UM-Chor1 were sensitive to all EGFR inhibitors, whereas the remaining cell lines were generally insensitive to these drugs. Afatinib was the only EGFR inhibitor with activity across the chordoma panel. We then investigated the molecular mechanisms behind the responses observed and found that the antiproliferative IC50s correlate with the unique ability of afatinib to promote degradation of EGFR and brachyury, an embryonic transcription factor considered a key driver of chordoma. Afatinib displayed potent antitumor efficacy in U-CH1, SF8894, CF322, and CF365 chordoma tumor models in vivo. In the panel analyzed, high EGFR phosphorylation and low AXL and STK33 expression correlated with higher sensitivity to afatinib and deserve further investigation as potential biomarkers of response. These data support the use of afatinib in clinical trials and provide the rationale for the upcoming European phase II study on afatinib in advanced chordoma. Mol Cancer Ther; 17(3); 603–13. ©2017 AACR.
Data from Altered Steroid Milieu in AI-Resistant Breast Cancer Facilitates AR Mediated Gene-Expression Associated with Poor Response to Therapy
Authors: Creevey Laura , Bleach Rachel , Madden Stephen F. , Toomey Sinead , Bane Fiona T. , Varešlija Damir , Hill Arnold D. , Young Leonie S. , McIlroy Marie
Published: April 2023

Abstract
Divergent roles for androgen receptor (AR) in breast cancer have been reported. Following aromatase inhibitor (AI) treatment, the conversion of circulating androgens into estrogens can be diminished by >99%. We wished to establish whether the steroid environment can dictate the role of AR and the implications of this for subsequent therapy. This study utilizes models of AI resistance to explore responsiveness to PI3K/mTOR and anti-AR therapy when cells are exposed to unconverted weak androgens. Transcriptomic alterations driven by androstenedione (4AD) were assessed by RNA-sequencing. AR and estrogen receptor (ER) recruitment to target gene promoters was evaluated using ChIP, and relevance to patient profiles was performed using publicly available data sets. Although BEZ235 showed decreased viability across AI-sensitive and -resistant cell lines, anti-AR treatment elicited a decrease in cell viability only in the AI-resistant model. Serum and glucocorticoid-regulated kinase 3 (SGK3) and cAMP-dependent protein kinase inhibitor β (PKIB) were confirmed to be regulated by 4AD and shown to be mediated by AR; crucially, reexposure to estradiol suppressed expression of these genes. Meta-analysis of transcript levels showed high expression of SGK3 and PKIB to be associated with poor response to endocrine therapy (HR = 2.551, P = 0.003). Furthermore, this study found levels of SGK3 to be sustained in patients who do not respond to AI therapy. This study highlights the importance of the tumor steroid environment. SGK3 and PKIB are associated with poor response to endocrine therapy and could have utility in tailoring therapeutic approaches.