bluuebunny/crossref_metadata_2025 huggingface dataset:
Data from CSPG4-Specific Immunity and Survival Prolongation in Dogs with Oral Malignant Melanoma Immunized with Human CSPG4 DNA
Authors: Riccardo Federica , Iussich Selina , Maniscalco Lorella , Lorda Mayayo Saray , La Rosa Giuseppe , Arigoni Maddalena , De Maria Raffaella , Gattino Francesca , Lanzardo Stefania , Lardone Elena , Martano Marina , Morello Emanuela , Prestigio Simone , Fiore Alessandra , Quaglino Elena , Zabarino Sara , Ferrone Soldano , Buracco Paolo , Cavallo Federica
Published: April 2023

Abstract
Purpose: Due to the many similarities with its human counterpart, canine malignant melanoma (cMM) is a valuable model in which to assess the efficacy of novel therapeutic strategies. The model is herein used to evaluate the immunogenicity, safety, and therapeutic efficacy of a human chondroitin sulfate proteoglycan-4 (hCSPG4) DNA-based vaccine. The fact that homology between hCSPG4 and cCSPG4 amino-acidic sequences stands at more than 80% provides the rationale for using an hCSPG4 DNA vaccine in the cMM model.

Experimental Design: Dogs with stage II–III surgically resected CSPG4-positive oral MM were subjected to monthly intramuscular plasmid administration, which was followed immediately by electroporation (electrovaccination) for at least 6, and up to 20, months. The immunogenicity, safety, and therapeutic efficacy of the vaccine have been evaluated.

Results: hCSPG4 electrovaccination caused no clinically relevant local or systemic side effects and resulted in significantly longer overall and disease-free survival times in 14 vaccinated dogs as compared with 13 nonvaccinated controls. All vaccinated dogs developed antibodies against both hCSPG4 and cCSPG4. Seven vaccinated dogs were also tested for a cCSPG4-specific T-cell response and only two gave a detectable interferon (IFN)γ response.

Conclusion: Xenogeneic electrovaccination against CSPG4 is able to overcome host unresponsiveness to the “self” antigen and seems to be effective in treating cMM, laying the foundation for its translation to a human clinical setting. Clin Cancer Res; 20(14); 3753–62. ©2014 AACR.
Data from A High-Throughput Pharmaceutical Screen Identifies Compounds with Specific Toxicity against BRCA2-Deficient Tumors
Authors: Evers Bastiaan , Schut Eva , van der Burg Eline , Braumuller Tanya M. , Egan David A. , Holstege Henne , Edser Pauline , Adams David J. , Wade-Martins Richard , Bouwman Peter , Jonkers Jos
Published: April 2023

Abstract
Purpose: Hereditary breast cancer is partly explained by germline mutations in BRCA1 and BRCA2. Although patients carry heterozygous mutations, their tumors have typically lost the remaining wild-type allele. Selectively targeting BRCA deficiency may therefore constitute an important therapeutic approach. Clinical trials applying this principle are underway, but it is unknown whether the compounds tested are optimal. It is therefore important to identify alternative compounds that specifically target BRCA deficiency and to test new combination therapies to establish optimal treatment strategies.

Experimental Design: We did a high-throughput pharmaceutical screen on BRCA2-deficient mouse mammary tumor cells and isogenic controls with restored BRCA2 function. Subsequently, we validated positive hits in vitro and in vivo using mice carrying BRCA2-deficient mammary tumors.

Results: Three alkylators—chlorambucil, melphalan, and nimustine—displayed strong and specific toxicity against BRCA2-deficient cells. In vivo, these showed heterogeneous but generally strong BRCA2-deficient antitumor activity, with melphalan and nimustine doing better than cisplatin and the poly-(ADP-ribose)-polymerase inhibitor olaparib (AZD2281) in this small study. In vitro drug combination experiments showed synergistic interactions between the alkylators and olaparib. Tumor intervention studies combining nimustine and olaparib resulted in recurrence-free survival exceeding 330 days in 3 of 5 animals tested.

Conclusions: We generated and validated a platform for identification of compounds with specific activity against BRCA2-deficient cells that translates well to the preclinical setting. Our data call for the re-evaluation of alkylators, especially melphalan and nimustine, alone or in combination with the poly-(ADP-ribose)-polymerase inhibitors, for the treatment of breast cancers with a defective BRCA pathway. Clin Cancer Res; 16(1); 99–108
Data from Novel Tumor Subgroups of Urothelial Carcinoma of the Bladder Defined by Integrated Genomic Analysis
Authors: Hurst Carolyn D. , Platt Fiona M. , Taylor Claire F. , Knowles Margaret A.
Published: April 2023

Abstract
Purpose: There is a need for improved subclassification of urothelial carcinoma (UC) at diagnosis. A major aim of this study was to search for novel genomic subgroups.

Experimental design: We assessed 160 tumors for genome-wide copy number alterations and mutation in genes implicated in UC. These comprised all tumor grades and stages and included 49 high-grade stage T1 (T1G3) tumors.

Results: Our findings point to the existence of genomic subclasses of the “gold-standard” grade/stage groups. The T1G3 tumors separated into 3 major subgroups that differed with respect to the type and number of copy number events and to FGFR3 and TP53 mutation status. We also identified novel regions of copy number alteration, uncovered relationships between molecular events, and elucidated relationships between molecular events and clinico-pathologic features. FGFR3 mutant tumors were more chromosomally stable than their wild-type counterparts and a mutually exclusive relationship between FGFR3 mutation and overrepresentation of 8q was observed in non-muscle-invasive tumors. In muscle-invasive (MI) tumors, metastasis was positively associated with losses of regions on 10q (including PTEN), 16q and 22q, and gains on 10p, 11q, 12p, 19p, and 19q. Concomitant copy number alterations positively associated with TP53 mutation in MI tumors were losses on 16p, 2q, 4q, 11p, 10q, 13q, 14q, 16q, and 19p, and gains on 1p, 8q, 10q, and 12q. Significant complexity was revealed in events affecting chromosome 9.

Conclusions: These findings may lead to improved biologic understanding and the development of prognostic biomarkers. Novel regions of copy number alteration may reveal potential therapeutic targets. Clin Cancer Res; 18(21); 5865–77. ©2012 AACR.
Data from BRCA2 Deep Intronic Mutation Causing Activation of a Cryptic Exon: Opening toward a New Preventive Therapeutic Strategy
Authors: Anczuków Olga , Buisson Monique , Léoné Mélanie , Coutanson Christine , Lasset Christine , Calender Alain , Sinilnikova Olga M. , Mazoyer Sylvie
Published: April 2023

Abstract
Purpose: Diagnostic screening of the BRCA1/2 genes in breast cancer families is mostly done on genomic DNA. For families with a very strong family history and no mutation identified in the coding sequences or the exon–intron boundaries, BRCA1/2 transcripts' analysis is an efficient approach to uncover gene inversion and pre-mRNA splicing defaults missed by conventional DNA-based protocols.

Experimental Design: We analyzed RNA from patients of negative BRCA families by reverse transcriptase PCR and identified an insertion in one family that we characterized by sequencing and by using a minigene splicing assay. More than 2,000 additional BRCA1/2 negative families were subsequently screened for this mutation using a dedicated PCR approach.

Results: Nine families were found to harbor a BRCA2 mutant transcript containing a 95-nucleotide cryptic exon between exons 12 and 13. This cryptic exon results from a new mutation located deep into intron 12, c.6937+594T > G, which reinforces the strength of a preexisting 5′ splice site, turning it into a perfect consensus sequence. It is systematically included in transcripts produced by the mutant allele in cells from mutation carriers or produced by a mutant splicing reporter minigene. The inclusion of the cryptic exon was prevented when we cotransfected the minigene with antisense oligonucleotides complementary to the 3′ or mutated 5′ splice sites.

Conclusion: This first deep intronic BRCA mutation emphasizes the importance of analyzing RNA to provide comprehensive BRCA1/2 diagnostic tests and opens the possibility of using antisense therapy in the future as an alternative strategy for cancer prevention. Clin Cancer Res; 18(18); 4903–9. ©2012 AACR.
Data from Hidden Dangers: A Cryptic Exon Disrupts BRCA2 mRNA
Authors: Fackenthal James D. , Lee Younghee , Olopade Olufunmilayo I.
Published: April 2023

Abstract
The first mutation that disrupts BRCA2 mRNA by including a novel, cryptic exon is reported in this issue. The mutation lies deep within an intron and would not have been detected by conventional screening methods. In the future, more mutations may be discovered by direct mRNA analysis. Clin Cancer Res; 18(18); 4865–7. ©2012 AACR.
Data from Targeting Fibroblast Growth Factor Receptor Signaling Inhibits Prostate Cancer Progression
Authors: Feng Shu , Shao Longjiang , Yu Wendong , Gavine Paul , Ittmann Michael
Published: April 2023

Abstract
Purpose: Extensive correlative studies in human prostate cancer as well as studies in vitro and in mouse models indicate that fibroblast growth factor receptor (FGFR) signaling plays an important role in prostate cancer progression. In this study, we used a probe compound for an FGFR inhibitor, which potently inhibits FGFR-1–3 and significantly inhibits FGFR-4. The purpose of this study is to determine whether targeting FGFR signaling from all four FGFRs will have in vitro activities consistent with inhibition of tumor progression and will inhibit tumor progression in vivo.

Experimental Design: Effects of AZ8010 on FGFR signaling and invasion were analyzed using immortalized normal prostate epithelial (PNT1a) cells and PNT1a overexpressing FGFR-1 or FGFR-4. The effect of AZ8010 on invasion and proliferation in vitro was also evaluated in prostate cancer cell lines. Finally, the impact of AZ8010 on tumor progression in vivo was evaluated using a VCaP xenograft model.

Results: AZ8010 completely inhibits FGFR-1 and significantly inhibits FGFR-4 signaling at 100 nmol/L, which is an achievable in vivo concentration. This results in marked inhibition of extracellular signal–regulated kinase (ERK) phosphorylation and invasion in PNT1a cells expressing FGFR-1 and FGFR-4 and all prostate cancer cell lines tested. Treatment in vivo completely inhibited VCaP tumor growth and significantly inhibited angiogenesis and proliferation and increased cell death in treated tumors. This was associated with marked inhibition of ERK phosphorylation in treated tumors.

Conclusions: Targeting FGFR signaling is a promising new approach to treating aggressive prostate cancer. Clin Cancer Res; 18(14); 3880–8. ©2012 AACR.
Data from Molecular Profiling Reveals Low- and High-Grade Forms of Primary Melanoma
Authors: Harbst Katja , Staaf Johan , Lauss Martin , Karlsson Anna , Måsbäck Anna , Johansson Iva , Bendahl Pär-Ola , Vallon-Christersson Johan , Törngren Therese , Ekedahl Henrik , Geisler Jürgen , Höglund Mattias , Ringnér Markus , Lundgren Lotta , Jirström Karin , Olsson Håkan , Ingvar Christian , Borg Åke , Tsao Hensin , Jönsson Göran
Published: April 2023

Abstract
Purpose: For primary melanomas, tumor thickness, mitotic rate, and ulceration are well-laid cornerstones of prognostication. However, a molecular exposition of melanoma aggressiveness is critically missing. We recently uncovered a four-class structure in metastatic melanoma, which predicts outcome and informs biology. This raises the possibility that a molecular structure exists even in the early stages of melanoma and that molecular determinants could underlie histophenotype and eventual patient outcome.

Experimental Design: We subjected 223 archival primary melanomas to a horizontally integrated analysis of RNA expression, oncogenic mutations at 238 lesions, histomorphometry, and survival data.

Results: Our previously described four-class structure that was elucidated in metastatic lesions was evident within the expression space of primary melanomas. Because these subclasses converged into two larger prognostic and phenotypic groups, we used the metastatic lesions to develop a binary subtype-based signature capable of distinguishing between “high” and “low” grade forms of the disease. The two-grade signature was subsequently applied to the primary melanomas. Compared with low-grade tumors, high-grade primary melanomas were significantly associated with increased tumor thickness, mitotic rate, ulceration (all P < 0.01), and poorer relapse-free (HR = 4.94; 95% CI, 2.84–8.59), and overall (HR = 3.66; 95% CI, 2.40–5.58) survival. High-grade melanomas exhibited elevated levels of proliferation and BRCA1/DNA damage signaling genes, whereas low-grade lesions harbored higher expression of immune genes. Importantly, the molecular-grade signature was validated in two external gene expression data sets.

Conclusions: We provide evidence for a molecular organization within melanomas, which is preserved across all stages of disease. Clin Cancer Res; 18(15); 4026–36. ©2012 AACR.
Data from Common PIK3CA Mutants and a Novel 3′ UTR Mutation Are Associated with Increased Sensitivity to Saracatinib
Authors: Arcaroli John J. , Quackenbush Kevin S. , Powell Rebecca W. , Pitts Todd M. , Spreafico Anna , Varella-Garcia Marileila , Bemis Lynne , Tan Aik Choon , Reinemann Jaclyn M. , Touban Basel M. , Dasari Arvind , Eckhardt S. Gail , Messersmith Wells A.
Published: April 2023

Abstract
Purpose: Dysregulation of the phosphoinositide 3-kinase (PI3K) and Src signaling pathways commonly occur in colorectal cancer. Mutations in the PIK3CA gene are associated with an increase in severity of disease and worse clinical outcomes. Elevated levels of Src have been identified in premalignant lesions and are suggested to play a central role in tumor progression. Because these pathways appear to enhance tumor growth and metastasis, molecularly targeted agents for both pathways are currently being evaluated in early-phase clinical trials.

Experimental Design: We used colorectal cancer cell lines and a patient-derived explant model to investigate the efficacy of saracatinib. Mutations in the PIK3CA were evaluated to examine the association between mutations in the PIK3CA gene and sensitivity to saracatinib.

Results: We have identified a subset of patients with a PIK3CA (exon 9 and 20) mutation with increased sensitivity to saracatinib. A novel 3′ untranslated region (UTR) mutation was also shown to be associated with increased sensitivity to saracatinib and have a reduced affinity for miR-520a and miR-525a. Importantly, we show that Src inhibition reduces the interaction between Src and p85, subsequently decreasing Akt-dependent signaling.

Conclusion: These results indicate that a personalized approach in targeting Src in PIK3CA-mutant patients with colorectal cancers may prove effective in a subset of patients with this genetic alteration. Clin Cancer Res; 18(9); 2704–14. ©2012 AACR.
Data from Effective Activity of Cytokine-Induced Killer Cells against Autologous Metastatic Melanoma Including Cells with Stemness Features
Authors: Gammaitoni Loretta , Giraudo Lidia , Leuci Valeria , Todorovic Maja , Mesiano Giulia , Picciotto Franco , Pisacane Alberto , Zaccagna Alessandro , Volpe Maria Giuseppa , Gallo Susanna , Caravelli Daniela , Giacone Elena , Venesio Tiziana , Balsamo Antonella , Pignochino Ymera , Grignani Giovanni , Carnevale-Schianca Fabrizio , Aglietta Massimo , Sangiolo Dario
Published: April 2023

Abstract
Purpose: We investigate the unknown tumor-killing activity of cytokine-induced killer (CIK) cells against autologous metastatic melanoma and the elusive subset of putative cancer stem cells (mCSC).

Experimental Design: We developed a preclinical autologous model using same patient-generated CIK cells and tumor targets to consider the unique biology of each patient/tumor pairing. In primary tumor cell cultures, we visualized and immunophenotypically defined a putative mCSC subset using a novel gene transfer strategy that exploited their exclusive ability to activate the promoter of stemness gene Oct4.

Results: The CIK cells from 10 patients with metastatic melanoma were successfully expanded (median, 23-fold; range, 11–117). Primary tumor cell cultures established and characterized from the same patients were used as autologous targets. Patient-derived CIK cells efficiently killed autologous metastatic melanoma [up to 71% specific killing (n = 26)]. CIK cells were active in vivo against autologous melanoma, resulting in delayed tumor growth, increased necrotic areas, and lymphocyte infiltration at tumor sites. The metastatic melanoma cultures presented an average of 11.5% ± 2.5% putative mCSCs, which was assessed by Oct4 promoter activity and stemness marker expression (Oct4, ABCG2, ALDH, MITF). Expression was confirmed on mCSC target molecules recognized by CIK cells (MIC A/B; ULBPs). CIK tumor killing activity against mCSCs was intense (up to 71%, n = 4) and comparable with results reported against differentiated metastatic melanoma cells (P = 0.8).

Conclusions: For the first time, the intense killing activity of CIK cells against autologous metastatic melanoma, including mCSCs, has been shown. These findings move clinical investigation of a new immunotherapy for metastatic melanoma, including mCSCs, closer. Clin Cancer Res; 19(16); 4347–58. ©2013 AACR.
Data from Developing Standards for Breakthrough Therapy Designation in Oncology
Authors: Horning Sandra J. , Haber Daniel A. , Selig Wendy K.D. , Ivy S. Percy , Roberts Samantha A. , Allen Jeff D. , Sigal Ellen V. , Sawyers Charles L.
Published: April 2023

Abstract
In July 2012, Congress passed the Food and Drug Administration Safety and Innovation Act (FDASIA). The Advancing Breakthrough Therapies for Patients Act was incorporated into a Title of FDASIA to expedite clinical development of new, potential “breakthrough” drugs or treatments that show dramatic responses in early-phase studies. Using this regulatory pathway, once a promising new drug candidate is designated as a “Breakthrough Therapy”, the U.S. Food and Drug Administration (FDA) and sponsor would collaborate to determine the best path forward to abbreviate the traditional three-phase approach to drug development. The breakthrough legislation requires that an FDA guidance be drafted that details specific requirements of the bill to aid FDA in implementing requirements of the Act. In this article, we have proposed criteria to define a product as a Breakthrough Therapy, and discussed critical components of the development process that would require flexibility in order to enable expedited development of a Breakthrough Therapy. Clin Cancer Res; 19(16); 4297–304. ©2013 AACR.
Data from Epigenetic Alteration of PRKCDBP in Colorectal Cancers and Its Implication in Tumor Cell Resistance to TNFα-Induced Apoptosis
Authors: Lee Jin-Hee , Kang Min-Ju , Han Hye-Yeon , Lee Min-Goo , Jeong Seong-In , Ryu Byung-Kyu , Ha Tae-Kyu , Her Nam-Goo , Han Jikhyon , Park Sun Jin , Lee Kil Yeon , Kim Hyo-Jong , Chi Sung-Gil
Published: April 2023

Abstract
Purpose:PRKCDBP is a putative tumor suppressor in which alteration has been observed in several human cancers. We investigated expression and function of PRKCDBP in colorectal cells and tissues to explore its candidacy as a suppressor in colorectal tumorigenesis.

Experimental Design: Expression and methylation status of PRKCDBP and its effect on tumor growth were evaluated. Transcriptional regulation by NF-κB signaling was defined by luciferase reporter and chromatin immunoprecipitation assays.

Results: PRKCDBP expression was hardly detectable in 29 of 80 (36%) primary tumors and 11 of 19 (58%) cell lines, and its alteration correlated with tumor stage and grade. Promoter hypermethylation was commonly found in cancers. PRKCDBP expression induced the G1 cell-cycle arrest and increased cellular sensitivity to various apoptotic stresses. PRKCDBP was induced by TNFα, and its level correlated with tumor cell sensitivity to TNFα-induced apoptosis. PRKCDBP induction by TNFα was disrupted by blocking NF-κB signaling while it was enhanced by RelA transfection. The PRKCDBP promoter activity was increased in response to TNFα, and this response was abolished by disruption of a κB site in the promoter. PRKCDBP delayed the formation and growth of xenograft tumors and improved tumor response to TNFα-induced apoptosis.

Conclusions:PRKCDBP is a proapoptotic tumor suppressor which is commonly altered in colorectal cancer by promoter hypermethylation, and its gene transcription is directly activated by NF-κB in response to TNFα. This suggests that PRKCDBP inactivation may contribute to tumor progression by reducing cellular sensitivity to TNFα and other stresses, particularly under chronic inflammatory microenvironment. Clin Cancer Res; 17(24); 7551–62. ©2011 AACR.
Data from Potent Antimyeloma Activity of a Novel ERK5/CDK Inhibitor
Authors: Álvarez-Fernández Stela , Ortiz-Ruiz María Jesús , Parrott Tracy , Zaknoen Sara , Ocio Enrique M. , San Miguel Jesús , Burrows Francis J. , Esparís-Ogando Azucena , Pandiella Atanasio
Published: April 2023

Abstract
Purpose: To analyze the antimyeloma potential of TG02, an ERK5/CDK inhibitory drug.

Experimental Design: Utilizing different multiple myeloma cell lines we determined the effect of TG02 over viability by MTT assays. The apoptotic effect over multiple myeloma patient samples was studied ex vivo by cytometry. The mechanism of action of TG02 was analyzed in the cell line MM1S, studying its effect on the cell cycle, the induction of apoptosis, and the loss of mitochondrial membrane potential by cytometry and Western blot. Two models of multiple myeloma xenograft were utilized to study the in vivo action of TG02.

Results: TG02 potently inhibited proliferation and survival of multiple myeloma cell lines, even under protective bone marrow niche conditions, and selectively induced apoptosis of primary patient-derived malignant plasma cells. TG02 displayed significant single-agent activity in two multiple myeloma xenograft models, and enhanced the in vivo activity of bortezomib and lenalidomide. Signaling analyses revealed that the drug simultaneously blocked the activity of CDKs 1, 2, and 9 as well as the MAP kinase ERK5 in MM1S cells, leading to cell-cycle arrest and rapid commitment to apoptosis. TG02 induced robust activation of both the intrinsic and extrinsic pathways of apoptosis, and depletion of XIAP and the key multiple myeloma survival protein Mcl-1.

Conclusions: TG02 is a promising new antimyeloma agent that is currently in phase I clinical trials in leukemia and multiple myeloma patients. Clin Cancer Res; 19(10); 2677–87. ©2013 AACR.
Data from A Long-term Prospective Population Pharmacokinetic Study on Imatinib Plasma Concentrations in GIST Patients
Authors: Eechoute Karel , Fransson Martin N. , Reyners An K. , de Jong Floris A. , Sparreboom Alex , van der Graaf Winette T.A. , Friberg Lena E. , Schiavon Gaia , Wiemer Erik A.C. , Verweij Jaap , Loos Walter J. , Mathijssen Ron H.J. , De Giorgi Ugo
Published: April 2023

Abstract
Purpose: Imatinib minimal (trough) plasma concentrations after one month of treatment have shown a significant association with clinical benefit in patients with gastrointestinal stromal tumors (GIST). Considering that a retrospective pharmacokinetic analysis has also suggested that imatinib clearance increases over time in patients with soft tissue sarcoma and GIST, the primary aim of this study was to assess systemic exposure to imatinib at multiple time points in a long-term prospective population pharmacokinetic study. As imatinib is mainly metabolized in the liver, our secondary aim was to elucidate the potential effects of the volume of liver metastases on exposure to imatinib.

Experimental Design: Full pharmacokinetic blood sampling was conducted in 50 patients with GIST on the first day of imatinib treatment, and after one, six, and 12 months. In addition, on day 14, and monthly during imatinib treatment, trough samples were taken. Pharmacokinetic analysis was conducted using a compartmental model. Volume of liver metastases was assessed by computed tomographic (CT) imaging.

Results: After 90 days of treatment, a significant decrease in imatinib systemic exposure of 29.3% compared with baseline was observed (P < 0.01). For every 100 cm3 increase of metastatic volume, a predicted decrease of 3.8% in imatinib clearance was observed.

Conclusions: This is the first prospective pharmacokinetic study in patients with GIST, showing a significant decrease of approximately 30% in imatinib exposure after long-term treatment. This means that future "trough level – clinical benefit" analyses should be time point specific. GIST liver involvement, however, has a marginal effect on imatinib clearance. Clin Cancer Res; 18(20); 5780–7. ©2012 AACR.
Data from How to Develop Treatments for Biologically Heterogeneous “Diseases”
Authors: Simon Richard M.
Published: April 2023

Abstract
The standard paradigm for the design of phase III clinical trials is not suitable for evaluation of molecularly targeted treatments in biologically heterogeneous groups of patients. Here, we comment on alternative clinical trial designs and propose a prospective discovery/evaluation framework for using tumor genomics in the design of phase III trials. Clin Cancer Res; 18(15); 4001–3. ©2012 AACR.
Data from The Rectal Cancer microRNAome – microRNA Expression in Rectal Cancer and Matched Normal Mucosa
Authors: Gaedcke Jochen , Grade Marian , Camps Jordi , Søkilde Rolf , Kaczkowski Bogumil , Schetter Aaron J. , Difilippantonio Michael J. , Harris Curtis C. , Ghadimi B. Michael , Møller Søren , Beissbarth Tim , Ried Thomas , Litman Thomas
Published: April 2023

Abstract
Purpose: miRNAs play a prominent role in a variety of physiologic and pathologic biologic processes, including cancer. For rectal cancers, only limited data are available on miRNA expression profiles, whereas the underlying genomic and transcriptomic aberrations have been firmly established. We therefore, aimed to comprehensively map the miRNA expression patterns of this disease.

Experimental Design: Tumor biopsies and corresponding matched mucosa samples were prospectively collected from 57 patients with locally advanced rectal cancers. Total RNA was extracted, and tumor and mucosa miRNA expression profiles were subsequently established for all patients. The expression of selected miRNAs was validated using semi-quantitative real-time PCR.

Results: Forty-nine miRNAs were significantly differentially expressed (log2-fold difference >0.5 and P < 0.001) between rectal cancer and normal rectal mucosa. The predicted targets for these miRNAs were enriched for the following pathways: Wnt, TGF-beta, mTOR, insulin, mitogen-activated protein kinase, and ErbB signaling. Thirteen of these 49 miRNAs seem to be rectal cancer-specific, and have not been previously reported for colon cancers: miR-492, miR-542-5p, miR-584, miR-483-5p, miR-144, miR-2110, miR-652, miR-375, miR-147b, miR-148a, miR-190, miR-26a/b, and miR-338-3p. Of clinical impact, miR-135b expression correlated significantly with disease-free and cancer-specific survival in an independent multicenter cohort of 116 patients.

Conclusion: This comprehensive analysis of the rectal cancer miRNAome uncovered novel miRNAs and pathways associated with rectal cancer. This information contributes to a detailed view of this disease. Moreover, the identification and validation of miR-135b may help to identify novel molecular targets and pathways for therapeutic exploitation. Clin Cancer Res; 18(18); 4919–30. ©2012 AACR.
Data from Synthetic miR-34a Mimics as a Novel Therapeutic Agent for Multiple Myeloma: In Vitro and In Vivo Evidence
Authors: Di Martino Maria T. , Leone Emanuela , Amodio Nicola , Foresta Umberto , Lionetti Marta , Pitari Maria R. , Cantafio Maria E. Gallo , Gullà Annamaria , Conforti Francesco , Morelli Eugenio , Tomaino Vera , Rossi Marco , Negrini Massimo , Ferrarini Manlio , Caraglia Michele , Shammas Masood A. , Munshi Nikhil C. , Anderson Kenneth C. , Neri Antonino , Tagliaferri Pierosandro , Tassone Pierfrancesco
Published: April 2023

Abstract
Purpose: Deregulated expression of miRNAs has been shown in multiple myeloma (MM). A promising strategy to achieve a therapeutic effect by targeting the miRNA regulatory network is to enforce the expression of miRNAs that act as tumor suppressor genes, such as miR-34a.

Experimental Design: Here, we investigated the therapeutic potential of synthetic miR-34a against human MM cells in vitro and in vivo.

Results: Either transient expression of miR-34a synthetic mimics or lentivirus-based miR-34a-stable enforced expression triggered growth inhibition and apoptosis in MM cells in vitro. Synthetic miR-34a downregulated canonic targets BCL2, CDK6, and NOTCH1 at both the mRNA and protein level. Lentiviral vector-transduced MM xenografts with constitutive miR-34a expression showed high growth inhibition in severe combined immunodeficient (SCID) mice. The anti-MM activity of lipidic-formulated miR-34a was further shown in vivo in two different experimental settings: (i) SCID mice bearing nontransduced MM xenografts; and (ii) SCID-synth-hu mice implanted with synthetic 3-dimensional scaffolds reconstituted with human bone marrow stromal cells and then engrafted with human MM cells. Relevant tumor growth inhibition and survival improvement were observed in mice bearing TP53-mutated MM xenografts treated with miR-34a mimics in the absence of systemic toxicity.

Conclusions: Our findings provide a proof-of-principle that formulated synthetic miR-34a has therapeutic activity in preclinical models and support a framework for development of miR-34a–based treatment strategies in MM patients. Clin Cancer Res; 18(22); 6260–70. ©2012 AACR.
Data from A Phase I Dose-Finding Trial of Recombinant Interleukin-21 and Rituximab in Relapsed and Refractory Low Grade B-cell Lymphoproliferative Disorders
Authors: Timmerman John M. , Byrd John C. , Andorsky David J. , Yamada Reiko E. , Kramer Janet , Muthusamy Natarajan , Hunder Naomi , Pagel John M.
Published: April 2023

Abstract
Purpose: We conducted a phase I study to determine the safety, maximum-tolerated dose (MTD), and efficacy of weekly bolus recombinant human interleukin-21 (rIL-21) plus rituximab in patients with indolent B-cell malignancies.

Experimental Design: One week after a lead-in rituximab dose, cohorts of three patients were treated with 30, 100, or 150 μg/kg rIL-21 weekly for four weeks, concurrent with four weekly doses of rituximab. Patients with stable disease or better were eligible for a second course of therapy.

Results: Twenty-one patients with relapsed small lymphocytic lymphoma/chronic lymphocytic leukemia (SLL/CLL, n = 11), follicular lymphoma (n = 9), or marginal zone lymphoma (n = 1) were enrolled, with 19 completing at least one course of therapy. The MTD for rIL-21 was 100 μg/kg, based on observed toxicities including nausea, vomiting, diarrhea, hypotension, edema, and hypophosphatemia. Clinical responses were seen in 8 of 19 evaluable patients (42%; 3 CR/CRu, 5 PR), with 4 of longer duration than the patient's previous response to rituximab-based treatment (median 9 months vs. 3 months).

Conclusions: Outpatient therapy of indolent B-cell malignancies with rituximab and weekly rIL-21 was well tolerated and clinically active, with durable complete remissions in a small subset of patients. Additional studies of rIL-21 and anti-CD20 antibodies in lymphoma and SLL/CLL are warranted. Clin Cancer Res; 18(20); 5752–60. ©2012 AACR.
Data from Molecular Pathways: Tumor Cells Co-opt the Brain-Specific Metabolism Gene CPT1C to Promote Survival
Authors: Reilly Patrick T. , Mak Tak W.
Published: April 2023

Abstract
The metabolic adaptations of cancer cells are receiving renewed attention as potential targets for therapeutic exploitation. Recent work has highlighted the importance of fatty acid catabolism through β-oxidation to cellular energy homeostasis. In this article, we describe recent preclinical studies suggesting that a gene usually expressed only in the brain, carnitine palmitoyltransferase (CPT)1C, promotes cancer cell survival and tumor growth. CTP1C confers rapamycin resistance on breast cancer cells, indicating that this gene may act in a pathway parallel to mTOR-enhanced glycolysis. Because of CPT1C's normally brain-restricted expression and the inability of most drugs to pass the blood–brain barrier, CPT1C may be an ideal candidate for specific small-molecule inhibition. We further speculate that concurrent targeting of CPT1C activity and glycolysis in tumor cells could be a highly effective anticancer approach. Clin Cancer Res; 18(21); 5850–5. ©2012 AACR.
Data from Double Down for a Double Win
Authors: Huang Pearl S.
Published: April 2023

Abstract
The rationale for using multiple inhibitors between and within the phosphoinositide 3-kinase/AKT/mTOR and RAS/MEK/ERK pathways is scientifically compelling, and a limited number of experimental agents are currently being tested in phase I combinations. Patient subpopulations, whose tumors are defined by genetic lesions, are showing promising responses to this approach. Clin Cancer Res; 18(8); 2124–6. ©2012 AACR.
Data from Analysis of Spontaneous Tumor-Specific CD4 T-cell Immunity in Lung Cancer Using Promiscuous HLA-DR Telomerase-Derived Epitopes: Potential Synergistic Effect with Chemotherapy Response
Authors: Godet Yann , Fabre Elizabeth , Dosset Magalie , Lamuraglia Michele , Levionnois Emeline , Ravel Patrice , Benhamouda Nadine , Cazes Aurélie , Le Pimpec-Barthes Françoise , Gaugler Beatrice , Langlade-Demoyen Pierre , Pivot Xavier , Saas Philippe , Maillère Bernard , Tartour Eric , Borg Christophe , Adotévi Olivier
Published: April 2023

Abstract
Purpose: To investigate the presence and impact of spontaneous telomerase-specific CD4 T-cell responses in cancer patients.

Experimental Design: A multistep approach was used to design novel pan-HLA-DR–restricted peptides from telomerase. T-cell clones isolated from cancer patients were used to characterize the polarization of telomerase-specific CD4 response. The presence of spontaneous CD4 T-cell response against telomerase was monitored in 84 metastatic non–small cell lung cancer (NSCLC) patients before first-line chemotherapy (CT) using IFN-γ ELISPOT assay. Then we analyzed the impact of the pretherapeutic telomerase-specific CD4 T immunity on clinical outcome in patients according to their respective response to CT.

Results: We described four novel telomerase-derived CD4 epitopes referred as universal cancer peptides (UCP) that effectively bind to most commonly found human MHC class II alleles. UCP-specific CD4 T-cell repertoire is present in human and UCP-specific CD4 T-cell clones generated from cancer patients exhibited high avidity and are Th1 polarized. Significant frequency (38%) of naturally occurring UCP-specific T-cell responses were detected before CT in advanced NSCLC but not in healthy volunteers. This response was shown to significantly increase overall survival (OS) of patients responding to CT (Median OS: 53 vs. 40 weeks, P = 0.034).

Conclusions: These results show for the first time a potential synergistic effect of telomerase-specific CD4 T-cell response with CT response in NSCLC and underline the potential role of tumor-specific CD4 T-cell response on the efficiency of conventional anticancer therapy. Clin Cancer Res; 18(10); 2943–53. ©2012 AACR.
Data from Genome-Wide miRNA Expression Profiling Identifies miR-9-3 and miR-193a as Targets for DNA Methylation in Non–Small Cell Lung Cancers
Authors: Heller Gerwin , Weinzierl Marlene , Noll Christian , Babinsky Valerie , Ziegler Barbara , Altenberger Corinna , Minichsdorfer Christoph , Lang György , Döme Balazs , End-Pfützenreuter Adelheid , Arns Britt-Madeleine , Grin Yuliya , Klepetko Walter , Zielinski Christoph C. , Zöchbauer-Müller Sabine
Published: April 2023

Abstract
Purpose: The major aim of this study was to investigate the role of DNA methylation (referred to as methylation) on miRNA silencing in non–small cell lung cancers (NSCLC).

Experimental Design: We conducted microarray expression analyses of 856 miRNAs in NSCLC A549 cells before and after treatment with the DNA methyltransferase inhibitor 5-aza-2′-deoxycytidine (Aza-dC) and with a combination of Aza-dC and the histone deacetylase inhibitor trichostatin A. miRNA methylation was determined in 11 NSCLC cell lines and in primary tumors and corresponding nonmalignant lung tissue samples of 101 patients with stage I–III NSCLC.

Results: By comparing microarray data of untreated and drug-treated A549 cells, we identified 33 miRNAs whose expression was upregulated after drug treatment and which are associated with a CpG island. Thirty (91%) of these miRNAs were found to be methylated in at least 1 of 11 NSCLC cell lines analyzed. Moreover, miR-9-3 and miR-193a were found to be tumor specifically methylated in patients with NSCLC. We observed a shorter disease-free survival of patients with miR-9-3 methylated lung squamous cell carcinoma (LSCC) than patients with miR-9-3 unmethylated LSCC by multivariate analysis [HR = 3.8; 95% confidence interval (CI), 1.3–11.2, P = 0.017] and a shorter overall survival of patients with miR-9-3 methylated LSCC than patients with miR-9-3 unmethylated LSCC by univariate analysis (P = 0.013).

Conclusions: Overall, our results suggest that methylation is an important mechanism for inactivation of certain miRNAs in NSCLCs and that miR-9-3 methylation may serve as a prognostic parameter in patients with LSCC. Clin Cancer Res; 18(6); 1619–29. ©2012 AACR.
Data from Combined Alloreactive CTL Cellular Therapy with Prodrug Activator Gene Therapy in a Model of Breast Cancer Metastatic to the Brain
Authors: Hickey Michelle J. , Malone Colin C. , Erickson Kate L. , Lin Amy , Soto Horacio , Ha Edward T. , Kamijima Shuichi , Inagaki Akihito , Takahashi Masamichi , Kato Yuki , Kasahara Noriyuki , Mueller Barbara M. , Kruse Carol A.
Published: April 2023

Abstract
Purpose: Individual or combined strategies of cellular therapy with alloreactive CTLs (alloCTL) and gene therapy using retroviral replicating vectors (RRV) encoding a suicide prodrug activating gene were explored for the treatment of breast tumors metastatic to the brain.

Experimental Design: AlloCTL, sensitized to the HLA of MDA-MB-231 breast cancer cells, were examined in vitro for antitumor functionality toward breast cancer targets. RRV encoding the yeast cytosine deaminase (CD) gene was tested in vivo for virus spread, ability to infect, and kill breast cancer targets when exposed to 5-fluorocytosine (5-FC). Individual and combination treatments were tested in subcutaneous and intracranial xenograft models with 231BR, a brain tropic variant.

Results: AlloCTL preparations were cytotoxic, proliferated, and produced IFN-γ when coincubated with target cells displaying relevant HLA. In vivo, intratumorally placed alloCTL trafficked through one established intracranial 231BR focus to another in contralateral brain and induced tumor cell apoptosis. RRV-CD efficiently spread in vivo, infected 231BR and induced their apoptosis upon 5-FC exposure. Subcutaneous tumor volumes were significantly reduced in alloCTL and/or gene therapy–treated groups compared to control groups. Mice with established intracranial 231BR tumors treated with combined alloCTL and RRV-CD had a median survival of 97.5 days compared with single modalities (50–83 days); all experimental treatment groups survived significantly longer than sham-treated groups (median survivals 31.5 or 40 days) and exhibited good safety/toxicity profiles.

Conclusion: The results indicate combining cellular and suicide gene therapies is a viable strategy for the treatment of established breast tumors in the brain. Clin Cancer Res; 19(15); 4137–48. ©2013 AACR.
Data from TP53 Disruptive Mutations Lead to Head and Neck Cancer Treatment Failure through Inhibition of Radiation-Induced Senescence
Authors: Skinner Heath D. , Sandulache Vlad C. , Ow Thomas J. , Meyn Raymond E. , Yordy John S. , Beadle Beth M. , Fitzgerald Alison L. , Giri Uma , Ang K. Kian , Myers Jeffrey N.
Published: April 2023

Abstract
Purpose: Mortality of patients with head and neck squamous cell carcinoma (HNSCC) is primarily driven by tumor cell radioresistance leading to locoregional recurrence (LRR). In this study, we use a classification of TP53 mutation (disruptive vs. nondisruptive) and examine impact on clinical outcomes and radiation sensitivity.

Experimental Design: Seventy-four patients with HNSCC treated with surgery and postoperative radiation and 38 HNSCC cell lines were assembled; for each, TP53 was sequenced and the in vitro radioresistance measured using clonogenic assays. p53 protein expression was inhibited using short hairpin RNA (shRNA) and overexpressed using a retrovirus. Radiation-induced apoptosis, mitotic cell death, senescence, and reactive oxygen species (ROS) assays were carried out. The effect of the drug metformin on overcoming mutant p53-associated radiation resistance was examined in vitro as well as in vivo, using an orthotopic xenograft model.

Results: Mutant TP53 alone was not predictive of LRR; however, disruptive TP53 mutation strongly predicted LRR (P = 0.03). Cell lines with disruptive mutations were significantly more radioresistant (P < 0.05). Expression of disruptive TP53 mutations significantly decreased radiation-induced senescence, as measured by SA-β-gal staining, p21 expression, and release of ROS. The mitochondrial agent metformin potentiated the effects of radiation in the presence of a disruptive TP53 mutation partially via senescence. Examination of our patient cohort showed that LRR was decreased in patients taking metformin.

Conclusions: Disruptive TP53 mutations in HNSCC tumors predicts for LRR, because of increased radioresistance via the inhibition of senescence. Metformin can serve as a radiosensitizer for HNSCC with disruptive TP53, presaging the possibility of personalizing HNSCC treatment. Clin Cancer Res; 18(1); 290–300. ©2011 AACR.
Data from The Antitumor Immunity of Ipilimumab: (T-cell) Memories to Last a Lifetime?
Authors: Postow Michael A. , Callahan Margaret K. , Wolchok Jedd D.
Published: April 2023

Abstract
Ipilimumab has shown an overall survival benefit in 2 randomized phase III studies. A minority of patients achieve long-term disease control, highlighting the potential of this immunotherapeutic approach. In ongoing efforts, investigators are continuing to characterize these patients' unique clinical courses and correlate their responses with underlying mechanisms of antitumor immunity. Clin Cancer Res; 18(7); 1821–3. ©2012 AACR.
Data from Inhibition of GSK3B Bypass Drug Resistance of p53-Null Colon Carcinomas by Enabling Necroptosis in Response to Chemotherapy
Authors: Grassilli Emanuela , Narloch Robert , Federzoni Elena , Ianzano Leonarda , Pisano Fabio , Giovannoni Roberto , Romano Gabriele , Masiero Laura , Leone Biagio Eugenio , Bonin Serena , Donada Marisa , Stanta Giorgio , Helin Kristian , Lavitrano Marialuisa
Published: April 2023

Abstract
Purpose: Evasion from chemotherapy-induced apoptosis due to p53 loss strongly contributes to drug resistance. Identification of specific targets for the treatment of drug-resistant p53-null tumors would therefore increase the effectiveness of cancer therapy.

Experimental Design: By using a kinase-directed short hairpin RNA library and HCT116p53KO drug-resistant colon carcinoma cells, glycogen synthase kinase 3 beta (GSK3B) was identified as a target whose silencing bypasses drug resistance due to loss of p53. p53-null colon cancer cell lines with different sets of mutations were used to validate the role of GSK3B in sustaining resistance and to characterize cell death mechanisms triggered by chemotherapy when GSK3B is silenced. In vivo xenograft studies were conducted to confirm resensitization of drug-resistant cells to chemotherapy upon GSK3 inhibition. Colon cancer samples from a cohort of 50 chemotherapy-treated stage II patients were analyzed for active GSK3B expression.

Results: Downregulation of GSK3B in various drug-resistant p53-null colon cancer cell lines abolished cell viability and colony growth after drug addition without affecting cell proliferation or cell cycle in untreated cells. Cell death of 5-fluorouracil (5FU)–treated p53-null GSK3B-silenced colon carcinoma cells occurred via PARP1-dependent and AIF-mediated but RIP1-independent necroptosis. In vivo studies showed that drug-resistant xenograft tumor mass was significantly reduced only when 5FU was given after GSK3B inhibition. Tissue microarray analysis of colon carcinoma samples from 5FU-treated patients revealed that GSK3B is significantly more activated in drug-resistant versus responsive patients.

Conclusions: Targeting GSK3B, in combination with chemotherapy, may represent a novel strategy for the treatment of chemotherapy-resistant tumors. Clin Cancer Res; 19(14); 3820–31. ©2013 AACR.
Data from Tumor-Specific T-cell Help Is Associated with Improved Survival in Melanoma
Authors: Woods Katherine , Cebon Jonathan
Published: April 2023

Abstract
Despite success with immune checkpoint inhibitors, clinical benefit from cancer vaccines remains elusive. Combined targeting of melanoma-specific CD4+ and CD8+ T-lymphocyte epitopes was associated with improved survival compared with targeting either alone, or when a nonspecific helper epitope was used. We discuss the potential role of antigen-specific CD4 help. Clin Cancer Res; 19(15); 4021–3. ©2013 AACR.
Data from Paracrine Receptor Activation by Microenvironment Triggers Bypass Survival Signals and ALK Inhibitor Resistance in EML4-ALK Lung Cancer Cells
Authors: Yamada Tadaaki , Takeuchi Shinji , Nakade Junya , Kita Kenji , Nakagawa Takayuki , Nanjo Shigeki , Nakamura Takahiro , Matsumoto Kunio , Soda Manabu , Mano Hiroyuki , Uenaka Toshimitsu , Yano Seiji
Published: April 2023

Abstract
Purpose: Cancer cell microenvironments, including host cells, can critically affect cancer cell behaviors, including drug sensitivity. Although crizotinib, a dual tyrosine kinase inhibitor (TKI) of ALK and Met, shows dramatic effect against EML4-ALK lung cancer cells, these cells can acquire resistance to crizotinib by several mechanisms, including ALK amplification and gatekeeper mutation. We determined whether microenvironmental factors trigger ALK inhibitor resistance in EML4-ALK lung cancer cells.

Experimental Design: We tested the effects of ligands produced by endothelial cells and fibroblasts, and the cells themselves, on the susceptibility of EML4-ALK lung cancer cell lines to crizotinib and TAE684, a selective ALK inhibitor active against cells with ALK amplification and gatekeeper mutations, both in vitro and in vivo.

Results:EML4-ALK lung cancer cells were highly sensitive to ALK inhibitors. EGF receptor (EGFR) ligands, such as EGF, TGF-α, and HB-EGF, activated EGFR and triggered resistance to crizotinib and TAE684 by transducing bypass survival signaling through Erk1/2 and Akt. Hepatocyte growth factor (HGF) activated Met/Gab1 and triggered resistance to TAE684, but not crizotinib, which inhibits Met. Endothelial cells and fibroblasts, which produce the EGFR ligands and HGF, respectively, decreased the sensitivity of EML4-ALK lung cancer cells to crizotinib and TAE684, respectively. EGFR-TKIs resensitized these cells to crizotinib and Met-TKI to TAE684 even in the presence of EGFR ligands and HGF, respectively.

Conclusions: Paracrine receptor activation by ligands from the microenvironment may trigger resistance to ALK inhibitors in EML4-ALK lung cancer cells, suggesting that receptor ligands from microenvironment may be additional targets during treatment with ALK inhibitors. Clin Cancer Res; 18(13); 3592–602. ©2012 AACR.