bluuebunny/crossref_metadata_2025 huggingface dataset:
Data from Inhibition of Histone Deacetylation Potentiates the Evolution of Acquired Temozolomide Resistance Linked to MGMT Upregulation in Glioblastoma Xenografts
Authors: Kitange Gaspar J. , Mladek Ann C. , Carlson Brett L. , Schroeder Mark A. , Pokorny Jenny L. , Cen Ling , Decker Paul A. , Wu Wenting , Lomberk Gwen A. , Gupta Shiv K. , Urrutia Raul A. , Sarkaria Jann N.
Published: April 2023

Abstract
Purpose: The therapeutic benefit of temozolomide in glioblastoma multiforme (GBM) is limited by resistance. The goal of this study was to elucidate mechanisms of temozolomide resistance in GBM.

Experimental Design: We developed an in vivo GBM model of temozolomide resistance and used paired parental and temozolomide-resistant tumors to define the mechanisms underlying the development of resistance and the influence of histone deacetylation (HDAC) inhibition.

Results: Analysis of paired parental and resistant lines showed upregulation of O6-methylguanine-DNA methyltransferase (MGMT) expression in 3 of the 5 resistant xenografts. While no significant change was detected in MGMT promoter methylation between parental and derivative-resistant samples, chromatin immunoprecipitation showed an association between MGMT upregulation and elevated acetylation of lysine 9 of histone H3 (H3K9-ac) and decreased dimethylation (H3K9-me2) in GBM12 and GBM14. In contrast, temozolomide resistance development in GBM22 was not linked to MGMT expression, and both parental and resistant lines had low H3K9-ac and high H3K9-me2 within the MGMT promoter. In the GBM12TMZ-resistant line, MGMT reexpression was accompanied by increased recruitment of SP1, C-JUN, NF-κB, and p300 within the MGMT promoter. Interestingly, combined treatment of GBM12 flank xenografts with temozolomide and the HDAC inhibitor suberoylanilide hydroxamic acid (SAHA) favored the evolution of temozolomide resistance by MGMT overexpression as compared with treatment with temozolomide alone.

Conclusion: This study shows, for the first time, a unique mechanism of temozolomide resistance development driven by chromatin-mediated MGMT upregulation and highlights the potential for epigenetically directed therapies to influence the mechanisms of resistance development in GBM. Clin Cancer Res; 18(15); 4070–9. ©2012 AACR.
Data from Quantitative Immunofluorescence Reveals the Signature of Active B-cell Receptor Signaling in Diffuse Large B-cell Lymphoma
Authors: Bogusz Agata M. , Baxter Richard H.G. , Currie Treeve , Sinha Papiya , Sohani Aliyah R. , Kutok Jeffery L. , Rodig Scott J.
Published: April 2023

Abstract
Purpose: B-cell receptor (BCR)–mediated signaling is important in the pathogenesis of a subset of diffuse large B-cell lymphomas (DLBCL) and the BCR-associated kinases SYK and BTK have recently emerged as potential therapeutic targets. We sought to identify a signature of activated BCR signaling in DLBCL to aid the identification of tumors that may be most likely to respond to BCR-pathway inhibition.

Experimental Design: We applied quantitative immunofluorescence (qIF) using antibodies to phosphorylated forms of proximal BCR signaling kinases LYN, SYK, and BTK and antibody to BCR-associated transcription factor FOXO1 on BCR-cross-linked formalin-fixed paraffin-embedded (FFPE) DLBCL cell lines as a model system and on two clinical cohorts of FFPE DLBCL specimens (n = 154).

Results: A robust signature of active BCR signaling was identified and validated in BCR-cross-linked DLBCL cell lines and in 71/154 (46%) of the primary DLBCL patient specimens. Further analysis of the primary biopsy samples revealed increased nuclear exclusion of FOXO1 among DLBCL with qIF evidence of active BCR signaling compared with those without (P = 0.004). Nuclear exclusion of FOXO1 was also detected in a subset of DLBCL without evidence of proximal BCR signaling suggesting that alternative mechanisms for PI3K/AKT activation may mediate FOXO1 subcellular localization in these cases.

Conclusion: This study establishes the feasibility of detecting BCR activation in primary FFPE biopsy specimens of DLBCL. It lays a foundation for future dissection of signal transduction networks in DLBCL and provides a potential platform for evaluating individual tumors in patients receiving novel therapies targeting the BCR pathway. Clin Cancer Res; 18(22); 6122–35. ©2012 AACR.
Data from The IL-18 Antagonist IL-18–Binding Protein Is Produced in the Human Ovarian Cancer Microenvironment
Authors: Carbotti Grazia , Barisione Gaia , Orengo Anna Maria , Brizzolara Antonella , Airoldi Irma , Bagnoli Marina , Pinciroli Patrizia , Mezzanzanica Delia , Centurioni Maria Grazia , Fabbi Marina , Ferrini Silvano
Published: April 2023

Abstract
Purpose: Interleukin (IL)-18 is an immune-enhancing cytokine, which induces IFN-γ production, T-helper 1 responses, and antitumor effects. In turn, IFN-γ stimulates IL-18–binding protein production, which blocks IL-18 activity. In view of the potential use of IL-18 in epithelial ovarian cancer (EOC) immunotherapy, here, we studied IL-18BP expression and its regulation by cytokines in EOC cells in vitro and in vivo.

Experimental Design: Expression and production of IL-18BP in EOC cell lines, primary ovarian carcinomas, and the corresponding normal tissues, patients' serum, and ascites were investigated by immunochemistry, ELISA, screening of gene expression profiles, and reverse-transcription PCR.

Results: Analysis of gene expression profiles revealed that IL18BP mRNA is increased in EOC tumors compared with normal ovary cells. Release of IL-18BP was detectable in EOC sera and to a greater extent in the ascites, indicating production at the tumor site. Indeed, immunochemical analyses on cells isolated from the ascites and on tumor sections indicated that IL-18BP is expressed in both tumor cells and tumor-associated leukocytes, which displayed a CD3−CD20−NKp46−CD13+CD14low phenotype. EOC cell lines do not constitutively express IL-18BP. However, its release is inducible both by IFN-γ stimulation in vitro and by xenotransplantation of EOC cells in immune-deficient mice, suggesting a role for the microenvironment. In vitro experiments and immunochemistry indicated that IL-27 is also involved in IL-18BP upregulation in EOC cell lines and primary cells through STAT1 activation. Together, these data indicate that IL-18BP, which is produced in EOC in response to microenvironmental factors, may inhibit endogenous or exogenous IL-18 activity. Clin Cancer Res; 19(17); 4611–20. ©2013 AACR.
Data from Aflibercept in Pediatric Solid Tumors: Moving Beyond the Trap
Authors: Chau Cindy H. , Figg William D.
Published: April 2023

Abstract
Angiogenesis plays a pivotal role in the growth and metastasis of adult and pediatric solid tumors. Clinical investigation of angiogenesis inhibitors is currently under way for childhood cancers. While the pediatric study of aflibercept provides a proof-of-principle, challenges remain in developing clinical endpoints and biomarkers of angiogenesis for pediatric trials. Clin Cancer Res; 18(18); 4868–71. ©2012 AACR.
Data from Dendritic Cell–Activating Vaccine Adjuvants Differ in the Ability to Elicit Antitumor Immunity Due to an Adjuvant-Specific Induction of Immunosuppressive Cells
Authors: Dang Yushe , Wagner Wolfgang M. , Gad Ekram , Rastetter Lauren , Berger Carmen M. , Holt Gregory E. , Disis Mary L.
Published: April 2023

Abstract
Purpose: We questioned whether the vaccine adjuvant combination of TLR-7 ligand agonist, imiquimod, with granulocyte macrophage colony-stimulating factor (GM-CSF) would result in enhanced dendritic cell recruitment and activation with increased antigen-specific immunity as compared with either adjuvant used alone.

Experimental Design: The adjuvant effects of GM-CSF and imiquimod were studied in ovalbumin (OVA) and MMTVneu transgenic mice using peptide-based vaccines. Type I immunity, serum cytokines, myeloid-derived suppressive cells (MDSC), and regulatory T cells (Treg) levels were examined.

Results: Both GM-CSF and imiquimod equally induced local accumulation and activation of dendritic cells. Both adjuvants effectively enhanced OVA-specific T-cell responses. We further evaluated the antitumor efficacy of adjuvant GM-CSF and imiquimod immunizing against murine insulin-like growth factor–binding protein-2 (IGFBP-2), a nonmutated oncoprotein overexpressed in the tumors of MMTVneu transgenic mice. Tumor growth was significantly inhibited in the mice receiving IGFBP-2 peptides with GM-CSF (P = 0.000), but not in imiquimod vaccine-treated groups (P = 0.141). Moreover, the addition of imiquimod to GM-CSF negated the antitumor activity of the vaccine when GM-CSF was used as the sole adjuvant. While GM-CSF stimulated significant levels of antigen-specific T-helper cell (TH)1, imiquimod induced elevated serum interleukin (IL)-10. Both MDSC and Tregs were increased in the imiquimod-treated but not GM-CSF–treated groups (P = 0.000 and 0.006, respectively). Depleting MDSC and Treg in animals immunized with imiquimod and IGFBP-2 peptides restored antitumor activity to the levels observed with vaccination using GM-CSF as the sole adjuvant.

Conclusion: Adjuvants may induce regulatory responses in the context of a self-antigen vaccine. Adjuvant triggered immunosuppression may limit vaccine efficacy and should be evaluated in preclinical models especially when contemplating combination approaches. Clin Cancer Res; 18(11); 3122–31. ©2012 AACR.
Data from Universal Cancer Peptide-Based Therapeutic Vaccine Breaks Tolerance against Telomerase and Eradicates Established Tumor
Authors: Dosset Magalie , Godet Yann , Vauchy Charline , Beziaud Laurent , Lone Yu Chun , Sedlik Christine , Liard Christelle , Levionnois Emeline , Clerc Bertrand , Sandoval Federico , Daguindau Etienne , Wain-Hobson Simon , Tartour Eric , Langlade-Demoyen Pierre , Borg Christophe , Adotévi Olivier
Published: April 2023

Abstract
Purpose: To evaluate CD4+ helper functions and antitumor effect of promiscuous universal cancer peptides (UCP) derived from telomerase reverse transcriptase (TERT).

Experimental Design: To evaluate the widespread immunogenicity of UCPs in humans, spontaneous T-cell responses against UCPs were measured in various types of cancers using T-cell proliferation and ELISPOT assays. The humanized HLA-DRB1*0101/HLA-A*0201 transgenic mice were used to study the CD4+ helper effects of UCPs on antitumor CTL responses. UCP-based antitumor therapeutic vaccine was evaluated using HLA-A*0201–positive B16 melanoma that express TERT.

Results: The presence of a high number of UCP-specific CD4+ T cells was found in the blood of patients with various types of cancer. These UCP-specific T cells mainly produce IFN-γ and TNF-α. In HLA transgenic mice, UCP vaccinations induced high avidity CD4+ TH1 cells and activated dendritic cells that produced interleukin-12. UCP-based vaccination breaks self-tolerance against TERT and enhances primary and memory CTL responses. Furthermore, the use of UCP strongly improves the efficacy of therapeutic vaccination against established B16-HLA-A*0201 melanoma and promotes tumor infiltration by TERT-specific CD8+ T cells.

Conclusions: Our results showed that UCP-based vaccinations strongly stimulate antitumor immune responses and could be used to design efficient immunotherapies in multiple types of cancers. Clin Cancer Res; 18(22); 6284–95. ©2012 AACR.
Data from Leukemia Cell-Rhabdovirus Vaccine: Personalized Immunotherapy for Acute Lymphoblastic Leukemia
Authors: Conrad David P. , Tsang Jovian , Maclean Meaghan , Diallo Jean-Simon , Le Boeuf Fabrice , Lemay Chantal G. , Falls Theresa J. , Parato Kelley A. , Bell John C. , Atkins Harold L.
Published: April 2023

Abstract
Purpose: Acute lymphoblastic leukemia (ALL) remains incurable in most adults. It has been difficult to provide effective immunotherapy to improve outcomes for the majority of patients. Rhabdoviruses induce strong antiviral immune responses. We hypothesized that mice administered ex vivo rhabdovirus-infected ALL cells [immunotherapy by leukemia-oncotropic virus (iLOV)] would develop robust antileukemic immune responses capable of controlling ALL.

Experimental Design: Viral protein production, replication, and cytopathy were measured in human and murine ALL cells exposed to attenuated rhabdovirus. Survival following injection of graded amounts of ALL cells was compared between cohorts of mice administered γ-irradiated rhabdovirus-infected ALL cells (iLOV) or multiple control vaccines to determine key immunotherapeutic components and characteristics. Host immune requirements were assessed in immunodeficient and bone marrow–transplanted mice or by adoptive splenocyte transfer from immunized donors. Antileukemic immune memory was ascertained by second leukemic challenge in long-term survivors.

Results: Human and murine ALL cells were infected and killed by rhabdovirus; this produced a potent antileukemia vaccine. iLOV protected mice from otherwise lethal ALL by developing durable leukemia-specific immune-mediated responses (P < 0.0001), which required an intact CTL compartment. Preexisting antiviral immunity augmented iLOV potency. Splenocytes from iLOV-vaccinated donors protected 60% of naïve recipients from ALL challenge (P = 0.0001). Injecting leukemia cells activated by, or concurrent with, multiple Toll-like receptor agonists could not reproduce the protective effect of iLOV. Similarly, injecting uninfected irradiated viable, apoptotic, or necrotic leukemia cells with/without concurrent rhabdovirus administration was ineffective.

Conclusion: Rhabdovirus-infected leukemia cells can be used to produce a vaccine that induces robust specific immunity against aggressive leukemia. Clin Cancer Res; 19(14); 3832–43. ©2013 AACR.
Data from Gliomas Promote Immunosuppression through Induction of B7-H1 Expression in Tumor-Associated Macrophages
Authors: Bloch Orin , Crane Courtney A. , Kaur Rajwant , Safaee Michael , Rutkowski Martin J. , Parsa Andrew T.
Published: April 2023

Abstract
Purpose: Gliomas are known to induce local and systemic immunosuppression, inhibiting T-cell–mediated cytotoxic responses to tumor growth. Tumor-associated macrophages are a significant component of the immune infiltrate in gliomas and may express immunosuppressive surface ligands, such as B7-H1.

Experimental Design: Tumor and peripheral blood samples from patients with glioblastoma (GBM) were analyzed by flow cytometry to evaluate the expression of B7-H1 in circulating and tumor-infiltrating macrophages. Human monocytes from healthy patients were stimulated with conditioned media from glioma cells to evaluate B7-H1 expression. Production of interleukin (IL)-10 by stimulated monocytes was measured by ELISA, and stimulation with IL-10 alone was evaluated for the ability to induce B7-H1 expression. The effect of inhibiting IL-10 and its receptor on glioma-induced B7-H1 expression in monocytes was evaluated.

Results: Circulating monocytes in patients with GBM had significantly increased expression of B7-H1 compared with healthy control patients. Tumor-associated macrophages from matched GBM tissue had even greater B7-H1 expression. Treatment of normal monocytes with glioma-conditioned media could significantly increase B7-H1 expression. Stimulation of monocytes with conditioned media resulted in substantial production of IL-10 and upregulation of the IL-10 receptor. Stimulation of monocytes with IL-10 alone could significantly increase B7-H1 expression, sufficient to induce T-cell apoptosis when cocultured with stimulated monocytes. Inhibition of IL-10 and the IL-10 receptor could knock down the effect of glioma media on B7-H1 by more than 50%.

Conclusions: Gliomas can upregulate B7-H1 expression in circulating monocytes and tumor-infiltrative macrophages through modulation of autocrine/paracrine IL-10 signaling, resulting in an immunosuppressive phenotype. Clin Cancer Res; 19(12); 3165–75. ©2013 AACR.
Data from miRNA-34b Inhibits Prostate Cancer through Demethylation, Active Chromatin Modifications, and AKT Pathways
Authors: Majid Shahana , Dar Altaf A. , Saini Sharanjot , Shahryari Varahram , Arora Sumit , Zaman Mohd Saif , Chang Inik , Yamamura Soichiro , Tanaka Yuichiro , Chiyomaru Takeshi , Deng Guoren , Dahiya Rajvir
Published: April 2023

Abstract
Purpose: miRNAs can act as oncomirs or tumor-suppressor miRs in cancer. This study was undertaken to investigate the status and role of miR-34b in prostate cancer.

Experimental Design: Profiling of miR-34b was carried out in human prostate cancer cell lines and clinical samples by quantitative real-time PCR and in situ hybridization. Statistical analyses were done to assess diagnostic/prognostic potential. Biological significance was elucidated by carrying out a series of experiments in vitro and in vivo.

Results: We report that miR-34b is silenced in human prostate cancer and the mechanism is through CpG hypermethylation. miR-34b directly targeted methyltransferases and deacetylases resulting in a positive feedback loop inducing partial demethylation and active chromatin modifications. miR-34b expression could predict overall and recurrence-free survival such that patients with high miR-34b levels had longer survival. Functionally, miR-34b inhibited cell proliferation, colony formation, migration/invasion, and triggered G0/G1 cell-cycle arrest and apoptosis by directly targeting the Akt and its downstream proliferative genes. miR-34b caused a decline in the mesenchymal markers vimentin, ZO1, N-cadherin, and Snail with an increase in E-cadherin expression, thus inhibiting epithelial-to-mesenchymal transition. Finally we showed the antitumor effect of miR-34b in vivo. MiR-34b caused a dramatic decrease in tumor growth in nude mice compared with cont-miR.

Conclusion: These findings offer new insight into the role of miR-34b in the inhibition of prostate cancer through demethylation, active chromatin modification, and Akt pathways and may provide a rationale for the development of new strategies targeting epigenetic regulation of miRNAs for the treatment of prostate cancer. Clin Cancer Res; 19(1); 73–84. ©2012 AACR.
Data from Outcomes of Phase II Clinical Trials with Single-Agent Therapies in Advanced/Metastatic Non–Small Cell Lung Cancer Published between 2000 and 2009
Authors: Janku Filip , Berry Donald A. , Gong Jing , Parsons Henrique A. , Stewart David J. , Kurzrock Razelle
Published: April 2023

Abstract
Purpose: We analyzed the outcomes of single-agent phase II clinical trials in non–small cell lung cancer (NSCLC) to determine trial parameters that predicted clinical activity.

Exoerimental Design: Data on response rate (RR), progression-free survival (PFS), and overall survival (OS) from all English language, single-agent phase II trials in advanced/metastatic NSCLC indexed by PubMed (January 2000 through December 2009) were abstracted.

Results: A total of 143 single-agent phase II trials (7,701 patients) were identified. The median RR was 10%, PFS 2.8 months, and OS 7.6 months. RR and PFS correlated with OS (r = 0.46, P < 0.001, r = 0.52, P < 0.001, respectively) and RR correlated with PFS (r = 0.61, P < 0.001). Treatment arms enriched for patients with molecular targets had a higher median RR (48.8% vs. 9.7%, P = 0.005), longer median PFS (6 vs. 2.8 months, P = 0.005), and OS (11.3 vs. 7.5 months, P = 0.05) as compared with those of unselected patients. In multivariate analysis, only studies enriched for patients with molecular targets or including drugs that eventually gained FDA/EMA approval were associated with a higher RR, and longer PFS/OS.

Conclusions: In phase II trials in NSCLC, RR and PFS correlated with OS. Studies enriched for patients with putative molecular drug targets were associated with higher therapeutic benefit as compared with those of unselected populations. Clin Cancer Res; 18(22); 6356–63. ©2012 AACR.
Data from Molecular Pathways: The Complexity of the Epigenome in Cancer and Recent Clinical Advances
Authors: Conte Mariarosaria , Altucci Lucia
Published: April 2023

Abstract
Human cancer is causally linked to genomic and epigenomic deregulations. Epigenetic abnormalities occurring within signaling pathways regulating proliferation, migration, growth, differentiation, transcription, and death signals may be critical in the progression of malignancies. Consequently, identification of epigenetic marks and their bioimplications in tumors represents a crucial step toward defining new therapeutic strategies both in cancer treatment and prevention. Alterations of writers, readers, and erasers in cancer may affect, for example, the methylation and acetylation state of huge areas of chromatin, suggesting that epi-based treatments may require “distinct” therapeutic strategies compared with “canonical” targeted treatments. Whereas anticancer treatments targeting histone deacetylase and DNA methylation have entered the clinic, additional chromatin modification enzymes have not yet been pharmacologically targeted for clinical use in patients. Thus, a greater insight into alterations occurring on chromatin modifiers and their impact in tumorigenesis represents a crucial advancement in exploiting epigenetic targeting in cancer prevention and treatment. Here, the interplay of the best known epi-mutations and how their targeting might be optimized are addressed. Clin Cancer Res; 18(20); 5526–34. ©2012 AACR.
Data from Modulation of the Tumor Microvasculature by Phosphoinositide-3 Kinase Inhibition Increases Doxorubicin Delivery In Vivo
Authors: Qayum Naseer , Im Jaehong , Stratford Michael R. , Bernhard Eric J. , McKenna W. Gillies , Muschel Ruth J.
Published: April 2023

Abstract
Purpose: Because effective drug delivery is often limited by inadequate vasculature within the tumor, the ability to modulate the tumor microenvironment is one strategy that may achieve better drug distribution. We have previously shown that treatment of mice bearing tumors with phosphoinositide-3 kinase (PI3K) inhibitors alters vascular structure in a manner analogous to vascular normalization and results in increased perfusion of the tumor. On the basis of that result, we asked whether inhibition of PI3K would improve chemotherapy delivery.

Experimental Design: Mice with xenografts using the cell line SQ20B bearing a hypoxia marker or MMTV-neu transgenic mice with spontaneous breast tumors were treated with the class I PI3K inhibitor GDC-0941. The tumor vasculature was evaluated by Doppler ultrasound, and histology. The delivery of doxorubicin was assessed using whole animal fluorescence, distribution on histologic sections, high-performance liquid chromatography on tumor lysates, and tumor growth delay.

Results: Treatment with GDC-0941 led to approximately three-fold increases in perfusion, substantially reduced hypoxia and vascular normalization by histology. Significantly increased amounts of doxorubicin were delivered to the tumors correlating with synergistic tumor growth delay. The GDC-0941 itself had no effect on tumor growth.

Conclusion: Inhibition of PI3K led to vascular normalization and improved delivery of a chemotherapeutic agent. This study highlights the importance of the microvascular effects of some novel oncogenic signaling inhibitors and the need to take those changes into account in the design of clinical trials many of which use combinations of chemotherapeutic agents. Clin Cancer Res; 18(1); 161–9. ©2011 AACR.
Data from Serum Cholesterol and mTOR Inhibitors: Surrogate Biomarker or Epiphenomenon?
Authors: Cho Daniel C. , Atkins Michael B.
Published: April 2023

Abstract
Lee and colleagues report that increase in serum cholesterol is associated with improved clinical outcomes in patients with renal cell carcinoma treated with temsirolimus. Although these findings must be validated prospectively, it should also be determined if this marker is a true mechanism-based toxicity or an epiphenomenon associated with therapy. Clin Cancer Res; 18(11); 2999–3001. ©2012 AACR.
Data from Stemness of B-cell Progenitors in Multiple Myeloma Bone Marrow
Authors: Boucher Kelly , Parquet Nancy , Widen Raymond , Shain Kenneth , Baz Rachid , Alsina Melissa , Koomen John , Anasetti Claudio , Dalton William , Perez Lia E.
Published: April 2023

Abstract
Purpose: In myeloma, B cells and plasma cells show a clonal relationship. Clonotypic B cells may represent a tumor-initiating compartment or cancer stem cell responsible for minimal residual disease in myeloma.

Experimental Design: We report a study of 58 patients with myeloma at time of diagnosis or relapse. B cells in bone marrow were evaluated by multicolor flow cytometry and sorting. Clonality was determined by light chain and/or immunoglobulin chain gene rearrangement PCR. We also determined aldehyde dehydrogenase activity and colony formation growth. Drug sensitivity was tested with conventional and novel agents.

Results: Marrow CD19+ cells express a light chain identical to plasma cells and are therefore termed light chain restricted (LCR). The LCR B-cell mass is small in both newly diagnosed and relapsed patients (≤1%). Few marrow LCR B cells (∼10%) are CD19+/CD34+, with the rest being more differentiated CD19+/CD34− B cells. Marrow LCR CD19+ B cells exhibit enhanced aldehyde dehydrogenase activity versus healthy controls. Both CD19+/CD34+ and CD19+/CD34− cells showed colony formation activity, with colony growth efficiency optimized when stroma-conditioned medium was used. B-cell progenitors showed resistance to melphalan, lenalidomide, and bortezomib. Panobinostat, a histone deacetylase inhibitor, induced apoptosis of LCR B cells and CD138+ cells. LCR B cells are CD117, survivin, and Notch positive.

Conclusions: We propose that antigen-independent B-cell differentiation stages are involved in disease origination and progression in myeloma. Furthermore, investigations of myeloma putative stem cell progenitors may lead to novel treatments to eradicate the potential reservoir of minimal residual disease. Clin Cancer Res; 18(22); 6155–68. ©2012 AACR.
Data from Molecular Profiling of Aromatase Inhibitor–Treated Postmenopausal Breast Tumors Identifies Immune-Related Correlates of Resistance
Authors: Dunbier Anita K. , Ghazoui Zara , Anderson Helen , Salter Janine , Nerurkar Ashutosh , Osin Peter , A'hern Roger , Miller William R. , Smith Ian E. , Dowsett Mitch
Published: April 2023

Abstract
Purpose: Estrogen withdrawal by treatment with aromatase inhibitors is the most effective form of endocrine therapy for postmenopausal estrogen receptor–positive (ER+) breast cancer. However, response to therapy varies markedly and understanding of the precise molecular effects of aromatase inhibitors and causes of resistance is limited. We aimed to identify in clinical breast cancer those genes and pathways most associated with resistance to aromatase inhibitors by examining the global transcriptional effects of AI treatment.

Experimental Design: Baseline and 2-week posttreatment biopsies were obtained from 112 postmenopausal women with ER+ breast cancer receiving neoadjuvant anastrozole. Gene expression data were obtained from 81 baseline and 2-week paired samples. Pathway analysis identified (i) the most prevalent changes in expression and (ii) the pretreatment genes/pathways most related to poor antiproliferative response.

Results: A total of 1,327 genes were differentially expressed after 2-week treatment (false discovery rate < 0.01). Proliferation-associated genes and classical estrogen-dependent genes were strongly downregulated whereas collagens and chemokines were upregulated. Pretreatment expression of an inflammatory signature correlated with antiproliferative response to anastrozole and this observation was validated in an independent study. Higher expression of immune-related genes such as SLAMF8 and TNF as well as lymphocytic infiltration were associated with poorer response (P < 0.001) and validated in an independent cohort.

Conclusions: The molecular response to aromatase inhibitor treatment varies greatly between patients consistent with the variable clinical benefit from aromatase inhibitor treatment. Higher baseline expression of an inflammatory signature is associated with poor antiproliferative response and should be assessed further as a novel biomarker and potential target for aromatase inhibitor-treated patients. Clin Cancer Res; 19(10); 2775–86. ©2013 AACR.
Data from Novel Targeting of Phospho-cMET Overcomes Drug Resistance and Induces Antitumor Activity in Multiple Myeloma
Authors: Moschetta Michele , Basile Antonio , Ferrucci Arianna , Frassanito Maria Antonia , Rao Luigia , Ria Roberto , Solimando Antonio Giovanni , Giuliani Nicola , Boccarelli Angelina , Fumarola Fabio , Coluccia Mauro , Rossini Bernardo , Ruggieri Simona , Nico Beatrice , Maiorano Eugenio , Ribatti Domenico , Roccaro Aldo M. , Vacca Angelo
Published: April 2023

Abstract
Purpose: The aim of the study was to verify the hypothesis that the cMet oncogene is implicated in chemio- and novel drug resistance in multiple myeloma.

Experimental Design: We have evaluated the expression levels of cMET/phospho-cMET (p-cMET) and the activity of the novel selective p-cMET inhibitor (SU11274) in multiple myeloma cells, either sensitive (RPMI-8226 and MM.1S) or resistant (R5 and MM.1R) to anti–multiple myeloma drugs, in primary plasma cells and in multiple myeloma xenograft models.

Results: We found that resistant R5 and MM.1R cells presented with higher cMET phosphorylation, thus leading to constitutive activation of cMET-dependent signaling pathways. R5 cells exhibited a higher susceptibility to the SU11274 inhibitory effects on viability, proliferation, chemotaxis, adhesion, and to its apoptogenic effects. SU11274 was able to revert drug resistance in R5 cells. R5 but not RPMI-8226 cells displayed cMET-dependent activation of mitogen-activated protein kinase pathway. The cMET and p-cMET expression was higher on plasma cells from patients with multiple myeloma at relapse or on drug resistance than on those from patients at diagnosis, complete/partial remission, or from patients with monoclonal gammopathy of unknown significance. Viability, chemotaxis, adhesion to fibronectin or paired bone marrow stromal cells of plasma cells from relapsed or resistant patients was markedly inhibited by SU11274. Importantly, SU11274 showed higher therapeutic activity in R5- than in RPMI-8226–induced plasmocytomas. In R5 tumors, it caused apoptosis and necrosis and reverted bortezomib resistance.

Conclusion: Our findings suggest that the cMET pathway is constitutively activated in relapsed and resistant multiple myeloma where it may also be responsible for induction of drug resistance, thus providing the preclinical rationale for targeting cMET in patients with relapsed/refractory multiple myeloma. Clin Cancer Res; 19(16); 4371–82. ©2013 AACR.
Data from Individual Patient-Specific Immunity against High-Grade Glioma after Vaccination with Autologous Tumor Derived Peptides Bound to the 96 KD Chaperone Protein
Authors: Crane Courtney A. , Han Seunggu J. , Ahn Brian , Oehlke Jessica , Kivett Valerie , Fedoroff Anne , Butowski Nicholas , Chang Susan M. , Clarke Jennifer , Berger Mitchel S. , McDermott Michael W. , Prados Michael D. , Parsa Andrew T.
Published: April 2023

Abstract
Purpose: Cancer immunotherapy offers hope of a highly specific nontoxic adjuvant treatment. Heat shock protein peptide complexes (HSPPCs) found in cancer cells carry tumor-specific antigenic proteins and can facilitate adaptive and innate immune responses. Here we show that peptides bound to a 96 kD chaperone protein (HSP-96) from brain tissue containing glioblastoma multiforme (GBM) can be used to safely immunize patients with recurrent GBM.

Experimental Design: Multimodality immunomonitoring was completed on 12 patients with recurrent GBM before and after immunization with an autologous HSPPC vaccine derived from surgically resected tumor. Clinical endpoints included safety assessments and overall survival.

Results: No adverse events attributable to the vaccine were found. Testing of peripheral blood leukocytes before and after vaccination revealed a significant peripheral immune response specific for the peptides bound to HSP-96, in 11 of the 12 patients treated. Brain biopsies of immune responders after vaccination revealed focal CD4, CD8, and CD56 IFNγ positive cell infiltrates, consistent with tumor site specific immune responses. Immune responders had a median survival of 47 weeks after surgery and vaccination, compared with 16 weeks for the single nonresponder.

Conclusions: These data provide the first evidence in humans of individual patient-specific immune responses against autologous tumor derived peptides bound to HSP-96. Clin Cancer Res; 19(1); 205–14. ©2012 AACR.
Data from In Vivo Phosphoantigen Levels in Bisphosphonate-Treated Human Breast Tumors Trigger Vγ9Vδ2 T-cell Antitumor Cytotoxicity through ICAM-1 Engagement
Authors: Benzaïd Ismahène , Mönkkönen Hannu , Bonnelye Edith , Mönkkönen Jukka , Clézardin Philippe
Published: April 2023

Abstract
Purpose: Nitrogen-containing bisphosphonates (N-BP) such as zoledronate and risedronate exhibit antitumor effects. They block the activity of farnesyl pyrophosphate synthase (FPPS) in the mevalonate pathway, leading to intracellular accumulation of mevalonate metabolites (IPP/ApppI), which are recognized as tumor phosphoantigens by Vγ9Vδ2 T cells. However, mechanisms responsible for Vγ9Vδ2 T-cell recognition of N-BP–treated tumors producing IPP/ApppI remain unclear.

Experimental Design: The effects of N-BPs on Vγ9Vδ2 T-cell expansion and anticancer activity were evaluated in vitro and in animal models of human breast cancers. The modalities of recognition of breast tumors by Vγ9Vδ2 T cells in N-BP–treated animals were also examined.

Results: We found a strong correlation between Vγ9Vδ2 T-cell anticancer activity and intracellular accumulation of IPP/ApppI in risedronate-treated breast cancer cells in vitro. In addition, following risedronate treatment of immunodeficient mice bearing human breast tumors, human Vγ9Vδ2 T cells infiltrated and inhibited growth of tumors that produced high IPP/ApppI levels but not those expressing low IPP/ApppI levels. The combination of doxorubicin with a N-BP improved, however, Vγ9Vδ2 T-cell cytotoxicity against breast tumors expressing low IPP/ApppI levels. Moreover, Vγ9Vδ2 T-cell cytotoxicity in mice treated with risedronate or zoledronate did not only depend on IPP/ApppI accumulation in tumors but also on expression of tumor cell surface receptor intercellular adhesion molecule-1 (ICAM-1), which triggered the recognition of N-BP–treated breast cancer cells by Vγ9Vδ2 T cells in vivo.

Conclusion: These findings suggest that N-BPs can have an adjuvant role in cancer therapy by activating Vγ9Vδ2 T-cell cytotoxicity in patients with breast cancer that produces high IPP/ApppI levels after N-BP treatment. Clin Cancer Res; 18(22); 6249–59. ©2012 AACR.
Data from The 2-Nitroimidazole EF5 Is a Biomarker for Oxidoreductases That Activate the Bioreductive Prodrug CEN-209 under Hypoxia
Authors: Wang Jingli , Foehrenbacher Annika , Su Jiechuang , Patel Rita , Hay Michael P. , Hicks Kevin O. , Wilson William R.
Published: April 2023

Abstract
Purpose: Benzotriazine-N-oxide bioreductive prodrugs such as tirapazamine and its improved analogue CEN-209 (SN30000) have potential for exploiting hypoxia in tumors. Here, we test the hypothesis that the 2-nitroimidazole EF5, in clinical development for both immunohistochemical and positron emission tomography imaging of hypoxia, can detect not only hypoxia but also the one-electron reductases required for activation of these hypoxia-targeted prodrugs.

Experimental Design: Aerobic and hypoxic covalent binding of [14C]-EF5 was determined in human tumor cell lines, including lines with overexpression of NADPH:cytochrome P450 oxidoreductase (CYPOR), and reductive metabolism of tirapazamine and CEN-209 by mass spectrometry. DNA damage response was measured by γH2AX formation. Bioreductive metabolism was modulated in HCT116 tumor xenografts by overexpression of CYPOR and breathing of hyperbaric oxygen or 10% oxygen.

Results: Overexpression of CYPOR induced similar 2- to 4-fold increases in EF5 binding and metabolic reduction of tirapazamine and CEN-209 in SiHa and HCT116 cell lines, and similar enhancement of γH2AX formation. EF5 binding and metabolic reduction of the prodrugs were highly correlated in a panel of 14 hypoxic tumor cell lines. In HCT116 xenografts, CYPOR overexpression also significantly increased EF5 binding and CEN-209 reduction, and modification of tumor hypoxia caused similar changes to the bioreductive activation of both agents, resulting in a strong correlation between EF5 binding and CEN209-induced DNA damage (R2 = 0.68, P < 0.0001) at the individual tumor level.

Conclusions: EF5 binding is a promising stratification biomarker for benzotriazine-N-oxide bioreductive prodrugs because of its potential for interrogating reductase activity as well as hypoxia in individual tumors. Clin Cancer Res; 18(6); 1684–95. ©2011 AACR.
Data from Influence of Biospecimen Variables on Proteomic Biomarkers in Breast Cancer
Authors: Meric-Bernstam Funda , Akcakanat Argun , Chen Huiqin , Sahin Aysegul , Tarco Emily , Carkaci Selin , Adrada Beatriz E. , Singh Gopal , Do Kim-Anh , Garces Zerzhinski M. , Mittendorf Elizabeth , Babiera Gildy , Bedrosian Isabelle , Hwang Rosa , Krishnamurthy Savitri , Symmans William F. , Gonzalez-Angulo Ana Maria , Mills Gordon B.
Published: April 2023

Abstract
Background: PI3K/Akt/mTOR signaling is being actively pursued as a therapeutic target for breast cancer. We sought to determine if tumor heterogeneity and biospecimen variables affect the evaluation of PI3K/Akt/mTOR pathway markers.

Methods: Intraoperative image-guided core-needle biopsies (CNB), and central and peripheral surgical tumor specimens were prospectively collected in 53 patients with invasive breast cancer. Specimens were assessed with reverse-phase protein arrays (RPPA) and immunohistochemistry (IHC).

Results: There was a moderate or strong correlation between the expression of 149 (97%) of the 154 different RPPA markers in the center and periphery. Correlation was higher for smaller tumors, in patients who did not undergo neoadjuvant therapy, and with shorter cold ischemia time. Of 154 markers, 132 (86%) were not statistically different between the center and periphery, and 97 (63%) were not different between the CNB and the surgical specimen (average of the central and peripheral specimen). pAkt S473 and PTEN had a significant correlation between central and peripheral specimens, and between CNB and surgical specimen. However, pAkt S473, pS6 S235/236, and pS6 240/244 levels were significantly higher in CNB than the central specimens both by RPPA and by IHC.

Conclusions: Most individual proteomic biomarkers studied do not have significant intratumoral heterogeneity. However, protein and phosphoprotein levels are affected by biospecimen type and other preanalytic variables. PI3K pathway activation is greater in CNB compared with postexcision surgical samples suggesting a potential loss of phosphorylation during surgical manipulation, or with cold ischemia of surgical specimens. Clin Cancer Res; 20(14); 3870–83. ©2014 AACR.
Data from Search for a Gene Expression Signature of Breast Cancer Local Recurrence in Young Women
Authors: Servant Nicolas , Bollet Marc A. , Halfwerk Hans , Bleakley Kevin , Kreike Bas , Jacob Laurent , Sie Daoud , Kerkhoven Ron M. , Hupé Philippe , Hadhri Rim , Fourquet Alain , Bartelink Harry , Barillot Emmanuel , Sigal-Zafrani Brigitte , van de Vijver Marc J.
Published: April 2023

Abstract
Purpose: A gene expression signature, predictive for local recurrence after breast-conserving treatment, has previously been identified from a series of 165 young patients with breast cancer. We evaluated this signature on both another platform and an independent series, compared its performance with other published gene-sets, and investigated the gene expression profile of a larger data set.

Experimental Design: Gene expression tumor profiles were obtained on 148 of the initial 165 Dutch patients and on an independent validation series of 195 French patients. Both unsupervised and supervised classifications were used to study the gene expression profile of the 343 breast cancers and to identify subgroups that differ for their risk of local recurrence.

Results: The previous local recurrence signature was validated across platforms. However, when applied to the French patients, the signature did not reproduce its reported performance and did not better classify the patients than other published gene sets. Hierarchical clustering of all 343 breast cancers did not show any grouping reflecting local recurrence status. Genes related to proliferation were found differentially expressed between patients with or without local recurrence only in triple-negative tumors. Supervised classification revealed no significant gene set predictive for local recurrence or able to outperform classification based on clinical variables.

Conclusions: Although the previously identified local recurrence signature was robust on another platform, we were neither able to validate it on an independent data set, nor able to define a strong gene expression classifier for local recurrence using a larger data set. We conclude that there are no significant differences in gene expression pattern in tumors from patients with and without local recurrence after breast-conserving treatment. Clin Cancer Res; 18(6); 1704–15. ©2012 AACR.
Data from EGFR Protein Expression in Non–Small Cell Lung Cancer Predicts Response to an EGFR Tyrosine Kinase Inhibitor—A Novel Antibody for Immunohistochemistry or AQUA Technology
Authors: Mascaux Celine , Wynes Murry W. , Kato Yasufumi , Tran Cindy , Asuncion Bernadette Reyna , Zhao Jason M. , Gustavson Mark , Ranger-Moore Jim , Gaire Fabien , Matsubayashi Jun , Nagao Toshitaka , Yoshida Koichi , Ohira Tatuso , Ikeda Norihiko , Hirsch Fred R.
Published: April 2023

Abstract
Introduction: Epidermal growth factor receptor (EGFR) protein expression in non–small cell lung cancer (NSCLC) is not recommended for predicting response to EGFR tyrosine kinase inhibitors (TKI) due to conflicting results, all using antibodies detecting EGFR external domain (ED). We tested the predictive value of EGFR protein expression for response to an EGFR TKI with an antibody that detects the intracellular domain (ID) and compared fluorescence-based Automated QUantitative Analysis (AQUA) technology to immunohistochemistry (IHC).

Methods: Specimens from 98 gefitinib-treated NSCLC Japanese patients were evaluated by IHC (n = 98 of 98) and AQUA technology (n = 70 of 98). EGFR ID (5B7)- and ED-specific antibodies (3C6 and 31G7) were compared.

Results: EGFR expression evaluated with 5B7 was significantly higher in responders versus nonresponders to gefitinib both with IHC and with AQUA. ED-specific antibodies did not significantly predict response. Using AQUA and ID-specific antibody resulted in the best prediction performance with a positive and negative predictive value (PPV/NPV) for responders of 50% and 87%, respectively. EGFR expression with ID-specific antibody and AQUA also predicted responders in EGFR-mutated patients. Increased EGFR expression with the ID antibody is associated with increased median progression free survival (PFS; 11.7 months vs. 5.0, log rank, P = 0.034) and overall survival (OS; 38.6 vs. 14.9, P = 0.040) from gefitinib therapy.

Conclusions: EGFR protein expression using an ID-specific antibody specifically predicts response to gefitinib in NSCLC patients, including in EGFR-mutated patients, and increased PFS/OS from gefitinib. These data suggest that the choice of diagnostic antibody and methodology matters to predict response and outcome to specific therapies. The potential clinical application needs further validation. Clin Cancer Res; 17(24); 7796–807. ©2011 AACR.
Data from IGKV3 Proteins as Candidate “Off-the-Shelf” Vaccines for Kappa-Light Chain–Restricted B-Cell Non-Hodgkin Lymphomas
Authors: Martorelli Debora , Guidoboni Massimo , De Re Valli , Muraro Elena , Turrini Riccardo , Merlo Anna , Pasini Elisa , Caggiari Laura , Romagnoli Luca , Spina Michele , Mortarini Roberta , Gasparotto Daniela , Mazzucato Mario , Carbone Antonino , Rosato Antonio , Anichini Andrea , Dolcetti Riccardo
Published: April 2023

Abstract
Purpose: An increasing set of B-cell non-Hodgkin lymphomas (B-NHL) show a biased usage of IGKV3-20 and IGKV3-15 immunoglobulin genes, a feature that could be exploited for the development of ready-to-use, broadly applicable cancer vaccines.

Experimental Design: The immunogenic properties of clonal IGKV3-20 and IGKV3-15 proteins were analyzed with particular focus on their ability to elicit cross-reactive responses against molecularly related IGKV proteins expressed by different B-cell lymphoproliferative disorders.

Results: IGK+ lymphoma patients show humoral and T-cell responses to IGKV3-20 and IGKV3-15 proteins and IGKV3-specific cytotoxic T lymphocytes (CTL) can be easily induced ex vivo. IGKV3-20–specific CTLs cross-react against different IGKV3 proteins, an effect mediated by the presence of 21 shared, sometimes promiscuous, T-cell epitopes, presented by common HLA class I allele products, thus assuring a broad HLA coverage of IGKV3-based vaccines. Many natural epitope variants are carried by IGK light chains expressed by a broad spectrum of B-NHLs and we show that IGKV3-20–specific CTLs cross-react also against several of these variant epitopes. Both humoral and CTL-specific responses were induced by KLH-conjugated IGKV3-20 protein in HLA-A2-transgenic mice and coinjection of IGKV3-20–specific CTLs with IGKV3-20+ or IGKV3-15+ lymphoma cells into SCID mice totally prevented tumor growth, thus confirming the ability of these effectors to mediate efficient and cross-reactive cytotoxic responses also in vivo.

Conclusions: These results provide the rationale to exploit IGKV3 proteins as “off-the-shelf” vaccines for a large fraction of lymphoma patients. Clin Cancer Res; 18(15); 4080–91. ©2012 AACR.
Data from PIK3CA/PTEN Mutations and Akt Activation As Markers of Sensitivity to Allosteric mTOR Inhibitors
Authors: Meric-Bernstam Funda , Akcakanat Argun , Chen Huiqin , Do Kim-Anh , Sangai Takafumi , Adkins Farrell , Gonzalez-Angulo Ana Maria , Rashid Asif , Crosby Katherine , Dong Mei , Phan Alexandria T. , Wolff Robert A. , Gupta Sanjay , Mills Gordon B. , Yao James
Published: April 2023

Abstract
Purpose: We sought to determine whether phosphoinositide 3-kinase (PI3K) pathway mutation or activation state and rapamycin-induced feedback loop activation of Akt is associated with rapamycin sensitivity or resistance.

Experimental Design: Cancer cell lines were tested for rapamycin sensitivity, Akt phosphorylation, and mTOR target inhibition. Mice injected with breast or neuroendocrine cancer cells and patients with neuroendocrine tumor (NET) were treated with rapalogs and Akt phosphorylation was assessed.

Results: Thirty-one cell lines were rapamycin sensitive (RS) and 12 were relatively rapamycin resistant (RR; IC50 > 100 nmol/L). Cells with PIK3CA and/or PTEN mutations were more likely to be RS (P = 0.0123). Akt phosphorylation (S473 and T308) was significantly higher in RS cells (P < 0.0001). Rapamycin led to a significantly greater pathway inhibition and greater increase in p-Akt T308 (P < 0.0001) and p-Akt S473 (P = 0.0009) in RS cells. Rapamycin and everolimus significantly increased Akt phosphorylation but inhibited growth in an in vivo NET model (BON). In patients with NETs treated with everolimus and octreotide, progression-free survival correlated with p-Akt T308 in pretreatment (R = 0.4762, P = 0.0533) and on-treatment tumor biopsies (R = 0.6041, P = 0.0102). Patients who had a documented partial response were more likely to have an increase in p-Akt T308 with treatment compared with nonresponders (P = 0.0146).

Conclusion: PIK3CA/PTEN genomic aberrations and high p-Akt levels are associated with rapamycin sensitivity in vitro. Rapamycin-mediated Akt activation is greater in RS cells, with a similar observation in patients with clinical responses on exploratory biomarker analysis; thus feedback loop activation of Akt is not a marker of resistance but rather may function as an indicator of rapamycin activity. Clin Cancer Res; 18(6); 1777–89. ©2012 AACR.