bluuebunny/crossref_metadata_2025 huggingface dataset:
Data from Meta-analysis of Neuroblastomas Reveals a Skewed ALK Mutation Spectrum in Tumors with MYCN Amplification
Authors: De Brouwer Sara , De Preter Katleen , Kumps Candy , Zabrocki Piotr , Porcu Michaël , Westerhout Ellen M. , Lakeman Arjan , Vandesompele Jo , Hoebeeck Jasmien , Van Maerken Tom , De Paepe Anne , Laureys Geneviève , Schulte Johannes H. , Schramm Alexander , Van Den Broecke Caroline , Vermeulen Joëlle , Van Roy Nadine , Beiske Klaus , Renard Marleen , Noguera Rosa , Delattre Olivier , Janoueix-Lerosey Isabelle , Kogner Per , Martinsson Tommy , Nakagawara Akira , Ohira Miki , Caron Huib , Eggert Angelika , Cools Jan , Versteeg Rogier , Speleman Frank
Published: April 2023

Abstract
Purpose: Activating mutations of the anaplastic lymphoma kinase (ALK) were recently described in neuroblastoma. We carried out a meta-analysis of 709 neuroblastoma tumors to determine their frequency and mutation spectrum in relation to genomic and clinical parameters, and studied the prognostic significance of ALK copy number and expression.

Experimental Design: The frequency and type of ALK mutations, copy number gain, and expression were analyzed in a new series of 254 neuroblastoma tumors. Data from 455 published cases were used for further in-depth analysis.

Results: ALK mutations were present in 6.9% of 709 investigated tumors, and mutations were found in similar frequencies in favorable [International Neuroblastoma Staging System (INSS) 1, 2, and 4S; 5.7%] and unfavorable (INSS 3 and 4; 7.5%) neuroblastomas (P = 0.087). Two hotspot mutations, at positions R1275 and F1174, were observed (49% and 34.7% of the mutated cases, respectively). Interestingly, the F1174 mutations occurred in a high proportion of MYCN-amplified cases (P = 0.001), and this combined occurrence was associated with a particular poor outcome, suggesting a positive cooperative effect between both aberrations. Furthermore, the F1174L mutant was characterized by a higher degree of autophosphorylation and a more potent transforming capacity as compared with the R1275Q mutant. Chromosome 2p gains, including the ALK locus (91.8%), were associated with a significantly increased ALK expression, which was also correlated with poor survival.

Conclusions: ALK mutations occur in equal frequencies across all genomic subtypes, but F1174L mutants are observed in a higher frequency of MYCN-amplified tumors and show increased transforming capacity as compared with the R1275Q mutants. Clin Cancer Res; 16(17); 4353–62. ©2010 AACR.
Data from Determination of Microvessel Density by Quantitative Real-time PCR in Esophageal Cancer: Correlation with Histologic Methods, Angiogenic Growth Factor Expression, and Lymph Node Metastasis
Authors: Loges Sonja , Clausen Henning , Reichelt Uta , Bubenheim Michael , Erbersdobler Andreas , Schurr Paulus , Yekebas Emre , Schuch Gunter , Izbicki Jakob , Pantel Klaus , Bokemeyer Carsten , Fiedler Walter
Published: April 2023

Abstract
Purpose: Angiogenesis and lymphangiogenesis are important steps in tumor growth and dissemination and are of prognostic importance in solid tumors. The determination of microvessel density (MVD) by immunohistology is subject to considerable variability between different laboratories and observers. We compared MVD determination by immunohistology and quantitative real-time PCR and correlated the results with clinical variables.

Experimental Design: The expression of endothelial antigens vascular endothelial cadherin (CD144), P1H12 (CD146), tie-2, and VEGFR-2, and lymphatic endothelial markers VEGFR-3, Prox, and LYVE was assessed by quantitative PCR (qPCR) in primary surgical samples. The expression of angiogenetic growth factors VEGF-A, VEGF-C, VEGF-D, angiopoietin-1, and angiopoietin-2 was quantified by PCR and correlated with MVD and clinical variables.

Results: The expression of endothelial antigens vascular endothelial cadherin (CD144), P1H12 (CD146), tie-2, and VEGFR-2 correlated with each other in 54 samples of primary esophageal cancer (P < 0.0001 for all comparisons). MVD determined immunohistologically by CD31 staining in a subgroup of 35 patients correlated significantly with the qPCR method. The expression of angiogenetic growth factors VEGF-A, VEGF-C, VEGF-D, angiopoietin-1, and angiopoietin-2 was significantly associated with MVD (P < 0.0001 for all comparisons). Analysis of the expression of lymphendothelial markers VEGFR-3, Prox, and LYVE revealed concordant results, indicating that quantification of lymphendothelial cells is possible by qPCR. The presence of lymph node metastasis on surgical specimens was significantly correlated with MVD (P < 0.003), VEGFR-2 (P < 0.048), and VEGF-C (P < 0.042) expression.

Conclusions: These results indicate that quantification of MVD by qPCR in surgical samples of esophageal carcinoma yields similar results with immunohistology. Interestingly, the extent of angiogenesis and lymphangiogenesis was not related in individual tumor samples. Lymph node metastases could be predicted by MVD and VEGF-C expression.
Data from Antitumor Efficacy of Edotecarin as a Single Agent and in Combination with Chemotherapy Agents in a Xenograft Model
Authors: Ciomei Marina , Croci Valter , Ciavolella Antonella , Ballinari Dario , Pesenti Enrico
Published: April 2023

Abstract
The novel indolocarbazole edotecarin (J-107088, formerly ED-749) differs from other topoisomerase I inhibitors both pharmacokinetically and pharmacodynamically. In vitro, it is more potent than camptothecins and has a variable cytotoxic activity in 31 different human cancer cell lines. Edotecarin also possesses greater than additive inhibitory effects on cell proliferation when used in combination with other agents tested in vitro against various cancer cell lines. The present in vivo studies were done to extend the in vitro findings to characterize the antitumor effects of edotecarin when used either alone or in combination with other agents (i.e., 5-fluorouracil, irinotecan, cisplatin, oxaliplatin, and SU11248) in the HCT-116 human colon cancer xenograft model. Treatment effects were based on the delay in onset of an exponential growth of tumors in drug-treated versus vehicle control-treated groups. In all studies, edotecarin was active both as a single agent and in combination with other agents. Combination therapy resulted in greater than additive effects, the extent of which depended on the specific dosage regimen. Toxicity in these experiments was minimal. Of all 359 treated mice, the six that died of toxicity were in the high-dose edotecarin/oxaliplatin group. The results suggest that edotecarin may serve as effective chemotherapy of colon cancer when used as a single agent, in combination with standard regimens and other topoisomerase inhibitors or with novel agents, such as the multitargeted tyrosine kinase inhibitor SU11248.
Data from Higher Expression of the Androgen-Regulated Gene PSA/HK3 mRNA in Prostate Cancer Tissues Predicts Biochemical Recurrence-Free Survival
Authors: Sterbis Joseph R. , Gao Chunling , Furusato Bungo , Chen Yongmei , Shaheduzzaman Syed , Ravindranath Lakshmi , Osborn David J. , Rosner Inger L. , Dobi Albert , McLeod David G. , Sesterhenn Isabell A. , Srivastava Shiv , Cullen Jennifer , Petrovics Gyorgy
Published: April 2023

Abstract
Purpose: Alterations of the androgen receptor (AR)-mediated signaling through numerous mechanisms are increasingly recognized in prostate cancer (CaP) progression. We hypothesized that the assessment of well-defined AR transcriptional targets (e.g., PSA/HK3 mRNA) in CaP tissues will provide in vivo readout of AR dysfunctions. Moreover, quantitative expression features of PSA/HK3 mRNA in prostate tumor cells may serve as a prognostic indicator of disease progression.

Experimental Design: Paired benign and malignant epithelial cells (242 specimens) were obtained from laser capture microdissection of frozen OCT-embedded tissue sections prepared from radical prostatectomy specimens of 121 patients. Quantitative expression of PSA/HK3 mRNA in the matched malignant and benign cells was analyzed by real-time reverse transcription-PCR.

Results: CaP cells express significantly lower PSA/HK3 mRNA levels than matched benign cells (P = 0.0133). Moreover, low PSA/HK3 mRNA expression in malignant cells was associated with increased risk of biochemical recurrence (P = 0.0217), as well as with time to recurrence (P = 0.0371), in patients with intermediate preoperative serum prostate-specific antigen levels (2-10 ng/mL). The expression of androgen-dependent genes in clinical samples correlates with each other in patients with higher expression of PSA/HK3 mRNA but not in patients with lower expression of PSA/HK3 mRNA reflecting AR pathway dysfunction.

Conclusions: Our study has unraveled a novel prognostic utility of quantitative measurements of PSA/HK3 mRNA reflecting AR transcriptional activity in CaP cells, which is independent of serum prostate-specific antigen. It also has potential in stratifying subsets of patients exhibiting progressive disease associated with dampened AR transcriptional functions who may be targeted by tailored therapeutic strategies.
Data from Targeting Aberrant PI3K/Akt Activation by PI103 Restores Sensitivity to TRAIL-Induced Apoptosis in Neuroblastoma
Authors: Opel Daniela , Naumann Ivonne , Schneider Maxi , Bertele Daniela , Debatin Klaus-Michael , Fulda Simone
Published: April 2023

Abstract
Purpose: Because we recently identified Akt activation as a novel poor prognostic indicator in neuroblastoma, we investigated whether phosphoinositide 3′-kinase (PI3K) inhibition sensitizes neuroblastoma cells for TRAIL-induced apoptosis.

Experimental Design: The effect of pharmacological or genetic inhibition of PI3K or mTOR was analyzed on apoptosis induction, clonogenic survival, and activation of apoptosis signaling pathways in vitro and in a neuroblastoma in vivo model. The functional relevance of individual Bcl-2 family proteins was examined by knockdown or overexpression experiments.

Results: The PI3K inhibitor PI103 cooperates with TRAIL to synergistically induce apoptosis (combination index < 0.1), to suppress clonogenic survival, and to reduce tumor growth in a neuroblastoma in vivo model. Similarly, genetic silencing of PI3K significantly increases TRAIL-mediated apoptosis, whereas genetic or pharmacological blockage of mTOR fails to potentiate TRAIL-induced apoptosis. Combined treatment with PI103 and TRAIL enhances cleavage of Bid and the insertion of tBid into mitochondrial membranes, and reduces phosphorylation of BimEL. Additionally, PI103 decreases expression of Mcl-1, XIAP, and cFLIP, thereby promoting Bax/Bak activation, mitochondrial perturbations, and caspase-dependent apoptosis. Knockdown of Bid or Noxa or overexpression of Bcl-2 rescues cells from PI103- and TRAIL-induced apoptosis, whereas Mcl-1 silencing potentiates apoptosis. Bcl-2 overexpression also inhibits cleavage of caspase-3, caspase-8, and Bid pointing to a mitochondria-driven feedback amplification loop.

Conclusions: PI103 primes neuroblastoma cells for TRAIL-induced apoptosis by shifting the balance toward proapoptotic Bcl-2 family members and increased mitochondrial apoptosis. Thus, PI3K inhibitors represent a novel promising approach to enhance the efficacy of TRAIL-based treatment protocols in neuroblastoma. Clin Cancer Res; 17(10); 3233–47. ©2011 AACR.
Data from Class I Histone Deacetylase Expression Has Independent Prognostic Impact in Human Colorectal Cancer: Specific Role of Class I Histone Deacetylases In vitro and In vivo
Authors: Weichert Wilko , Röske Annika , Niesporek Silvia , Noske Aurelia , Buckendahl Ann-Christin , Dietel Manfred , Gekeler Volker , Boehm Markus , Beckers Thomas , Denkert Carsten
Published: April 2023

Abstract
Purpose: Recently, several studies reported a strong functional link between histone deacetylases (HDAC) and the development of tumors of the large intestine. However, despite the importance of these molecules, comparably little is known on expression patterns and functions of specific HDAC isoforms in colorectal cancer.

Experimental Design: We characterized class I HDAC isoform expression patterns in a cohort of 140 colorectal carcinomas by immunohistochemistry. In addition, effects of HDAC inhibition by valproic acid and suberoylanilide hydroxamic acid, and specific HDAC isoform knockdown by short interfering RNA, were investigated in a cell culture model.

Results: We found class I HDACs highly expressed in a subset of colorectal carcinomas with positivity for HDAC1 in 36.4%, HDAC2 in 57.9%, and HDAC3 in 72.9% of cases. Expression was significantly enhanced in strongly proliferating (P = 0.002), dedifferentiated (P = 0.022) tumors. High HDAC expression levels implicated significantly reduced patient survival (P = 0.001), with HDAC2 expression being an independent survival prognosticator (hazard ratio, 2.6; P = 0.03). Short interfering RNA–based inhibition of HDAC1 and HDAC2 but not HDAC3 suppressed growth of colon cancer cells in vitro, although to a lesser extent than chemical HDAC inhibitors did.

Conclusions: The strong prognostic impact of HDAC isoforms in colorectal cancer, the interactions of HDACs with tumor cell proliferation and differentiation in vivo, and our finding that HDACs are differentially expressed in colorectal tumors suggest that the evaluation of HDAC expression in clinical trials for HDAC inhibitors might help to identify a patient subgroup who will exceptionally profit from such a treatment.
Data from The Suppression of Aurora-A/STK15/BTAK Expression Enhances Chemosensitivity to Docetaxel in Human Esophageal Squamous Cell Carcinoma
Authors: Tanaka Eiji , Hashimoto Yosuke , Ito Tetsuo , Kondo Kan , Higashiyama Motoshige , Tsunoda Shigeru , Ortiz Cristian , Sakai Yoshiharu , Inazawa Johji , Shimada Yutaka
Published: April 2023

Abstract
Purpose: We previously reported that the expression of Aurora-A was frequently up-regulated in human esophageal squamous cell carcinoma (ESCC) tissues as well as cell lines and the up-regulation contributed to a poor prognosis. In this study, we assessed the possibility of Aurora-A suppression as a therapeutic target for ESCC using ESCC cell lines.

Experimental Design: We established subclones using vector-based short hairpin RNA (shRNA). Then, we investigated the effect of Aurora-A suppression on proliferation and cell cycle changes in vitro. Next, chemosensitivity against docetaxel was investigated by tetrazolium salt–based proliferation assay (WST assay) and cell number determinations, and furthermore, the type of cell death induced by docetaxel was analyzed by flow cytometry. Finally, to examine the effect of Aurora-A shRNA on proliferation and chemosensitivity against docetaxel in vivo, a s.c. tumor formation assay in nude mice was done.

Results: We established two genetically different stable cell lines (510 A and 1440 A) in which levels of Aurora-A were reduced. Cell growth was inhibited by 38.7% in 510 A and by 24.3% in 1440 A in vitro compared with empty vector–transfected controls (510 m and 1440 m), and this growth inhibition was mediated through G2-M arrest as confirmed by flow cytometry. Next, in a WST assay, the IC50 for Aurora-A shRNA-transfected cells was lower than that of empty vector–transfected cells (510 A, 2.7 × 10−7 mol/L; 510 m, 4.8 × 10−7 mol/L; 1440 A, 2.6 × 10−7 mol/L; 1440 m, 4.9 × 10−7 mol/L). In addition, 0.3 nmol/L docetaxel induced a notable level of apoptosis in Aurora-A shRNA-transfected cells compared with empty vector–transfected cells. In the assay of s.c. tumors in nude mice, tumor growth in 510 A was inhibited by 36.1% compared with that in 510 m, and in tumors treated with docetaxel, the suppression of Aurora-A resulted in 44.0% tumor growth suppression in vivo.

Conclusions: These results indicated that Aurora-A might play an important role in chemosensitivity to docetaxel, and the suppression of its expression might be a potential therapeutic target for ESCC.
Data from Antitumor Efficacy of PKI-587, a Highly Potent Dual PI3K/mTOR Kinase Inhibitor
Authors: Mallon Robert , Feldberg Larry R. , Lucas Judy , Chaudhary Inder , Dehnhardt Christoph , Santos Efren Delos , Chen Zecheng , dos Santos Osvaldo , Ayral-Kaloustian Semiramis , Venkatesan Aranapakam , Hollander Irwin
Published: April 2023

Abstract
Purpose: The aim of this study was to show preclinical efficacy and clinical development potential of PKI-587, a dual phosphoinositide 3-kinase (PI3K)/mTOR inhibitor.

Experimental Design: In vitro class 1 PI3K enzyme and human tumor cell growth inhibition assays and in vivo five tumor xenograft models were used to show efficacy.

Results: In vitro, PKI-587 potently inhibited class I PI3Ks (IC50 vs. PI3K-α = 0.4 nmol/L), PI3K-α mutants, and mTOR. PKI-587 inhibited growth of 50 diverse human tumor cell lines at IC50 values of less than 100 nmol/L. PKI-587 suppressed phosphorylation of PI3K/mTOR effectors (e.g., Akt), and induced apoptosis in human tumor cell lines with elevated PI3K/mTOR signaling. MDA-MB-361 [breast; HER2+, PIK3CA mutant (E545K)] was particularly sensitive to this effect, with cleaved PARP, an apoptosis marker, induced by 30 nmol/L PKI-587 at 4 hours.

In vivo, PKI-587 inhibited tumor growth in breast (MDA-MB-361, BT474), colon (HCT116), lung (H1975), and glioma (U87MG) xenograft models. In MDA-MB-361 tumors, PKI-587 (25 mg/kg, single dose i.v.) suppressed Akt phosphorylation [at threonine(T)308 and serine(S)473] for up to 36 hours, with cleaved PARP (cPARP) evident up to 18 hours. PKI-587 at 25 mg/kg (once weekly) shrank large (∼1,000 mm3) MDA-MB-361 tumors and suppressed tumor regrowth. Tumor regression correlated with suppression of phosphorylated Akt in the MDA-MB-361 model. PKI-587 also caused regression in other tumor models, and efficacy was enhanced when given in combination with PD0325901 (MEK 1/2 inhibitor), irinotecan (topoisomerase I inhibitor), or HKI-272 (neratinib, HER2 inhibitor).

Conclusion: Significant antitumor efficacy and a favorable pharmacokinetic/safety profile justified phase 1 clinical evaluation of PKI-587. Clin Cancer Res; 17(10); 3193–203. ©2011 AACR.
Data from Prognostic Value of Tumor Architecture, Tumor-Associated Vascular Characteristics, and Expression of Angiogenic Molecules in Pancreatic Endocrine Tumors
Authors: Takahashi Yu , Akishima-Fukasawa Yuri , Kobayashi Noritoshi , Sano Tsuyoshi , Kosuge Tomoo , Nimura Yuji , Kanai Yae , Hiraoka Nobuyoshi
Published: April 2023

Abstract
Purpose: It is difficult to predict the biological behavior of pancreatic endocrine tumors (PETs). Our aim was to evaluate the prognostic significance of certain variables in PETs.

Experimental Design: The following variables were examined in 37 patients with PETs and then compared with other clinicopathologic characteristics: histologic tumor structure; microvessel density (MVD) measured by three different methods, including a unique method involving calculation of solid area MVD; endothelial proliferation; and the immunohistochemical expression of vascular endothelial growth factor-A and CXC chemokine CXCL-12. Intratumoral vascular structures were analyzed by double immunofluorescence using 30-μm-thick sections.

Results: The presence of focal and intensive solid growth of tumor cells (large solid nests; P = 0.003), low solid area MVD (P = 0.002), a high endothelial cell proliferation index (EPI; P = 0.005), and high expression of CXCL-12 in PET cells (P = 0.018) were significant unfavorable prognostic indicators. The predominant structure of the overall tumor histology and the expression of vascular endothelial growth factor-A did not separate aggressive PETs. In areas of focal solid growth, tumor-associated blood vessels had obviously low MVD and high EPI, and their structures were poorly formed with highly abnormal features, in comparison with other areas. High expression of CXCL-12 in tumor cells was significantly associated with variables representing tumor growth, hematogenous tumor spread, low MVD, high EPI, and the presence of large solid nests.

Conclusions: This study has provided novel findings on the prognostic features of tumor architecture and tumor-associated angiogenesis in PETs. CXCL-12 is the first candidate molecule in association with neoangiogenesis in PETs.
Data from Activation of CCR9/CCL25 in Cutaneous Melanoma Mediates Preferential Metastasis to the Small Intestine
Authors: Amersi Farin F. , Terando Alicia M. , Goto Yasufumi , Scolyer Richard A. , Thompson John F. , Tran Andy N. , Faries Mark B. , Morton Donald L. , Hoon Dave S.B.
Published: April 2023

Abstract
Purpose: Specific chemokines and their respective receptors have been implicated in distant tumor cell metastasis. Cutaneous melanoma has a distinct pattern of metastasis, preferentially targeting the submucosa of the small intestine. However, the underlying pathogenic mechanism remains unknown. Migration of CCR9(+) lymphocytes to the small intestine is known to occur in response to the chemoattractant effects of CCL25 (thymus-expressed chemokine). The integrin heterodimers αβ are also known to be important mediators of cellular adhesion. We hypothesize that the mechanism of small intestinal metastasis by melanoma is via the CCR9-CCL25 axis and specific integrins.

Experimental Design: Quantitative reverse transcription-PCR, flow cytometry, and immunohistochemistry were used to assess melanoma tumors for CCR9 and CCL25. Integrin expression was assessed using flow cytometry. CCR9 expression by quantitative reverse transcription-PCR was assessed in primary (n = 23) and metastatic (n = 198) melanomas, and melanoma lines derived from small intestinal metastases (n = 23).

Results: We showed CCR9 expression in 88 of 102 paraffin-embedded metastatic melanomas from the small intestine, 8 of 8 melanoma lines derived from metastases in the small intestine, and 0 of 96 metastatic melanomas from other sites. In vitro migration and invasion studies done on CCR9(+) melanoma lines showed migration in response to CCL25 that was inhibited by anti-CCR9 antibody or by short interfering RNA CCR9. Flow cytometric analysis confirmed CCR9 expression by melanomas to the small intestine and showed concomitant α4β1 integrin expression.

Conclusions: Our findings show that functionally active CCR9 on melanoma cells facilitates metastasis to the small intestine. The CCR9-CCL25 axis may explain the high incidence of melanoma metastasis to this specific location.
Data from Prognostic DNA Methylation Biomarkers in Ovarian Cancer
Authors: Wei Susan H. , Balch Curtis , Paik Henry H. , Kim Yoo-Sung , Baldwin Rae Lynn , Liyanarachchi Sandya , Li Lang , Wang Zailong , Wan Joseph C. , Davuluri Ramana V. , Karlan Beth Y. , Gifford Gillian , Brown Robert , Kim Sun , Huang Tim H-M. , Nephew Kenneth P.
Published: April 2023

Abstract
Purpose: Aberrant DNA methylation, now recognized as a contributing factor to neoplasia, often shows definitive gene/sequence preferences unique to specific cancer types. Correspondingly, distinct combinations of methylated loci can function as biomarkers for numerous clinical correlates of ovarian and other cancers.

Experimental Design: We used a microarray approach to identify methylated loci prognostic for reduced progression-free survival (PFS) in advanced ovarian cancer patients. Two data set classification algorithms, Significance Analysis of Microarray and Prediction Analysis of Microarray, successfully identified 220 candidate PFS-discriminatory methylated loci. Of those, 112 were found capable of predicting PFS with 95% accuracy, by Prediction Analysis of Microarray, using an independent set of 40 advanced ovarian tumors (from 20 short-PFS and 20 long-PFS patients, respectively). Additionally, we showed the use of these predictive loci using two bioinformatics machine-learning algorithms, Support Vector Machine and Multilayer Perceptron.

Conclusion: In this report, we show that highly prognostic DNA methylation biomarkers can be successfully identified and characterized, using previously unused, rigorous classifying algorithms. Such ovarian cancer biomarkers represent a promising approach for the assessment and management of this devastating disease.
Data from Targeting of Bone-Derived Insulin-Like Growth Factor-II by a Human Neutralizing Antibody Suppresses the Growth of Prostate Cancer Cells in a Human Bone Environment
Authors: Kimura Taichi , Kuwata Takeshi , Ashimine Satoshi , Yamazaki Manabu , Yamauchi Chisako , Nagai Kanji , Ikehara Akashi , Feng Yang , Dimitrov Dimiter S. , Saito Seiichi , Ochiai Atsushi
Published: April 2023

Abstract
Purpose: Advanced prostate cancer frequently involves the bone, where the insulin-like growth factor (IGF)-II is abundant. However, the importance of IGF-II in bone metastasis from prostate cancer is uncertain. The present study was aimed at examining the therapeutic importance of targeting IGF-II in bone metastases from prostate cancer.

Experimental Design: We investigated whether inhibiting IGF-II using a human neutralizing antibody (m610) suppresses the growth of prostate cancer cells in a human bone environment. Human MDA PCa 2b prostate cancer cells were inoculated into human adult bone implanted into mammary fat pad of nonobese diabetic/severe combined immunodeficient mice or inoculated into mammary fat pad of the mice without human bone implantation. The mice were treated with m610 or a control antibody (m102.4) once weekly for 4 weeks immediately after inoculation with MDA PCa 2b cells.

Results: Histomorphologic examination indicated that m610 treatment significantly decreased the MDA PCa 2b tumor area in the human bone compared with the control. Ki-67 immunostaining revealed that the percentage of proliferating cancer cells in the m610-treated bone tumor sections was significantly lower than that in the control. m610 had no effect on MDA PCa 2b tumor growth in the absence of implanted human bone. m610 prevented the in vitro IGF-II–induced proliferation of MDA PCa 2b cells.

Conclusions: Our results indicate that IGF-II plays an important role in the prostate cancer cell growth in human bone, suggesting that targeting it by neutralizing antibodies offers a new therapeutic strategy for bone metastasis from prostate cancer. Clin Cancer Res; 16(1); 121–29
Data from Gene Expression Differences Associated with Human Papillomavirus Status in Head and Neck Squamous Cell Carcinoma
Authors: Slebos Robbert J.C. , Yi Yajun , Ely Kim , Carter Jesse , Evjen Amy , Zhang Xueqiong , Shyr Yu , Murphy Barbara M. , Cmelak Anthony J. , Burkey Brian B. , Netterville James L. , Levy Shawn , Yarbrough Wendell G. , Chung Christine H.
Published: April 2023

Abstract
Human papillomavirus (HPV) is associated with a subset of head and neck squamous cell carcinoma (HNSCC). Between 15% and 35% of HNSCCs harbor HPV DNA. Demographic and exposure differences between HPV-positive (HPV+) and negative (HPV−) HNSCCs suggest that HPV+ tumors may constitute a subclass with different biology, whereas clinical differences have also been observed. Gene expression profiles of HPV+ and HPV− tumors were compared with further exploration of the biological effect of HPV in HNSCC. Thirty-six HNSCC tumors were analyzed using Affymetrix Human 133U Plus 2.0 GeneChip and for HPV by PCR and real-time PCR. Eight of 36 (22%) tumors were positive for HPV subtype 16. Statistical analysis using Significance Analysis of Microarrays based on HPV status as a supervising variable resulted in a list of 91 genes that were differentially expressed with statistical significance. Results for a subset of these genes were verified by real-time PCR. Genes highly expressed in HPV+ samples included cell cycle regulators (p16INK4A, p18, and CDC7) and transcription factors (TAF7L, RFC4, RPA2, and TFDP2). The microarray data were also investigated by mapping genes by chromosomal location (DIGMAP). A large number of genes on chromosome 3q24-qter had high levels of expression in HPV+ tumors. Further investigation of differentially expressed genes may reveal the unique pathways in HPV+ tumors that may explain the different natural history and biological properties of these tumors. These properties may be exploited as a target of novel therapeutic agents in HNSCC treatment.
Data from Valproic Acid at Therapeutic Plasma Levels May Increase 5-Azacytidine Efficacy in Higher Risk Myelodysplastic Syndromes
Authors: Voso Maria Teresa , Santini Valeria , Finelli Carlo , Musto Pellegrino , Pogliani Enrico , Angelucci Emanuele , Fioritoni Giuseppe , Alimena Giuliana , Maurillo Luca , Cortelezzi Agostino , Buccisano Francesco , Gobbi Marco , Borin Lorenza , Di Tucci Anna , Zini Gina , Petti Maria Concetta , Martinelli Giovanni , Fabiani Emiliano , Fazi Paola , Vignetti Marco , Piciocchi Alfonso , Liso Vincenzo , Amadori Sergio , Leone Giuseppe
Published: April 2023

Abstract
Purpose: Epigenetic changes play a role and cooperate with genetic alterations in the pathogenesis of myelodysplastic syndromes (MDS). We conducted a phase II multicenter study on the combination of the DNA-methyltransferase inhibitor 5-azacytidine (5-AZA) and the histone deacetylase inhibitor valproic acid (VPA) in patients with higher risk MDS.

Experimental Design: We enrolled 62 patients with MDS (refractory anemia with excess blasts, 39 patients; refractory anemia with excess blasts in transformation, 19 patients; and chronic myelomanocytic leukemia (CMML), 4 patients) and an International Prognostic Scoring System (IPSS) rating of Intermediate-2 (42 patients) or high (20 patients). VPA was given to reach a plasma concentration of >50 μg/mL, then 5-AZA was added s.c. at 75 mg/m2 for 7 days in eight monthly cycles.

Results: The median overall survival was 14.4 months. At a median follow-up of 12 months (range, 0.7-21.0), the disease progressed in 20 patients, with 21% cumulative incidence of progression. Of 26 patients who completed eight cycles, 30.7% obtained complete or partial remission, 15.4% had a major hematologic improvement, whereas 38.5% showed stable disease. Drug-related toxicity was mild. Favorable prognostic factors for survival were IPSS Intermediate-2 and plasma VPA of ≥50 μg/mL (log rank = 0.013 and 0.007, respectively). Analysis of polymorphisms important for the metabolism of the drugs used in the trial showed that carriers of the CYP2C19*2 variant of cytochrome P450 required higher VPA doses to achieve the target VPA plasma concentration of 50 μg/mL on day 1 of 5-AZA treatment (P = 0.0021).

Conclusion: Our data show that the 5-AZA/VPA combination is active and safe in patients with MDS with a poor prognosis. Achievement of VPA therapeutic levels may indeed increase 5-AZA efficacy.
Data from Detection of Tumor DNA at the Margins of Colorectal Cancer Liver Metastasis
Authors: Holdhoff Matthias , Schmidt Kerstin , Diehl Frank , Aggrawal Nishant , Angenendt Philipp , Romans Katharine , Edelstein Daniel L. , Torbenson Michael , Kinzler Kenneth W. , Vogelstein Bert , Choti Michael A. , Diaz Luis A.
Published: April 2023

Abstract
Purpose: Defining an adequate resection margin of colorectal cancer liver metastases is essential for optimizing surgical technique. We have attempted to evaluate the resection margin through a combination of histopathologic and genetic analyses.

Experimental Design: We evaluated 88 samples of tumor margins from 12 patients with metastatic colon cancer who each underwent partial hepatectomy of one to six liver metastases. Punch biopsies of surrounding liver tissue were obtained at 4, 8, 12, and 16 mm from the tumor border. DNA from these biopsies was analyzed by a sensitive PCR-based technique, called BEAMing, for mutations of KRAS, PIK3CA, APC, or TP53 identified in the corresponding tumor.

Results: Mutations were identified in each patient's resected tumor and used to analyze the 88 samples circumscribing the tumor-normal border. Tumor-specific mutant DNA was detectable in surrounding liver tissue in 5 of these 88 samples, all within 4 mm of the tumor border. Biopsies that were 8, 12, and 16 mm from the macroscopic visible margin were devoid of detectable mutant tumor DNA and of microscopically visible cancer cells. Tumors with a significant radiologic response to chemotherapy were not associated with any increase in mutant tumor DNA in beyond 4 mm of the main tumor.

Conclusions: Mutant tumor-specific DNA can be detected beyond the visible tumor margin, but never beyond 4 mm, even in patients whose tumors were larger prior to chemotherapy. These data provide a rational basis for determining the extent of surgical excision required in patients undergoing resection of liver metastases. Clin Cancer Res; 17(11); 3551–7. ©2011 AACR.
Data from A Gene Expression–Based Predictor for Myeloma Patients at High Risk of Developing Bone Disease on Bisphosphonate Treatment
Authors: Wu Ping , Walker Brian A. , Brewer Daniel , Gregory Walter M. , Ashcroft John , Ross Fiona M. , Jackson Graham H. , Child Anthony J. , Davies Faith E. , Morgan Gareth J.
Published: April 2023

Abstract
Purpose: Myeloma bone disease impairs quality of life and is associated with impaired survival. Even with effective bisphosphonate treatment, a significant proportion of patients still develop skeletal-related events (SRE). Identifying such patients at presentation would allow treatment modification.

Experimental Design: To investigate the molecular basis of bone disease at presentation and to develop a predictive signature for patients at high risk of developing SREs on bisphosphonates, 261 presenting myeloma samples were analyzed by global gene expression profiling. The derived “SRE gene signature” was complemented by the integration of associated clinical parameters to generate an optimal predictor.

Results: Fifty genes were significantly associated with presenting bone disease, including the WNT signaling antagonist DKK1 and genes involved in growth factor signaling and apoptosis. Higher serum calcium level and the presence of bone disease and hyperdiploidy at presentation were associated with high risk of SRE development. A gene signature derived from the fourteen genes overexpressed in the SRE group was able to identify patients at high risk of developing an SRE on treatment. These genes either belonged to the IFN-induced family or were involved in cell signaling and mitosis. Multivariate logistic model selection yielded an optimal SRE predictor comprising seven genes and calcium level, which was validated as an effective predictor in a further set of patients.

Conclusions: The simple expression-based SRE predictor can effectively identify individuals at high risk of developing bone disease while being on bisphosphonates. This predictor could assist with developing future trials on novel therapies aimed at reducing myeloma bone disease. Clin Cancer Res; 17(19); 6347–55. ©2011 AACR.
Data from Clinical Responses in a Phase II Study Using Adoptive Transfer of Short-term Cultured Tumor Infiltration Lymphocytes in Metastatic Melanoma Patients
Authors: Besser Michal J. , Shapira-Frommer Ronnie , Treves Avraham J. , Zippel Dov , Itzhaki Orit , Hershkovitz Liat , Levy Daphna , Kubi Adva , Hovav Einat , Chermoshniuk Natalia , Shalmon Bruria , Hardan Izhar , Catane Raphael , Markel Gal , Apter Sara , Ben-Nun Alon , Kuchuk Iryna , Shimoni Avichai , Nagler Arnon , Schachter Jacob
Published: April 2023

Abstract
Purpose: Adoptive cell therapy with autologous tumor-infiltrating lymphocytes (TIL) has shown promising results in metastatic melanoma patients. Although objective response rates of over 50% have been reported, disadvantages of this approach are the labor-intensive TIL production and a very high drop-out rate of enrolled patients, limiting its widespread applicability.

Previous studies showed a clear correlation between short TIL culture periods and clinical response. Therefore, we used a new TIL production technique using unselected, minimally cultured, bulk TIL (Young-TIL). The use of Young-TIL is not restricted to human leukocyte antigen (HLA)-A2 patients.

The purpose of this study is to explore the efficacy and toxicity of adoptively transferred Young-TIL following lympho-depleting chemotherapy in metastatic melanoma patients, refractory to interleukin-2 and chemotherapy.

Experimental Design: Young-TIL cultures for 90% of the patients were successfully generated, enabling the treatment of most enrolled patients. We report here the results of 20 evaluated patients.

Results: Fifty percent of the patients achieved an objective clinical response according to the Response Evaluation Criteria in Solid Tumors, including two ongoing complete remissions (20+, 4+ months) and eight partial responses (progression-free survival: 18+, 13+, 10+, 9, 6+, 4, 3+, and 3 months). All responders are currently alive. Four additional patients showed disease stabilization. Side effects were transient and manageable.

Conclusion: We showed that lympho-depleting chemotherapy followed by transfer of short-term cultured TIL can mediate tumor regression in 50% of metastatic melanoma with manageable toxicity. The convincing clinical results combined with the simplification of the process may thus have a major effect on cell therapy of cancer. Clin Cancer Res; 16(9); 2646–55. ©2010 AACR.
Data from Successful Eradication of Established Peritoneal Ovarian Tumors in SCID-Beige Mice following Adoptive Transfer of T Cells Genetically Targeted to the MUC16 Antigen
Authors: Chekmasova Alena A. , Rao Thapi D. , Nikhamin Yan , Park Kay J. , Levine Douglas A. , Spriggs David R. , Brentjens Renier J.
Published: April 2023

Abstract
Purpose: Most patients diagnosed with ovarian cancer will ultimately die from their disease. For this reason, novel approaches to the treatment of this malignancy are needed. Adoptive transfer of a patient's own T cells, genetically modified ex vivo through the introduction of a gene encoding a chimeric antigen receptor (CAR) targeted to a tumor-associated antigen, is a novel approach to the treatment of ovarian cancer.

Experimental Design: We have generated several CARs targeted to the retained extracellular domain of MUC16, termed MUC-CD, an antigen expressed on most ovarian carcinomas. We investigate the in vitro biology of human T cells retrovirally transduced to express these CARs by coculture assays on artificial antigen-presenting cells as well as by cytotoxicity and cytokine release assays using the human MUC-CD+ ovarian tumor cell lines and primary patient tumor cells. Further, we assess the in vivo antitumor efficacy of MUC-CD–targeted T cells in SCID-Beige mice bearing peritoneal human MUC-CD+ tumor cell lines.

Results: CAR-modified, MUC-CD–targeted T cells exhibited efficient MUC-CD–specific cytolytic activity against both human ovarian cell and primary ovarian carcinoma cells in vitro. Furthermore, expanded MUC-CD–targeted T cells infused through either i.p. injection or i.v. infusion into SCID-Beige mice bearing orthotopic human MUC-CD+ ovarian carcinoma tumors either delayed progression or fully eradicated disease.

Conclusion: These promising preclinical studies justify further investigation of MUC-CD–targeted T cells as a potential therapeutic approach for patients with high-risk MUC16+ ovarian carcinomas. Clin Cancer Res; 16(14); 3594–606. ©2010 AACR.
Data from Prediction of Postoperative Recurrence-Free Survival in Non–Small Cell Lung Cancer by Using an Internationally Validated Gene Expression Model
Authors: Mitra Ranjana , Lee Jinseon , Jo Jisuk , Milani Monica , McClintick Jeanette N. , Edenberg Howard J. , Kesler Kenneth A. , Rieger Karen M. , Badve Sunil , Cummings Oscar W. , Mohiuddin Ahmed , Thomas Dafydd G. , Luo Xianghua , Juliar Beth E. , Li Lang , Mesaros Clementina , Blair Ian A. , Srirangam Anjaiah , Kratzke Robert A. , McDonald Clement J. , Kim Jhingook , Potter David A.
Published: April 2023

Abstract
Purpose: This study was performed to discover prognostic genomic markers associated with postoperative outcome of stage I to III non–small cell lung cancer (NSCLC) that are reproducible between geographically distant and demographically distinct patient populations.

Experimental Design: American patients (n = 27) were stratified on the basis of recurrence and microarray profiling of their tumors was performed to derive a training set of 44 genes. A larger Korean patient validation cohort (n = 138) was also stratified by recurrence and screened for these genes. Four reproducible genes were identified and used to construct genomic and clinicogenomic Cox models for both cohorts.

Results: Four genomic markers, DBN1 (drebrin 1), CACNB3 (calcium channel beta 3), FLAD1 (PP591; flavin adenine dinucleotide synthetase), and CCND2 (cyclin D2), exhibited highly significant differential expression in recurrent tumors in the training set (P < 0.001). In the validation set, DBN1, FLAD1 (PP591), and CACNB3 were significant by Cox univariate analysis (P ≤ 0.035), whereas only DBN1 was significant by multivariate analysis. Genomic and clinicogenomic models for recurrence-free survival (RFS) were equally effective for risk stratification of stage I to II or I to III patients (all models P < 0.0001). For stage I to II or I to III patients, 5-year RFS of the low- and high-risk patients was approximately 70% versus 30% for both models. The genomic model for overall survival of stage I to III patients was improved by addition of pT and pN stage (P < 0.0013 vs. 0.010).

Conclusion: A 4-gene prognostic model incorporating the multivariate marker DBN1 exhibits potential clinical utility for risk stratification of stage I to III NSCLC patients. Clin Cancer Res; 17(9); 2934–46. ©2011 AACR.
Data from Vandetanib Restores Head and Neck Squamous Cell Carcinoma Cells' Sensitivity to Cisplatin and Radiation In Vivo and In Vitro
Authors: Sano Daisuke , Matsumoto Fumihiko , Valdecanas David R. , Zhao Mei , Molkentine David P. , Takahashi Yoko , Hanna Ehab Y. , Papadimitrakopoulou Vali , Heymach John , Milas Luka , Myers Jeffrey N.
Published: April 2023

Abstract
Purpose: We investigated whether vandetanib, an inhibitor of the tyrosine kinase activities of vascular endothelial growth factor receptor-2 (VEGFR-2), epidermal growth factor receptor (EGFR), and rearranged during transfection (RET), could augment the antitumor activity of radiation with or without cisplatin in preclinical in vitro and in vivo models of human head and neck squamous cell carcinoma (HNSCC).

Experimental Design: OSC-19 and HN5 HNSCC cells that were cisplatin and radioresistant were treated with vandetanib, cisplatin, and radiation alone or in combination in vitro and in vivo using an orthotopic nude mouse model. Treatment effects were assessed using clonogenic survival assay, tumor volume, bioluminescence imaging, tumor growth delay, survival, microvessel density, tumor and endothelial cell apoptosis, and EGFR and Akt phosphorylation data.

Results: Vandetanib plus cisplatin radiosensitized HNSCC cells in vitro and in vivo. The combination treatment with vandetanib, cisplatin, and radiation was superior to the rest of treatments (including the double combinations) in antitumoral effects, prolonging survival, decreasing cervical lymph node metastases in vivo. It also increased both tumor and tumor-associated endothelial cell apoptosis and decreased microvessel density in vivo. An analysis of tumor growth delay data revealed that vandetanib plus cisplatin enhanced radioresponse in vivo. All vandetanib-containing treatments inhibited EGFR and Akt phosphorylation in vitro and in vivo.

Conclusion: The addition of vandetanib to combination therapy with cisplatin and radiation was able to effectively overcome cisplatin and radioresistance in in vitro and in vivo models of HNSCC. Further study of this regimen in clinical trials may be warranted. Clin Cancer Res; 17(7); 1815–27. ©2011 AACR.
Data from Targeted Inhibition of Inducible Nitric Oxide Synthase Inhibits Growth of Human Melanoma In vivo and Synergizes with Chemotherapy
Authors: Sikora Andrew G. , Gelbard Alexander , Davies Michael A. , Sano Daisuke , Ekmekcioglu Suhendan , Kwon John , Hailemichael Yared , Jayaraman Padmini , Myers Jeffrey N. , Grimm Elizabeth A. , Overwijk Willem W.
Published: April 2023

Abstract
Purpose: Aberrant expression of inflammatory molecules, such as inducible nitric oxide (NO) synthase (iNOS), has been linked to cancer, suggesting that their inhibition is a rational therapeutic approach. Whereas iNOS expression in melanoma and other cancers is associated with poor clinical prognosis, in vitro and in vivo studies suggest that iNOS and NO can have both protumor and antitumor effects. We tested the hypothesis that targeted iNOS inhibition would interfere with human melanoma growth and survival in vivo in a preclinical model.

Experimental Design: We used an immunodeficient non-obese diabetic/severe combined immunodeficient xenograft model to test the susceptibility of two different human melanoma lines to the orally-given iNOS-selective small molecule antagonist N6-(1-iminoethyl)-l-lysine-dihydrochloride (L-nil) with and without cytotoxic cisplatin chemotherapy.

Results: L-nil significantly inhibited melanoma growth and extended the survival of tumor-bearing mice. L-nil treatment decreased the density of CD31+ microvessels and increased the number of apoptotic cells in tumor xenografts. Proteomic analysis of melanoma xenografts with reverse-phase protein array identified alterations in the expression of multiple cell signaling and survival genes after L-nil treatment. The canonical antiapoptotic protein Bcl-2 was downregulated in vivo and in vitro after L-nil treatment, which was associated with increased susceptibility to cisplatin-mediated tumor death. Consistent with this observation, combination therapy with L-nil plus cisplatin in vivo was more effective than either drug alone, without increased toxicity.

Conclusions: These data support the hypothesis that iNOS and iNOS-derived NO support tumor growth in vivo and provide convincing preclinical validation of targeted iNOS inhibition as therapy for solid tumors. Clin Cancer Res; 16(6); 1834–44
Data from A Gene Expression Signature from Human Breast Cancer Cells with Acquired Hormone Independence Identifies MYC as a Mediator of Antiestrogen Resistance
Authors: Miller Todd W. , Balko Justin M. , Ghazoui Zara , Dunbier Anita , Anderson Helen , Dowsett Mitch , González-Angulo Ana M. , Mills Gordon B. , Miller William R. , Wu Huiyun , Shyr Yu , Arteaga Carlos L.
Published: April 2023

Abstract
Purpose: Although most patients with estrogen receptor α (ER)-positive breast cancer initially respond to endocrine therapy, many ultimately develop resistance to antiestrogens. However, mechanisms of antiestrogen resistance and biomarkers predictive of such resistance are underdeveloped.

Experimental Design: We adapted four ER+ human breast cancer cell lines to grow in an estrogen-depleted medium. A gene signature of estrogen independence was developed by comparing expression profiles of long-term estrogen-deprived (LTED) cells to their parental counterparts. We evaluated the ability of the LTED signature to predict tumor response to neoadjuvant therapy with an aromatase inhibitor and disease outcome following adjuvant tamoxifen. We utilized Gene Set Analysis (GSA) of LTED cell gene expression profiles and a loss-of-function approach to identify pathways causally associated with resistance to endocrine therapy.

Results: The LTED gene expression signature was predictive of high tumor cell proliferation following neoadjuvant therapy with anastrozole and letrozole, each in different patient cohorts. This signature was also predictive of poor recurrence-free survival in two studies of patients treated with adjuvant tamoxifen. Bioinformatic interrogation of expression profiles in LTED cells revealed a signature of MYC activation. The MYC activation signature and high MYC protein levels were both predictive of poor outcome following tamoxifen therapy. Finally, knockdown of MYC inhibited LTED cell growth.

Conclusions: A gene expression signature derived from ER+ breast cancer cells with acquired hormone independence predicted tumor response to aromatase inhibitors and associated with clinical markers of resistance to tamoxifen. Activation of the MYC pathway was associated with this resistance. Clin Cancer Res; 17(7); 2024–34. ©2011 AACR.