bluuebunny/crossref_metadata_2025 huggingface dataset:
Messenger RNA Expression Ratios among Four Genes Predict Subtypes of Renal Cell Carcinoma and Distinguish Oncocytoma from Carcinoma
Authors: Chen Yao-Tseng , Tu Jiangling J. , Kao Jean , Zhou Xi K. , Mazumdar Madhu
Published: September 2005

Purpose: Morphologic distinction among clear cell, papillary, and chromophobe types of renal cell carcinoma (RCC) can be difficult, as is the differential diagnosis between oncocytoma and RCC. Whether these renal tumors can be distinguished by their mRNA expression profile of a few selected genes was examined.

Experimental Design: The expression of four genes in renal tumor was evaluated by quantitative reverse transcription-PCR: carbonic anhydrase IX (CA9), methylacyl-CoA racemase (AMACR), parvalbumin (PVALB), and chloride channel kb (CLCNKB). Thirty-one fresh-frozen and 63 formalin-fixed, paraffin-embedded tumor specimens were analyzed.

Results: CA9 expression was highest in clear cell carcinoma and lowest in chromophobe RCC and in oncocytoma. AMACR expression was highest in papillary RCC, and CLCNKB was highest in chromophobe RCC/oncocytoma. PVALB was highest in chromophobe RCC, variable in oncocytoma, and low in clear cell and papillary types. Similar findings were observed in fresh-frozen and formalin-fixed specimens. The mRNA expression ratios among these genes (i.e., CA9/AMACR and AMACR/CLCNKB ratios) further accentuate the gene expression differences among these tumors, and a molecular diagnostic algorithm was established. This algorithm accurately classified the 31 fresh-frozen tumors into 14 clear cell, 5 papillary, 6 chromophobe, and 6 oncocytomas. In the formalin-fixed group, the molecular criteria accurately classified the cases into 15 clear cell, 16 papillary, and 32 in the chromophobe/oncocytoma group but could only separate some, but not all, oncocytomas from chromophobe RCC.

Conclusions: RNA expression ratios based on the four-gene panel can accurately classify subtypes of RCC as well as help distinguish some oncocytomas from chromophobe RCC.
Phase 1 Study of ABT-751, a Novel Microtubule Inhibitor, in Patients with Refractory Hematologic Malignancies
Authors: Yee Karen W.L. , Hagey Anne , Verstovsek Srdan , Cortes Jorge , Garcia-Manero Guillermo , O'Brien Susan M. , Faderl Stefan , Thomas Deborah , Wierda William , Kornblau Steven , Ferrajoli Alessandra , Albitar Maher , McKeegan Evelyn , Grimm David R. , Mueller Toby , Holley-Shanks Rhonda R. , Sahelijo Leonardo , Gordon Gary B. , Kantarjian Hagop M. , Giles Francis J.
Published: September 2005

Purpose: ABT-751 is an oral antimitotic agent that binds to the colchicine site on β-tubulin. A phase 1 study was conducted to determine the maximum tolerated dose and toxicities of ABT-751 in patients with advanced myelodysplastic syndrome and relapsed or refractory acute leukemias.

Study Design: Thirty-two patients were treated: nine with 100 (n = 3), 125 (n = 3), or 150 mg/m2 (n = 3) of ABT-751 given orally once daily for 7 days every 3 weeks and 23 with 75 (n = 3), 100 (n = 3), 125 (n = 5), 150 (n = 5), 175 (n = 3), or 200 mg/m2 (n = 4) of ABT-751 given orally once daily for 21 days every 4 weeks. Consenting patients had pharmacogenetic sampling and enumeration of circulating endothelial cells (CEC).

Results: Dose-limiting toxicity consisted of ileus in one patient at 200 mg/m2, with a subsequent patient developing grade 2 constipation at the same dose level. One patient with relapsed acute myelogenous leukemia achieved a complete remission that was sustained for 2 months. Four other patients had transient hematologic improvements, consisting of a decrease in peripheral blood blasts and improvements in platelet counts. CEC number was reduced in three patients with a concomitant reduction in peripheral blasts. A previously undescribed nonsynonymous single nucleotide polymorphism, encoding Ala185Thr, was identified in exon 4 of the β-tubulin gene, TUBB, in three other patients. The recommended phase 2 dose in hematologic malignancies is 175 mg/m2 daily orally for 21 days every 4 weeks.

Conclusion: Further assessment of ABT-751, especially in combination with other agents, in patients with acute leukemias is warranted.
Induction of Higher-Avidity Human CTLs by Vector-Mediated Enhanced Costimulation of Antigen-Presenting Cells
Authors: Yang Sixun , Tsang Kwong-Yok , Schlom Jeffrey
Published: September 2005

The efficacy of antigen-specific CD8+ CTLs depends not only on the quantity of CTLs generated but also perhaps, more importantly, on the avidity of the CTLs. To date, however, no strategy has been shown to preferentially induce higher-avidity human CTLs. In the present study, antigen-presenting cells (APC) generated from human peripheral blood mononuclear cells were infected with a recombinant avipox vector (rF-) containing the transgenes for a triad of costimulatory molecules (human B7.1, intercellular adhesion molecule-1, and LFA-3, designated as rF-TRICOM) and then used to elicit peptide-specific CTLs from autologous T cells. Compared with peptide-pulsed noninfected APCs or peptide-pulsed APCs infected with wild-type vector, peptide-pulsed APCs infected with rF-TRICOM induced not only more CTLs but also higher-avidity CTLs; this was shown by tetramer staining, tetramer dissociation, IFN-γ production, and cytolytic assays. Peptide-pulsed rF-TRICOM-infected dendritic cells were also shown to induce CTLs with a >10-fold higher avidity than CTLs induced using CD40L-matured dendritic cells; the use of peptide-pulsed CD40L-matured dendritic cells infected with rF-TRICOM as APCs induced CTLs of even greater avidity. To our knowledge, these studies are the first to show a methodology to induce higher-avidity human CTLs and have implications for the development of more efficient vaccines for a range of human cancers.
Multiple Mechanisms of Telomere Maintenance Exist in Liposarcomas
Authors: Johnson Jay E. , Varkonyi Robert J. , Schwalm Jaclyn , Cragle Ryan , Klein-Szanto Andres , Patchefsky Arthur , Cukierman Edna , von Mehren Margaret , Broccoli Dominique
Published: September 2005

Purpose: Telomeres are specialized nucleoprotein complexes that protect and confer stability upon chromosome ends. Loss of telomere function as a consequence of proliferation-associated sequence attrition results in genome instability, which may facilitate carcinogenesis by generating growth-promoting mutations. However, unlimited cellular proliferation requires the maintenance of telomeric DNA; thus, the majority of tumor cells maintain their telomeres either through the activity of telomerase or via a mechanism known as alternative lengthening of telomeres (ALT). Recent data suggest that constitutive telomere maintenance may not be required in all tumor types. Here we assess the role and requirement of telomere maintenance in liposarcoma.

Experimental Design: Tumor samples were analyzed with respect to telomerase activity, telomere length, and the presence of ALT-specific subcellular structures, ALT-associated promyelocytic leukemia nuclear bodies. This multiassay assessment improved the accuracy of categorization.

Results: Our data reveal a significant incidence (24%) of ALT-positive liposarcomas, whereas telomerase is used at a similar frequency (27%). A large number of tumors (49%) do not show characteristics of telomerase or ALT. In addition, telomere length was always shorter in recurrent disease, regardless of the telomere maintenance mechanism.

Conclusions: These results suggest that approximately one half of liposarcomas either employ a novel constitutively active telomere maintenance mechanism or lack such a mechanism. Analysis of recurrent tumors suggests that liposarcomas can develop despite limiting or undetectable activity of a constitutively active telomere maintenance mechanism.
Combining Etoposide and Dexrazoxane Synergizes with Radiotherapy and Improves Survival in Mice with Central Nervous System Tumors
Authors: Hofland Kenneth Francis , Thougaard Annemette Vinding , Dejligbjerg Marielle , Jensen Lars H. , Kristjansen Paul E.G. , Rengtved Pia , Sehested Maxwell , Jensen Peter Buhl
Published: September 2005

Purpose: The treatment of patients with brain metastases is presently ineffective, but cerebral chemoradiotherapy using radiosensitizing agents seems promising. Etoposide targets topoisomerase II, resulting in lethal DNA breaks; such lesions may increase the effect of irradiation, which also depends on DNA damage. Coadministration of the topoisomerase II catalytic inhibitor dexrazoxane in mice allows for more than 3-fold higher dosing of etoposide. We hypothesized that dexrazoxane combined with escalated etoposide doses might improve the efficacy of cerebral radiotherapy.

Experimental Design: Mice with cerebrally inoculated Ehrlich ascites tumor (EHR2) cells were treated with combinations of etoposide + dexrazoxane + cerebral radiotherapy. Similar chemotherapy and radiation combinations were investigated by clonogenic assays using EHR2 cells, and by DNA double-strand break assay through quantification of phosphorylated histone H2AX (γH2AX).

Results: Escalated etoposide dosing (90 mg/kg) combined with dexrazoxane (125 mg/kg) and cerebral radiotherapy (10 Gy × 1) increased the median survival by 60% (P = 0.001) without increased toxicity, suggesting that escalated etoposide levels may indeed represent a new strategy for improving radiotherapy. Interestingly, 125 mg/kg dexrazoxane combined with normal etoposide doses (34 mg/kg) also increased survival from radiotherapy, but only by 27% (P = 0.002). This indicates a direct dexrazoxane modulation of the combined effects of etoposide and radiation in brain tumors. Further, in vitro, concurrent dexrazoxane, etoposide, and irradiation significantly increased DNA double-strand breaks.

Conclusion: Combining etoposide (high or normal doses) and dexrazoxane synergizes with cerebral radiotherapy and significantly improves survival in mice with central nervous system tumors. This regimen may thus improve radiation therapy of central nervous system tumors.
Induction of CD4+ and CD8+ T-Cell Responses to the Human Stromal Antigen, Fibroblast Activation Protein: Implication for Cancer Immunotherapy
Authors: Fassnacht Martin , Lee Jaewoo , Milazzo Caterina , Boczkowski David , Su Zhen , Nair Smita , Gilboa Eli
Published: September 2005

Purpose: The propensity of tumor cells to escape immune elimination could limit, if not defeat, the long-term benefits of effective immunotherapeutic protocols. Immunologic targeting of tumor stroma could significantly reduce the ability of tumors to evade immune elimination. Murine studies have shown that inducing immunity against angiogenesis-associated products engenders potent antitumor immunity without significant pathology. It is, however, not known whether T cells corresponding to stromal products are present in humans. In this study, we describe a method to screen for human stromal products that have not triggered significant tolerance and could therefore serve as candidate antigens for cancer immunotherapy.

Experimental Design: To identify candidates for human stromal antigens, we used an in vitro–screening method to determine whether dendritic cells transfected with mRNA encoding products, which are overexpressed in the tumor stroma, are capable of stimulating cytotoxic CD8+ (CTL) responses from human peripheral blood mononuclear cells.

Results: CTL responses could be consistently generated against fibroblast activation protein (FAP) but not against matrix metalloproteinase-9 (MMP-9) or MMP-14. To enhance the immunogenicity of the mRNA-translated FAP product, a lysosomal targeting signal derived from lysosome-associated membrane protein-1 (LAMP-1) was fused to the COOH terminus of FAP to redirect the translated product into the class II presentation pathway. Dendritic cells transfected with mRNA encoding the FAP-LAMP fusion product stimulated enhanced CD4+ and CD8+ T-cell responses.

Conclusion: This study identifies FAP, a protease preferentially expressed in tumor-associated fibroblasts, as a candidate human stromal antigen to target in the setting of cancer immunotherapy, and shows that differential expression of stromal products is not a sufficient criteria to indicate its immunogenicity in a vaccination setting.
Effects of Exercise Training on Antitumor Efficacy of Doxorubicin in MDA-MB-231 Breast Cancer Xenografts
Authors: Jones Lee W. , Eves Neil D. , Courneya Kerry S. , Chiu Brian K. , Baracos Vickie E. , Hanson John , Johnson Lorelei , Mackey John R.
Published: September 2005

Purpose: Exercise is becoming readily accepted as a beneficial adjunct therapy to maintain or enhance quality of life in breast cancer patients undergoing adjuvant chemotherapy. An essential precursor to these studies is to investigate whether exercise modulates the antitumor efficacy of chemotherapeutic agents.

Experimental Design: Athymic female mice were transplanted with MDA-MB-231 breast xenografts and randomly assigned to one of four groups (n = 21 per group): (a) control, (b) exercise-only, (c) doxorubicin-only, or (d) exercise + doxorubicin. Exercise groups performed progressive treadmill running up to 18 m/min at 0% grade for 45 minutes, 5 d/wk for 8 weeks.

Results: Tumor growth delay was significantly longer in the doxorubicin-only and exercise + doxorubicin groups compared with the control (median 42 versus 25 days, P = 0.0082; 36 versus 25 days, P = 0.029, respectively) and exercise-only groups (median 42 versus 25 days, P = 0.029; 36 versus 25 days, P = 0.080, respectively). There was no significant difference between the doxorubicin-only and exercise + doxorubicin groups (median 42 versus 36 days, P = 0.33), suggesting that moderate intensity exercise does not significantly influence doxorubicin-induced tumor growth delay.

Conclusion: These studies are essential to fully understand the safety and application of exercise as a supportive intervention in cancer control.
Gemcitabine Selectively Eliminates Splenic Gr-1+/CD11b+ Myeloid Suppressor Cells in Tumor-Bearing Animals and Enhances Antitumor Immune Activity
Authors: Suzuki Eiji , Kapoor Veena , Jassar Arminder Singh , Kaiser Larry R. , Albelda Steven M.
Published: September 2005

Purpose: Myeloid suppressor (Gr-1+/CD11b+) cells accumulate in the spleens of tumor-bearing mice where they contribute to immunosuppression by inhibiting the function of CD8+ T cells and by promoting tumor angiogenesis. Elimination of these myeloid suppressor cells may thus significantly improve antitumor responses and enhance effects of cancer immunotherapy, although to date few practical options exist.

Experimental Design: The effect of the chemotherapy drug gemcitabine on the number of (Gr-1+/CD11b+) cells in the spleens of animals bearing large tumors derived from five cancer lines grown in both C57Bl/6 and BALB/c mice was analyzed. Suppressive activity of splenocytes from gemcitabine-treated and control animals was measured in natural killer (NK) cell lysis and Winn assays. The impact of myeloid suppressor cell activity was determined in an immunogene therapy model using an adenovirus expressing IFN-β.

Results: This study shows that the chemotherapeutic drug gemcitabine, given at a dose similar to the equivalent dose used in patients, was able to dramatically and specifically reduce the number of myeloid suppressor cells found in the spleens of animals bearing large tumors with no significant reductions in CD4+ T cells, CD8+ T cells, NK cells, macrophages, or B cells. The loss of myeloid suppressor cells was accompanied by an increase in the antitumor activity of CD8+ T cells and activated NK cells. Combining gemcitabine with cytokine immunogene therapy using IFN-β markedly enhanced antitumor efficacy.

Conclusions: These results suggest that gemcitabine may be a practical strategy for the reduction of myeloid suppressor cells and should be evaluated in conjunction with a variety of immunotherapy approaches.
Study Design Issues and Early Stage Non–Small Cell Lung Cancer
Authors: Schiller Joan H.
Published: September 2005

The most common type of clinical trial in oncology enrolls patients with advanced disease. End points for phase II trials in advanced disease typically include response rate or time to progression, based upon the presumption that these may serve as surrogates for the ultimate end point of improved survival. Unique problems arise with the design of adjuvant trials, for which response rates clearly are not appropriate end points. In addition, survival is much longer, making rapid completion of phase III trials difficult. For lung cancer, the proportion of patients with stage I and II disease is much lower than with other types of cancer, and the absolute number of these patients is relatively low. To conduct large adjuvant trials in lung cancer more efficiently, we need to optimize our trials as much as possible. “Targeted therapies,” by definition, inhibit a specific target, thus offering the theoretical advantage of enriching the patient population by restricting enrollment to patients whose tumor expresses the target of interest. Education of health care professionals regarding the benefits of adjuvant therapies, to decrease a sense of nihilism and to increase referrals, and international collaborations may also be necessary to increase accrual.
Localized Non–Small Cell Lung Cancer: Adjuvant Radiotherapy in the Era of Effective Systemic Therapy
Authors: Bogart Jeffrey A. , Aronowitz Jesse N.
Published: September 2005

Investigators in Europe, Canada, and the United States recently established a definitive role for adjuvant systemic chemotherapy following resection of early-stage non–small cell lung cancer (NSCLC). This was no small accomplishment, as upward of 20 randomized trials had previously been conducted. The role of postoperative radiotherapy (PORT) has been studied with far less vigor in the modern era. A 1998 meta-analysis of randomized trials suggesting that PORT was detrimental to survival included studies of doubtful quality. The value of PORT should be considered in the same context as recent chemotherapy trials. Advances in imaging have improved the accuracy of staging, patient selection, and target definition. Modern dosimetry and accelerator technologies have advanced the capacity to deliver radiation to the target with less tissue toxicity. Evolving philosophies in dosing and fractionation should improve the therapeutic ratio. Finally, it is reasonable to assume that the importance of local control will be enhanced in the setting of better systemic therapies. We will review the data on PORT and address critical issues in the design of trials to assess the role of modern radiotherapy in the integrated approach to management of early-stage NSCLC.
Clinical Issues in the Management of Early Lung Cancer
Authors: Mulshine James L.
Published: September 2005

Lung cancer is commonly diagnosed after metastatic spread, when therapies are rarely curative, providing an impetus for continued research directed at exploring approaches for cost-effective early lung cancer detection. Recently published pilot studies across three continents support a benefit of spiral computed tomography (CT) in detecting earlier stage non–small cell lung cancer. Improved resolution of early lung cancer is a result of significant changes in CT imaging hardware and software. The status and implications of these developments are reviewed. Many aspects of the management of screening for early lung cancer could be informed by optimizing the downstream clinical management of potential lung cancers identified by CT screening. The first and most critical issue is whether or not this improved detection rate is clearly associated with a reduction in lung cancer–related mortality. However, other related issues such as cost-benefit evaluations are also considered. If smaller, truly localized primary cancer can be routinely detected, then options for less morbid interventions would also be desirable. The rapid improvement in resolution and cost of spiral CT has provided a powerful impetus to reconsider the possibilities for achieving safe, economical, and meaningful early lung cancer detection.
Integration of Novel Therapeutics into Combined Modality Therapy of Locally Advanced Non–Small Cell Lung Cancer
Authors: Gandara David , Narayan Samir , Lara Primo N. , Goldberg Zelanna , Davies Angela , Lau Derrick H.M. , Mack Philip , Gumerlock Paul , Vijayakumar Srinivasan
Published: September 2005

Novel therapeutic agents (NTA) directed against a wide array of newly described molecular targets are now entering clinical investigation, many in the treatment of non–small cell lung cancer (NSCLC). The great majority of these clinical trials have been directed toward patients with advanced stage (metastatic) disease. More recently, study of NTAs has turned toward earlier-stage disease. Locally advanced, or stage III, NSCLC represents a large and heterogeneous group of patients and several clinically distinct substages. During the last 15 years, randomized clinical trials have shown improved survival with sequential chemoradiation compared with radiation alone and, more recently, the superiority of concurrent versus sequential chemoradiation. As NTAs have increasingly shown clinical activity against NSCLC, questions of how to incorporate them into clinical trials in stage III disease, whether they should be given together with radiotherapy, substituting for chemotherapy, or whether they should be added to current chemoradiation strategies, all remain as issues. Here, we describe conceptual issues, preclinical rationale, and ongoing or planned clinical trials incorporating NTAs into current treatment paradigms for unresectable stage III NSCLC.
Computed Tomography Screening for Lung Cancer: Principles and Results
Authors: Henschke Claudia I.
Published: September 2005

In modern healthcare, one of the most public, most important, and at the same time complicated and scientifically demanding topics is screening for a cancer. Mammographic screening for breast cancer has been, in the last few years, a particularly hotly disputed topic in scientific and public policy circles, with similar confusion and frustration widely reported and thus disseminated by the mass media. Most remarkably, this debacle has taken place against the backdrop of, and despite,a truly enormous amount of completed research designed to address the usefulness of such screening. To avoid a “mammography” debacle for lung cancer, the fundamental principles of screening need to be presented as we have come to think of them. Although it is generally accepted that low-dose computed tomography (CT) screening leads to early diagnosis of lung cancer in a high percentage of the cases, the logical consequence of the current knowledge is that annual CT screening prevents death from lung cancer. Thus, it is not whether CT screening is effective, rather the magnitude of its benefit which needs to be determined. We will illustrate our approach by discussing the Early Lung Cancer Action Project and its New York and International sequels.
Laparoscopic and Partial Nephrectomy
Authors: Novick Andrew C.
Published: September 2005

Radical nephrectomy is the gold standard curative operation for patients with localized renal cell carcinoma (RCC). Since its introduction in 1990, laparoscopic radical nephrectomy is being increasingly done at numerous institutions worldwide. In the hands of experienced laparoscopic urological surgeons and with adherence to established principles of open radical nephrectomy, laparoscopic radical nephrectomy is now a standard of care for patients with T1–3a N0 M0 RCC. Intermediate-term outcome data indicate equivalent cancer-free survival to open radical nephrectomy in such cases. Nephron-sparing surgery (NSS) is now an established approach for patients with localized RCC when there is a clinically relevant need to preserve renal function. NSS is also indicated in patients with a single, small, unilateral, localized RCC when the opposite kidney is completely normal. The technical success rate with NSS for RCC is excellent, and long-term patient survival free of cancer is comparable with that obtained after radical nephrectomy. We recently reviewed the results of NSS in 107 patients with localized sporadic RCC treated at the Cleveland Clinic before 1988 who were followed up for a minimum of 10 years. Long-term preservation of renal function was achieved in 93% of patients, and the 10-year cancer specific survival rate was 73%. Although open surgical partial nephrectomy remains the gold standard for nephron-sparing treatment of RCC, laparoscopic partial nephrectomy is now available in selected cases. The optimal indications for laparoscopic NSS are in patients with a relatively small and peripheral renal tumor. In such cases, laparoscopic NSS is proving to be an effective, minimally invasive therapeutic approach with respect to renal functional outcome, with additional advantages of reduced postoperative narcotic use, earlier hospital discharge, and a faster convalescence. The laparoscopic approach is associated with longer warm renal ischemia time, more major intraoperative complications, and more postoperative urological complications. Continued efforts are required to develop laparoscopic renal hypothermia techniques and to facilitate intrarenal suturing while minimizing the warm ischemia time.
Dendritic Cell-Tumor Fusion Vaccines for Renal Cell Carcinoma
Authors: Avigan David
Published: September 2005

Renal cell carcinoma is a malignant disease that demonstrates resistance to standard chemotherapeutic agents. A promising area of investigation is the use of cancer vaccines to educate host immunity to specifically target and eliminate malignant cells. Dendritic cells (DCs) are potent antigen-presenting cells that are uniquely effective in generating primary immune responses. DCs that are manipulated to present tumor antigens induce antitumor immunity in animal models and preclinical human studies. A myriad of strategies have been developed to effectively load tumor antigen onto DCs, including the introduction of individual peptides, proteins, or tumor-specific genes, as well as the use of whole tumor cells as a source of antigen. A promising approach for the design of cancer vaccines involves the fusion of whole tumor cells with DCs. The DC-tumor fusion presents a spectrum of tumor-associated antigens to helper and cytotoxic T-cell populations in the context of DC-mediated costimulatory signals. In animal models, vaccination with DC-tumor fusions resulted in protection from tumor challenge and regression of established metastatic disease. We have conducted phase 1 dose escalation studies in which patients with metastatic breast and renal cancer underwent vaccination with DC-tumor fusions. Twenty-three patients underwent vaccination with autologous DC-tumor fusions. Vaccination was well tolerated without substantial treatment-related toxic effects. Immunologic responses and disease regression were observed in a subset of patients. Future studies will explore the effect of DC maturation and cytokine adjuvants on vaccine potency.
Genetic Basis of Cancer of the Kidney
Authors: Linehan W. Marston , Vasselli James , Srinivasan Ramaprasad , Walther McClellan M. , Merino Maria , Choyke Peter , Vocke Cathy , Schmidt Laura , Isaacs Jennifer S. , Glenn Gladys , Toro Jorge , Zbar Berton , Bottaro Donald , Neckers Len
Published: September 2005

Studies during the past two decades have shown that kidney cancer is not a single disease; it is made up of a number of different types of cancer that occur in this organ. Clear cell renal carcinoma is characterized by mutation of the VHL gene. The VHL gene product forms a heterotrimeric complex with elongin C, elongin B, and Cul-2 to target hypoxia-inducible factors 1 and 2α for ubiquitin-mediated degradation. VHL−/− clear cell renal carcinoma overexpresses epidermal growth factor receptor and transforming growth factor α. Both hypoxia-inducible factor 1α and the epidermal growth factor receptor are potential therapeutic targets in clear cell renal carcinoma. Studies of the hereditary form of renal cell carcinoma (RCC) associated with hereditary papillary renal carcinoma (HPRC) determined that the c-Met proto-oncogene on chromosome 7 is the gene for HPRC and for a number of sporadic papillary RCCs. The HPRC c-Met mutations are activating mutations in the tyrosine kinase domain of the gene. The gene for a new form of hereditary RCC (Birt Hogg Dubé syndrome) associated with cutaneous tumors, lung cysts, and colon polyps or cancer has recently been identified. Studies are currently under way to determine what type of gene BHD is and how damage to this gene leads to kidney cancer. Individuals affected with hereditary leiomyomatosis renal cell carcinoma are at risk for the development of cutaneous leiomyomas, uterine leiomyomas (fibroids), and type 2 papillary RCC. The HLRC gene has been found to be the Krebs cycle enzyme, fumarate hydratase. Studies are under way to understand the downstream pathway of this cancer gene.
Comparative Preclinical Pharmacokinetic and Metabolic Studies of the Combretastatin Prodrugs Combretastatin A4 Phosphate and A1 Phosphate
Authors: Kirwan Ian G. , Loadman Paul M. , Swaine David J. , Anthoney D. Alan , Pettit George R. , Lippert John W. , Shnyder Steve D. , Cooper Patricia A. , Bibby Mike C.
Published: September 2005

Purpose: Combretastatin A4 phosphate (CA4P) and its structural analog, combretastatin A1 phosphate (CA1P), are soluble prodrugs capable of interacting with tubulin and causing rapid vascular shutdown within tumors. CA4P has completed Phase I clinical trials, but recent preclinical studies have shown that CA1P displays a greater antitumor effect than the combretastatin A4 (CA4) analog at equal doses. The aim of this study, therefore, is to compare pharmacokinetics and metabolism of the two compounds to determine whether pharmacokinetics plays a role in their differential activity.

Experimental Design: NMRI mice bearing MAC29 tumors received injection with either CA4P or CA1P at a therapeutic dose of 150 mg·kg−1, and profiles of both compounds and their metabolites analyzed by a sensitive and specific liquid chromatography/mass spectroscopy method.

Results: The metabolic profile of both compounds is complex, with up to 14 metabolites being detected for combretastatin A1 (CA1) in the plasma. Many of these metabolites have been identified by liquid chromatography/mass spectroscopy. Initial studies, however, focused on the active components CA4 and CA1, where plasma and tumor areas under the curve were 18.4 and 60.1 μg·h·ml−1 for CA4, and 10.4 and 13.1 μg·h·ml−1 for CA1, respectively. In vitro metabolic comparisons of the two compounds strongly suggest that CA1 is metabolized to a more reactive species than the CA4.

Conclusions: Although in vitro studies suggest that variable rates of tumor-specific prodrug dephosphorylation may explain these differences in pharmacokinetics profiles, the improved antitumor activity and altered pharmacokinetic profile of CA1 may be due to the formation of a more reactive metabolite.
Expression of Nuclear Factor-κB and IκBα Proteins in Prostatic Adenocarcinomas
Authors: Ross Jeffrey S. , Kallakury Bhaskar V. S. , Sheehan Christine E. , Fisher Hugh A. G. , Kaufman Ronald P. , Kaur Prabhjot , Gray Karen , Stringer Bradley
Published: September 2005

Purpose: The nuclear transcription factor nuclear factor-κB (NFκB) and its inhibitor, IκB, regulate the transcription of various genes involved in cell proliferation, adhesion, and survival. The NFκB transcription factor complex plays a role in cancer development and progression through its influence on apoptosis. More recently, NFκB has been shown to be activated in human and androgen-independent prostate cancer cells. To our knowledge, this is the first study demonstrating the prognostic significance of NFκB immunoreactivity in prostate adenocarcinomas (PACs).

Experimental Design: Using prostatectomy specimens, we performed immunohistochemical staining for NFκB and IκBα (Santa Cruz Biotechnology) on formalin-fixed, paraffin-embedded sections obtained from 136 patients with PAC. Cytoplasmic and nuclear immunoreactivity was scored for intensity and distribution, and results were correlated with preoperative serum prostate-specific antigen, tumor grade, stage, DNA ploidy (Feulgen spectroscopy), and biochemical disease recurrence.

Results: Forty-nine percent of PACs overexpressed cytoplasmic NFκB, and 63% showed decreased IκB expression. Cytoplasmic NFκB overexpression correlated with advanced tumor stage (P = 0.048), aneuploidy (P = 0.022), and biochemical disease recurrence (P = 0.001). When we compared the means for the NFκB-positive and -negative subgroups, NFκB overexpression correlated with preoperative serum prostate-specific antigen (P = 0.04) and DNA index (P = 0.05). Fifteen percent of PACs expressed nuclear NFκB, which correlated with high tumor grade (P = 0.001) and advanced stage (P = 0.05). Decreased IκBα expression correlated with high tumor grade (P = 0.015). On multivariate analysis, tumor stage (P = 0.043) and NFκB overexpression (P = 0.006) were independent predictors of biochemical recurrence.

Conclusion: These results support a role for NFκB pathway proteins in the tumorigenesis of PACs. The findings are also consistent with reported experimental studies suggesting a new strategy of combined chemotherapy and specific NFκB blockade in decreasing the rate of disease relapse.
M34 Actin Regulatory Protein Is a Sensitive Diagnostic Marker for Early- and Late-Stage Mammary Carcinomas
Authors: Soriano Zoraida , Pardee Joel D.
Published: September 2005

Purpose: At present, there is no available molecular marker that reliably detects the earliest stages of epithelial transformation in the majority of patients affected with incipient breast carcinoma. Here we introduce M34 protein, a mammalian actin filament regulatory protein, as a highly sensitive and easily detected positive cellular marker for both early and late stages of breast carcinoma.

Experimental Design: In this study, 24 human lactation duct neoplasms from postmenopausal women, including fibroadenoma, ductal carcinoma in situ, intraductal lobular papilloma, and metastatic adenocarcinoma, were analyzed for the presence of M34 protein by histochemical staining of paraffin and fresh-frozen sections.

Results: All 24 neoplasias tested positive for M34, whereas none of the 4 normal breast tissues stained for the protein. M34 identification was strongly positive for transformed epithelium in all tumor types tested. Twelve precancerous lesions of fibroadenoma (n = 4), intraductal papilloma (n = 4), and incipient ductal carcinoma in situ (n = 4) all showed high levels of M34 staining, suggesting that precancerous tumors, as well as the earliest stages of mammary carcinoma, can be sensitively detected. Furthermore, anti-M34 antibody selectively stained all 12 advanced-stage metastatic adenocarcinoma cell masses and micrometastases in axillary lymph nodes tested. Single-cell micrometastases embedded in connective tissue or lymph node parenchyma could be clearly resolved by M34 with horseradish peroxidase staining. Lymphocytes, normal ductal endothelium, and vascular endothelial cells were M34-negative, as were muscle, nerve, and adipose tissues. Low-level M34 staining was detected in connective tissue fibroblasts, macrophages, and neutrophils.

Conclusions: To our knowledge, no previously reported markers have shown high sensitivity of detection for both the earliest and most advanced stages of breast carcinoma. Consequently, M34 appears uniquely suited for diagnosis of the earliest stages of lactation duct transformation as well as for advanced-stage mammary carcinoma metastases in surgical margins and axillary lymph nodes.
The Metabolism of Pyrazoloacridine (NSC 366140) by Cytochromes P450 and Flavin Monooxygenase in Human Liver Microsomes
Authors: Reid Joel M. , Walker Denise L. , Miller Jennifer K. , Benson Linda M. , Tomlinson Andrew J. , Naylor Stephen , Blajeski April L. , LoRusso Patricia M. , Ames Matthew M.
Published: September 2005

Pyrazoloacridine (PZA) is an experimental antitumor agent presently under investigation for treatment of solid tumors on the basis of its unique mechanism of action and selectivity for human solid tumor xenograft in mice. Using capillary electrophoresis coupled with electrospray ionization mass spectrometry, we have identified three oxidative PZA metabolites, 9-desmethyl-PZA, N-demethyl-PZA, and PZA N-oxide. The cytochrome P450 (CYP) isoforms involved in PZA metabolism were characterized by studies with CYP chemical inhibitors, correlation of marker activities for selected CYPs with formation of the metabolites using a human liver panel, and PZA metabolism by cDNA-expressed CYPs. 9-Desmethyl-PZA formation was catalyzed by CYP1A2, whereas N-demethyl-PZA formation was catalyzed by CYP3A4. PZA N-oxide formation was catalyzed by flavin monooxygenase (FMO) rather than CYP, as determined by studies with chemical inhibitors of FMO and metabolism by cDNA-expressed human flavin monooxygenase. After administration of [10b-14C]PZA to mice, six urinary metabolites were detected by high-performance liquid chromatography UV and radiochromatograms including 9-desmethyl-PZA, N-demethyl-PZA, and PZA N-oxide. Trace concentrations of 9-desmethyl-PZA and PZA N-oxide were detected in mouse plasma. PZA N-oxide and N-demethyl-PZA were detected in urine from patients after PZA administration. PZA, 9-desmethyl-PZA, and PZA N-oxide inhibited growth of A375 human melanoma cells. IC50 values were 0.17, 0.11, and 7.0 μm, respectively, for the three molecules.
Microsatellite Instability Predicts Poor Short-Term Survival in Patients with Advanced Breast Cancer after High-Dose Chemotherapy and Autologous Stem-Cell Transplantation
Authors: Wild Peter J. , Reichle Albrecht , Andreesen Reinhard , Röckelein Georg , Dietmaier Wolfgang , Rüschoff Josef , Blaszyk Hagen , Hofstädter Ferdinand , Hartmann Arndt
Published: September 2005

Purpose: The purpose is to define molecular prognostic factors in patients with advanced breast cancer treated with high-dose chemotherapy (HDCT) and autologous stem cell transplantation (ASCT).

Experimental Design: Thirty-nine patients with breast cancer and extensive lymph node (level III) and/or systemic metastases from a prospective single-center study of sequential HDCT/ASCT were studied. Microsatellite analysis was performed after laser microdissection using 15 markers selected for sensitive detection of microsatellite instability (MSI) in breast cancer. Exons 5–9 of the P53 gene were directly sequenced. Expression of P53, HER-2/neu, and the mismatch repair proteins hMSH2 and hMLH1 was evaluated by immunohistochemistry.

Results: MSI of at least three markers was detected in 13 of 39 patients (33%) and was predominantly found at tetranucleotide markers. All MSI-positive tumors showed normal expression of hMSH2 and hMLH1. Complete sequence analysis of exons 5–9 of the P53gene was successful in 34 cases; 18% (n = 6) revealed a mutation. Overexpression of HER-2/neu and P53 was observed in 7 (22%) and 12 (46%) of 26 evaluated cases, respectively. The presence of MSI strongly correlated with shorter overall survival (OS; P = 0.0004) and progression-free survival (PFS; P = 0.02). None of the other investigated clinical or molecular factors correlated with OS in univariate analyses, with the exception of menopausal status and previous adjuvant chemotherapy. Testing various multivariate Cox regression models, MSI remained a highly significant, independent, and adverse risk factor for OS.

Conclusions: MSI is frequent in advanced breast cancer and could be an indicator of chemotherapy resistance and poor prognosis in breast cancer patients treated with HDCT/ASCT.
Conditionally Replicative Adenovirus with Tropism Expanded towards Integrins Inhibits Osteosarcoma Tumor Growth
in Vitro
and
in Vivo
Authors: Witlox Adhiambo M. , van Beusechem Victor W. , Molenaar Bonnie , Bras Hans , Schaap Gerard R. , Alemany Ramon , Curiel David T. , Pinedo Herbert M. , Wuisman Paul I. J. M. , Gerritsen Winald R.
Published: September 2005

Purpose: The clinical course of osteosarcoma (OS) demands the development of new therapeutic options. Conditionally replicative adenoviruses (CRAds) represent promising agents for the treatment of solid tumors, because CRAds have an intrinsic replication capacity that allows in situ amplification and extensive tumor infection through lateral spread. The CRAd AdΔ24 has been developed to replicate selectively in cells with a defective retinoblastoma (Rb) pathway. Because genetic alterations in the Rb pathway are frequently observed in OS, AdΔ24 might be useful in the treatment of this cancer.

Experimental Design: Because the lack of Coxsackie adenovirus receptor on OS cells limits the efficacy of CRAd treatment, we explored alternative target molecules on OS. Insertion of an Arg-Gly-Asp (RGD-4C) integrin-targeting motif into the adenovirus fiber knob expanded tropism toward the ανβ3 and ανβ5 integrins. The oncolytic capacity of the CRAd Ad5-Δ24RGD was tested on primary OS cells in vitro and in vivo.

Results: The ανβ3and ανβ5 integrins are abundantly expressed on OS cells. RGD-mediated infection augmented adenovirus infection of primary OS cells by two orders of magnitude. Ad5-Δ24RGD was shown to be highly active in killing human OS cell lines, as well as primary cell cultures. Furthermore, intratumoral injections with Ad5-Δ24RGD into established human OS xenografts that were derived directly from a patient with OS refractory for chemotherapeutic treatment caused a significant tumor-growth delay. Furthermore, adenoviral particles could be detected in tumor cells 25 days posttumor injection.

Conclusions: Targeting adenovirus toward integrins rendered OS cells more sensitive to killing by Ad5-Δ24RGD. These findings suggest that Ad5-Δ24RGD has potential for use in OS treatment.
Quantitative Detection of Circulating Tumor Cells in Cutaneous and Ocular Melanoma and Quality Assessment by Real-Time Reverse Transcriptase-Polymerase Chain Reaction
Authors: Keilholz Ulrich , Goldin-Lang Petra , Bechrakis Nikolaos E. , Max Nicole , Letsch Anne , Schmittel Alexander , Scheibenbogen Carmen , Heufelder Karin , Eggermont Alexander , Thiel Eckhard
Published: September 2005

Purpose: Inconsistent reports on the detection of melanoma cells in peripheral blood by reverse transcriptase-PCR (RT-PCR) have resulted in uncertainty on the prognostic value of circulating melanoma cells.

Experimental Design: We developed real-time RT-PCR assays for quantitation of tyrosinase, MelanA/MART1, and gp100 and for porphobilinogen deaminase housekeeping gene. Melanoma tissue (n = 18), peripheral blood samples from healthy donors (n = 21), and patients with cutaneous (n = 122) and uveal (n = 64) melanoma from our institution were analyzed. For quality control, an additional 251 samples from ongoing multicenter studies were compared with in-house samples.

Results: Tyrosinase was not detected in healthy donor blood samples. For the two other markers, cutoff values had to be defined to distinct patient samples from controls. Patients with stage IV uveal and cutaneous melanoma expressed all three markers more frequently and at higher levels in peripheral blood as compared with earlier stages. The variation of expression was 4 logs and correlated with tumor load and serum lactate dehydrogenase. In 2 of 3 uveal melanoma patients, detection of circulating tumor cells preceded the development of liver metastases. The diagnostic sensitivity was optimal in blood samples containing >0.1pg/μl porphobilinogen deaminase (95.7% of in-house samples and 57.4% of multicenter samples).

Conclusions: Real-time RT-PCR is able to quantitatively define the quality of a sample and provides quantitative data for melanoma markers. Disparities in the results of previous studies may be attributable to undetected differences in sample quality. The prognostic relevance of this assay is currently under evaluation in several prospective randomized trials.
Concomitant Overexpression of Cyclooxygenase-2 in HER-2–Positive on Smad4-Reduced Human Gastric Carcinomas Is Associated with a Poor Patient Outcome
Authors: Okano Hirokazu , Shinohara Hisashi , Miyamoto Akiko , Takaori Kyoichi , Tanigawa Nobuhiko
Published: September 2005

Purpose: The expression of cyclooxygenase-2 (COX-2) is known to be involved in gastric carcinogenesis and tumor progression, but little is known about the mechanisms responsible for the up-regulation of COX-2. We examined the involvement of two growth factor-signaling systems, HER-2 and transforming growth factor (TGF)-β, in the induction of COX-2 in human gastric cancer tissue.

Experimental Design: COX-2 expression was detected by immunohistochemistry in surgical specimens obtained from 166 patients with advanced gastric cancer; possible correlations between the expression of COX-2 and the expression of HER-2, TGF-β1, and Smad4, an intracellular mediator that transmits the TGF-β signal, were then analyzed.

Results: COX-2 protein was overexpressed in 91 (54.8%) tumors; COX-2 overexpression was correlated with a differentiated histologic type, deep invasion, and positive lymph node metastasis. COX-2 was frequently overexpressed in HER-2–positive tumors (19 of 22, 86.4%) and in Smad4-reduced tumors (67 of 104, 64.4%) but irrelevant to the TGF-β1 expression status. The expression levels of COX-2 and HER-2 and the reduction in Smad4 were all associated with a poor patient outcome. A multivariate analysis demonstrated a significantly poor outcome for the concomitant overexpression of COX-2 in patients with Smad4-reduced tumors.

Conclusions: These results support the possibility that signal transduction via HER-2 and the TGF-β/Smad system may be implicated in COX-2 expression and that the reduction of Smad4 may be, in part, of causal significance in the TGF-β-initiated overexpression of COX-2, which is associated with a poor prognosis for patients with gastric cancer.
Phenylbutyrate and Phenylacetate Induce Differentiation and Inhibit Proliferation of Human Medulloblastoma Cells
Authors: Li Xiao-Nan , Parikh Suhag , Shu Qin , Jung Hye-Lim , Chow Chi-Wan , Perlaky Laszlo , Leung Hon-Chiu Eastwood , Su Jack , Blaney Susan , Lau Ching C.
Published: September 2005

Purpose: Phenylbutyrate (PB) and phenylacetate (PA) have antiproliferative and differentiation-inducing effects in malignant tumors, and had been evaluated in Phase I/II clinical trials. This study was undertaken to evaluate their antitumor activities in medulloblastomas.

Experimental Design: The biological effects of PB and PA, ranging from 0.1 mm to 3 mm, on two medulloblastoma cell lines (DAOY and D283-MED) were examined using various long-term in vitro and in vivo assays for morphology, proliferation, differentiation, anchorage-independent growth, apoptosis, and tumorigenicity.

Results: PB and PA can both induce morphological changes and suppress proliferation in a time- and dose-dependent manner. These effects were more pronounced with PB and became irreversible in D283-MED cells after continuous exposure to 3 mm PB for 28 days. Both PB and PA were able to increase expression of glial marker glial fibriliary acidic protein and neuronal marker synaptophysin in two cell lines. For anchorage-independent growth, PB showed a more significant suppression than PA in D283-MED cells. PB caused more pronounced cell cycle arrest and remarkably reduced tumorigenicity in D283-MED cells than in DAOY cells. Apoptosis was readily induced in D283-MED cells with either low dose of PB or short-term treatment. In contrast, much higher concentrations of PB or longer treatment were required to achieve similar effect with DAOY cells. PB induced increased histones H3 acetylation in both cell lines, but histone H4 acetylation was only observed in D283-MED cells.

Conclusions: PB, through induction of hyperacetylation of histone H3 and H4, is a much more potent antitumor agent than PA. 283-MED cells are more responsive to PB than DAOY cells, which may be dependent on their original state of differentiation as well as the changes of histone H4 acetylation status.