Publications by authors named "Anna Maria Calcagno"

25 Publications

  • Page 1 of 1

An Analysis of Recent FDA Oncology Scientific Publications.

Oncologist 2020 03 26;25(3):266-270. Epub 2019 Nov 26.

Oncology Center of Excellence, U.S. Food and Drug Administration, Silver Spring, Maryland, USA.

In addition to its primary regulatory role, the Office of Hematology and Oncology Products at the U.S. Food and Drug Administration (FDA) is engaged in many forms of scientific authorship. During the period of 2010 to 2018, FDA oncology staff contributed to 356 publications in the scientific literature. Here, we collaborated with analysts in the Office of Program Planning, Analysis, and Evaluation at the National Institute of General Medical Sciences, National Institutes of Health (NIH), to present a series of analyses aimed at quantifying the characteristics and potential impact of these contributions, as well as characterizing the areas of work addressed. We found that FDA oncology papers are enriched for high-impact publications and have about two times the number of citations as an average NIH-funded paper. Further impact of the publications was measured based on the presence of 65 publications that were cited by guidelines and 12 publications cited by publicly listed clinical trials. The results seen here are promising in determining the impact of FDA oncology publication work but prompt further investigation into longer-term impacts, such as the influence of this work on other regulatory activities at FDA. IMPLICATIONS FOR PRACTICE: This article describes the first comprehensive study of scientific publications produced by U.S. Food and Drug Administration (FDA) oncology staff. The analysis illustrates that staff are highly engaged in publishing in the scientific literature in addition to completing regulatory review work. Publications are generally in clinical medicine, consistent with the large number of medical oncologists working at the Office of Hematology and Oncology Products (OHOP). OHOP publications generally focus either on communicating important regulatory work (approval summaries) or highlighting regulatory science issues to encourage dialogue with the scientific community (commentaries, reviews, and expert working papers). The analysis also suggests that several FDA oncology publications may influence clinical guidelines, but further work is needed to evaluate impact.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1634/theoncologist.2019-0503DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7066718PMC
March 2020

An Analysis of Recent FDA Oncology Scientific Publications.

Oncologist 2019 Nov 26. Epub 2019 Nov 26.

Oncology Center of Excellence, U.S. Food and Drug Administration, Silver Spring, Maryland, USA.

In addition to its primary regulatory role, the Office of Hematology and Oncology Products at the U.S. Food and Drug Administration (FDA) is engaged in many forms of scientific authorship. During the period of 2010 to 2018, FDA oncology staff contributed to 356 publications in the scientific literature. Here, we collaborated with analysts in the Office of Program Planning, Analysis, and Evaluation at the National Institute of General Medical Sciences, National Institutes of Health (NIH), to present a series of analyses aimed at quantifying the characteristics and potential impact of these contributions, as well as characterizing the areas of work addressed. We found that FDA oncology papers are enriched for high-impact publications and have about two times the number of citations as an average NIH-funded paper. Further impact of the publications was measured based on the presence of 65 publications that were cited by guidelines and 12 publications cited by publicly listed clinical trials. The results seen here are promising in determining the impact of FDA oncology publication work but prompt further investigation into longer-term impacts, such as the influence of this work on other regulatory activities at FDA. IMPLICATIONS FOR PRACTICE: This article describes the first comprehensive study of scientific publications produced by U.S. Food and Drug Administration (FDA) oncology staff. The analysis illustrates that staff are highly engaged in publishing in the scientific literature in addition to completing regulatory review work. Publications are generally in clinical medicine, consistent with the large number of medical oncologists working at the Office of Hematology and Oncology Products (OHOP). OHOP publications generally focus either on communicating important regulatory work (approval summaries) or highlighting regulatory science issues to encourage dialogue with the scientific community (commentaries, reviews, and expert working papers). The analysis also suggests that several FDA oncology publications may influence clinical guidelines, but further work is needed to evaluate impact.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1634/theoncologist.2019-0503DOI Listing
November 2019

Using database linkages to measure innovation, commercialization, and survival of small businesses.

Eval Program Plann 2019 12 3;77:101710. Epub 2019 Sep 3.

National Institute of General Medical Sciences, National Institutes of Health, MSC 6200, 45 Center Drive, Bethesda MD 20892-6200, USA. Electronic address:

Here, we report the results of an outcomes evaluation of the Small Business Innovation Research (SBIR) and Small Business Technology Transfer (STTR) Programs at the National Institute of General Medical Sciences (NIGMS). Since the programs' inception, assessments of the SBIR/STTR programs at several federal agencies have utilized surveys of former grantees as the primary source of data. Response rates have typically been low, making non-response bias a potential threat to the validity of some of these studies' results. Meanwhile, the availability of large publicly-available datasets continues to grow and methods of text mining and linking databases continue to improve. By linking NIGMS grant funding records, U.S. Patent and Trademark Office data, and business intelligence databases, we explored innovation, commercialization and survival for recipients of NIGMS SBIR/STTR funding. In doing so, we were able to more completely assess several key outcomes of the NIGMS SBIR/STTR program. Our evaluation demonstrated that the NIGMS program performed above baseline expectations along all dimensions, and comparably to other federal agency SBIR/STTR grant programs. In addition, we show that the use of extant data increasingly is a viable, less expensive, and more reliable approach to gathering data for evaluation studies.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.evalprogplan.2019.101710DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6815254PMC
December 2019

Geographically-related outcomes of U.S. funding for small business research and development: Results of the research grant programs of a component of the National Institutes of Health.

Eval Program Plann 2019 12 31;77:101696. Epub 2019 Jul 31.

National Institute of General Medical Sciences, National Institutes of Health, Bethesda, MD, 20892, United States. Electronic address:

This article examines the geographic distribution of funding for the U.S. Small Business Innovation Research (SBIR) and Small Business Technology Transfer (STTR) programs sponsored by the National Institute of General Medical Sciences (NIGMS). Despite a significant investment in SBIR/STTR and an interest in increasing geographic diversity in the institute's research portfolio, there has not been an assessment of the distribution of NIGMS's SBIR/STTR funding, outcomes associated with that investment, and relationships between the two. The geographic distribution of NIGMS' SBIR/STTR funding was highly concentrated in a small number of states, with a high correlation between each state's funding and its number of small scientific research and development businesses. Affiliation with a major research university was correlated with several measures of innovation and firm success. Our findings are consistent with earlier research showing that economic activity in research and development and research output tend to cluster in geographic regions where knowledge can be generated and shared more efficiently. These findings lend support to an investment strategy for small business research and development that creates networks between major research universities and small businesses.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.evalprogplan.2019.101696DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6815264PMC
December 2019

NIH funding longevity by gender.

Proc Natl Acad Sci U S A 2018 07 16;115(31):7943-7948. Epub 2018 Jul 16.

National Institute of General Medical Sciences, National Institutes of Health, Bethesda, MD 20892

Women have achieved parity with men among biomedical science degree holders but remain underrepresented in academic positions. The National Institutes of Health (NIH)-the world's largest public funder of biomedical research-receives less than one-third of its new grant applications from women. Correspondingly, women compose less than one-third of NIH research grantees, even though they are as successful as men in obtaining first-time grants. Our study examined women's and men's NIH funding trajectories over time ( = 34,770), exploring whether women remain funded at the same rate as men after receiving their first major research grants. A survival analysis demonstrated a slightly lower funding longevity for women. We next examined gender differences in application, review, and funding outcomes. Women individually held fewer grants, submitted fewer applications, and were less successful in renewing grants-factors that could lead to gender differences in funding longevity. Finally, two adjusted survival models that account for initial investigator characteristics or subsequent application behavior showed no gender differences, suggesting that the small observed longevity differences are affected by both sets of factors. Overall, given men's and women's generally comparable funding longevities, the data contradict the common assumption that women experience accelerated attrition compared with men across all career stages. Women's likelihood of sustaining NIH funding may be better than commonly perceived. This suggests a need to explore women's underrepresentation among initial NIH grantees, as well as their lower rates of new and renewal application submissions.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1073/pnas.1800615115DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6077749PMC
July 2018

A Gene Expression Signature Associated with Overall Survival in Patients with Hepatocellular Carcinoma Suggests a New Treatment Strategy.

Mol Pharmacol 2016 Feb 14;89(2):263-72. Epub 2015 Dec 14.

Laboratory of Cell Biology (J-P.G., J.P.M., C-P.W., A.M.C., S.V.A., M.M.G.) and Laboratory of Experimental Carcinogenesis (J.B.A., S.S.T.), Center for Cancer Research, National Cancer Institute, and Bioinformatics and Computational Biosciences Branch, Office of Cyber Infrastructure and Computational Biology, Office of Science Management and Operations, National Institute of Allergy and Infectious Diseases (S.V.), National Institutes of Health, Bethesda, Maryland; and the Viral Technologies Group and Molecular Detection Group, Protein Expression Laboratory, Frederick National Laboratory for Cancer Research, National Institutes of Health, Frederick, Marylanld (R.K.B., K.P., W.E.B.)

Despite improvements in the management of liver cancer, the survival rate for patients with hepatocellular carcinoma (HCC) remains dismal. The survival benefit of systemic chemotherapy for the treatment of liver cancer is only marginal. Although the reasons for treatment failure are multifactorial, intrinsic resistance to chemotherapy plays a primary role. Here, we analyzed the expression of 377 multidrug resistance (MDR)-associated genes in two independent cohorts of patients with advanced HCC, with the aim of finding ways to improve survival in this poor-prognosis cancer. Taqman-based quantitative polymerase chain reaction revealed a 45-gene signature that predicts overall survival (OS) in patients with HCC. Using the Connectivity Map Tool, we were able to identify drugs that converted the gene expression profiles of HCC cell lines from ones matching patients with poor OS to profiles associated with good OS. We found three compounds that convert the gene expression profiles of three HCC cell lines to gene expression profiles associated with good OS. These compounds increase histone acetylation, which correlates with the synergistic sensitization of those MDR tumor cells to conventional chemotherapeutic agents, including cisplatin, sorafenib, and 5-fluorouracil. Our results indicate that it is possible to modulate gene expression profiles in HCC cell lines to those associated with better outcome. This approach also increases sensitization of HCC cells toward conventional chemotherapeutic agents. This work suggests new treatment strategies for a disease for which few therapeutic options exist.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1124/mol.115.101360DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4727122PMC
February 2016

Multidrug resistance-linked gene signature predicts overall survival of patients with primary ovarian serous carcinoma.

Clin Cancer Res 2012 Jun 5;18(11):3197-206. Epub 2012 Apr 5.

Laboratory of Cell Biology, Center for Cancer Research, National Cancer Institute, Bethesda, MD 20892, USA.

Purpose: This study assesses the ability of multidrug resistance (MDR)-associated gene expression patterns to predict survival in patients with newly diagnosed carcinoma of the ovary. The scope of this research differs substantially from that of previous reports, as a very large set of genes was evaluated whose expression has been shown to affect response to chemotherapy.

Experimental Design: We applied a customized TaqMan low density array, a highly sensitive and specific assay, to study the expression profiles of 380 MDR-linked genes in 80 tumor specimens collected at initial surgery to debulk primary serous carcinoma. The RNA expression profiles of these drug resistance genes were correlated with clinical outcomes.

Results: Leave-one-out cross-validation was used to estimate the ability of MDR gene expression to predict survival. Although gene expression alone does not predict overall survival (OS; P = 0.06), four covariates (age, stage, CA125 level, and surgical debulking) do (P = 0.03). When gene expression was added to the covariates, we found an 11-gene signature that provides a major improvement in OS prediction (log-rank statistic P < 0.003). The predictive power of this 11-gene signature was confirmed by dividing high- and low-risk patient groups, as defined by their clinical covariates, into four specific risk groups on the basis of expression levels.

Conclusion: This study reveals an 11-gene signature that allows a more precise prognosis for patients with serous cancer of the ovary treated with carboplatin- and paclitaxel-based therapy. These 11 new targets offer opportunities for new therapies to improve clinical outcome in ovarian cancer.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1158/1078-0432.CCR-12-0056DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3376649PMC
June 2012

Redefining the relevance of established cancer cell lines to the study of mechanisms of clinical anti-cancer drug resistance.

Proc Natl Acad Sci U S A 2011 Nov 8;108(46):18708-13. Epub 2011 Nov 8.

Laboratory of Cell Biology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892, USA.

Although in vitro models have been a cornerstone of anti-cancer drug development, their direct applicability to clinical cancer research has been uncertain. Using a state-of-the-art Taqman-based quantitative RT-PCR assay, we investigated the multidrug resistance (MDR) transcriptome of six cancer types, in established cancer cell lines (grown in monolayer, 3D scaffold, or in xenograft) and clinical samples, either containing >75% tumor cells or microdissected. The MDR transcriptome was determined a priori based on an extensive curation of the literature published during the last three decades, which led to the enumeration of 380 genes. No correlation was found between clinical samples and established cancer cell lines. As expected, we found up-regulation of genes that would facilitate survival across all cultured cancer cell lines evaluated. More troubling, however, were data showing that all of the cell lines, grown either in vitro or in vivo, bear more resemblance to each other, regardless of the tissue of origin, than to the clinical samples they are supposed to model. Although cultured cells can be used to study many aspects of cancer biology and response of cells to drugs, this study emphasizes the necessity for new in vitro cancer models and the use of primary tumor models in which gene expression can be manipulated and small molecules tested in a setting that more closely mimics the in vivo cancer microenvironment so as to avoid radical changes in gene expression profiles brought on by extended periods of cell culture.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1073/pnas.1111840108DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3219108PMC
November 2011

Clinical relevance of multidrug resistance gene expression in ovarian serous carcinoma effusions.

Mol Pharm 2011 Dec 15;8(6):2080-8. Epub 2011 Jul 15.

Laboratory of Cell Biology, Center for Cancer Research, National Cancer Institute, NIH, Bethesda, Maryland 20892, USA.

The presence of tumor cells in effusions within serosal cavities is a clinical manifestation of advanced-stage cancer and is generally associated with poor survival. Identifying molecular targets may help to design efficient treatments to eradicate these aggressive cancer cells and improve patient survival. Using a state-of-the-art TaqMan-based qRT-PCR assay, we investigated the multidrug resistance (MDR) transcriptome of 32 unpaired ovarian serous carcinoma effusion samples obtained at diagnosis or at disease recurrence following chemotherapy. MDR genes were selected a priori based on an extensive curation of the literature published during the last three decades. We found three gene signatures with a statistically significant correlation with overall survival (OS), response to treatment [complete response (CR) vs other], and progression free survival (PFS). The median log-rank p-values for the signatures were 0.023, 0.034, and 0.008, respectively. No correlation was found with residual tumor status after cytoreductive surgery, treatment (with or without chemotherapy) and stage defined according to the International Federation of Gynecology and Obstetrics. Further analyses demonstrated that gene expression alone can effectively predict the survival outcome of women with ovarian serous carcinoma (OS, log-rank p = 0.0000; and PFS, log-rank p = 0.002). Interestingly, the signature for overall survival is the same in patients at first presentation and those who had chemotherapy and relapsed. This pilot study highlights two new gene signatures that may help in optimizing the treatment for ovarian carcinoma patients with effusions.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1021/mp200240aDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3224865PMC
December 2011

Prolonged drug selection of breast cancer cells and enrichment of cancer stem cell characteristics.

J Natl Cancer Inst 2010 Nov 8;102(21):1637-52. Epub 2010 Oct 8.

Laboratory of Cell Biology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892, USA.

Background: Cancer stem cells are presumed to have virtually unlimited proliferative and self-renewal abilities and to be highly resistant to chemotherapy, a feature that is associated with overexpression of ATP-binding cassette transporters. We investigated whether prolonged continuous selection of cells for drug resistance enriches cultures for cancer stem-like cells.

Methods: Cancer stem cells were defined as CD44+/CD24⁻ cells that could self-renew (ie, generate cells with the tumorigenic CD44+/CD24⁻ phenotype), differentiate, invade, and form tumors in vivo. We used doxorubicin-selected MCF-7/ADR cells, weakly tumorigenic parental MCF-7 cells, and MCF-7/MDR, an MCF-7 subline with forced expression of ABCB1 protein. Cells were examined for cell surface markers and side-population fractions by microarray and flow cytometry, with in vitro invasion assays, and for ability to form mammospheres. Xenograft tumors were generated in mice to examine tumorigenicity (n = 52). The mRNA expression of multidrug resistance genes was examined in putative cancer stem cells and pathway analysis of statistically significantly differentially expressed genes was performed. All statistical tests were two-sided.

Results: Pathway analysis showed that MCF-7/ADR cells express mRNAs from ABCB1 and other genes also found in breast cancer stem cells (eg, CD44, TGFB1, and SNAI1). MCF-7/ADR cells were highly invasive, formed mammospheres, and were tumorigenic in mice. In contrast to parental MCF-7 cells, more than 30% of MCF-7/ADR cells had a CD44+/CD24⁻ phenotype, could self-renew, and differentiate (ie, produce CD44+/CD24⁻ and CD44+/CD24+ cells) and overexpressed various multidrug resistance-linked genes (including ABCB1, CCNE1, and MMP9). MCF-7/ADR cells were statistically significantly more invasive in Matrigel than parental MCF-7 cells (MCF-7 cells = 0.82 cell per field and MCF-7/ADR = 7.51 cells per field, difference = 6.69 cells per field, 95% confidence interval = 4.82 to 8.55 cells per field, P < .001). No enrichment in the CD44+/CD24⁻ or CD133+ population was detected in MCF-7/MDR.

Conclusion: The cell population with cancer stem cell characteristics increased after prolonged continuous selection for doxorubicin resistance.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1093/jnci/djq361DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2970576PMC
November 2010

Analysis of expression of drug resistance-linked ABC transporters in cancer cells by quantitative RT-PCR.

Methods Mol Biol 2010 ;637:121-32

Laboratory of Cell Biology, Center for Cancer Research, National Cancer Institute, NIH, DHHS, Bethesda, MD, USA.

Quantitative real-time PCR (qRT-PCR) boasts many advantages over microarrays. For instance, very low amounts of total RNA are required to yield highly accurate and reproducible detection of transcript levels. As a consequence, qRT-PCR has become a popular technique for assessing gene expression levels and is now the gold standard. In this chapter, qRT-PCR using two distinct chemistries, SYBR Green and TaqMan, are described. We compare ABC transporter levels in various drug-resistant cancer cell lines by employing each method. SYBR Green yields reproducible results; nevertheless, TaqMan chemistry is superior to SYBR Green, as it displays higher specificity and sensitivity. Gene expression analysis by qRT-PCR is a powerful technique and shows potential as a diagnostic tool for predicting drug response in cancer patients.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1007/978-1-60761-700-6_6DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3108025PMC
July 2010

Molecular mechanisms of drug resistance in single-step and multi-step drug-selected cancer cells.

Methods Mol Biol 2010 ;596:77-93

Laboratory of Cell Biology, Center for Cancer Research, National Cancer Institute, NIH, DHHS, Bethesda, MD, USA.

Multidrug resistance (MDR) remains one of the key determinants in chemotherapeutic success of cancer patients. Often, acquired resistance is mediated by the overexpression of ATP-binding cassette (ABC) drug transporters. To study the mechanisms involved in the MDR phenotype, investigators have generated a variety of in vitro cell culture models using both multi-step and single-step drug selections. Sublines produced from multi-step selections have led to the discovery of several crucial drug transporters including ABCB1, ABCC1, and ABCG2. Additionally, a number of mechanisms causing gene overexpression have been elucidated. To more closely mimic in vivo conditions, investigators have also established MDR sublines with single-step drug selections. Here, we examine some of the multi-step and single-step selected cell lines generated to elucidate the mechanisms involved in the development of MDR in cancer cells.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1007/978-1-60761-416-6_5DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3099236PMC
January 2010

Evaluation of current methods used to analyze the expression profiles of ATP-binding cassette transporters yields an improved drug-discovery database.

Mol Cancer Ther 2009 Jul 7;8(7):2057-66. Epub 2009 Jul 7.

Laboratory of Cell Biology, Center for Cancer Research, National Cancer Institute, NIH, Bethesda, MD 20892-4256, USA.

The development of multidrug resistance (MDR) to chemotherapy remains a major challenge in the treatment of cancer. Resistance exists against every effective anticancer drug and can develop by multiple mechanisms. These mechanisms can act individually or synergistically, leading to MDR, in which the cell becomes resistant to a variety of structurally and mechanistically unrelated drugs in addition to the drug initially administered. Although extensive work has been done to characterize MDR mechanisms in vitro, the translation of this knowledge to the clinic has not been successful. Therefore, identifying genes and mechanisms critical to the development of MDR in vivo and establishing a reliable method for analyzing highly homologous genes from small amounts of tissue is fundamental to achieving any significant enhancement in our understanding of MDR mechanisms and could lead to treatments designed to circumvent it. In this study, we use a previously established database that allows the identification of lead compounds in the early stages of drug discovery that are not ATP-binding cassette (ABC) transporter substrates. We believe this can serve as a model for appraising the accuracy and sensitivity of current methods used to analyze the expression profiles of ABC transporters. We found two platforms to be superior methods for the analysis of expression profiles of highly homologous gene superfamilies. This study also led to an improved database by revealing previously unidentified substrates for ABCB1, ABCC1, and ABCG2, transporters that contribute to MDR.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1158/1535-7163.MCT-09-0256DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2736804PMC
July 2009

Reversal of ABC drug transporter-mediated multidrug resistance in cancer cells: evaluation of current strategies.

Curr Mol Pharmacol 2008 Jun;1(2):93-105

Laboratory of Cell Biology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892, USA.

Overexpression of ATP-binding cassette (ABC) drug transporters that actively efflux a variety of amphipathic compounds can cause multidrug resistance (MDR) in cancer cells, which is a major obstacle in the success of cancer chemotherapy. The development of synthetic small molecule compounds or the identification of natural products that block ABC transporter-mediated efflux has been the conventional approach used to combat MDR. The strategy of using chemosensitizers, however, has not been successful in clinical cancer chemotherapy. Therefore, alternative approaches to identify or to synthesize compounds that can induce selective toxicity in cancer cells overexpressing one or more ABC transporters have been undertaken. This review summarizes the recent advances in identifying strategies to restore sensitivity to chemotherapeutics in multidrug resistant cancer cells.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2600768PMC
http://dx.doi.org/10.2174/1874467210801020093DOI Listing
June 2008

Brca1 breast tumors contain distinct CD44+/CD24- and CD133+ cells with cancer stem cell characteristics.

Breast Cancer Res 2008 1;10(1):R10. Epub 2008 Feb 1.

Laboratory of Human Carcinogenesis, Center for Cancer Research, National Cancer Institute, 9000 Rockville Pike, Bethesda, Maryland 20892, USA.

Introduction: Whether cancer stem cells occur in BRCA1-associated breast cancer and contribute to therapeutic response is not known.

Methods: We generated and characterized 16 cell lines from five distinct Brca1deficient mouse mammary tumors with respect to their cancer stem cell characteristics.

Results: All cell lines derived from one tumor included increased numbers of CD44+/CD24- cells, which were previously identified as human breast cancer stem cells. All cell lines derived from another mammary tumor exhibited low levels of CD44+/CD24- cells, but they harbored 2% to 5.9% CD133+ cells, which were previously associated with cancer stem cells in other human and murine tumors. When plated in the absence of attachment without presorting, only those cell lines that were enriched in either stem cell marker formed spheroids, which were further enriched in cells expressing the respective cancer stem cell marker. In contrast, cells sorted for CD44+/CD24- or CD133+ markers lost their stem cell phenotype when cultured in monolayers. As few as 50 to 100 CD44+/CD24- or CD133+ sorted cells rapidly formed tumors in nonobese diabetic/severe combined immunodeficient mice, whereas 50-fold to 100-fold higher numbers of parental or stem cell depleted cells were required to form few, slow-growing tumors. Expression of stem cell associated genes, including Oct4, Notch1, Aldh1, Fgfr1, and Sox1, was increased in CD44+/CD24- and CD133+ cells. In addition, cells sorted for cancer stem cell markers and spheroid-forming cells were significantly more resistant to DNA-damaging drugs than were parental or stem cell depleted populations, and they were sensitized to the drugs by the heat shock protein-90 inhibitor 17-DMAG (17-dimethylaminoethylamino-17-demethoxygeldanamycin hydrochloride).

Conclusion: Brca1-deficient mouse mammary tumors harbor heterogeneous cancer stem cell populations, and CD44+/CD24- cells represent a population that correlates with human breast cancer stem cells.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1186/bcr1855DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2374965PMC
June 2008

Evidence for dual mode of action of a thiosemicarbazone, NSC73306: a potent substrate of the multidrug resistance linked ABCG2 transporter.

Mol Cancer Ther 2007 Dec;6(12 Pt 1):3287-96

Laboratory of Cell Biology, Center for Cancer Research, National Cancer Institute, NIH, Bethesda, Maryland 20892-4256, USA.

Multidrug resistance due to reduced drug accumulation is a phenomenon predominantly caused by the overexpression of members of the ATP-binding cassette (ABC) transporters, including ABCB1 (P-glycoprotein), ABCG2, and several ABCC family members [multidrug resistance-associated protein (MRP)]. We previously reported that a thiosemicarbazone derivative, NSC73306, is cytotoxic to carcinoma cells that overexpress functional P-glycoprotein, and it resensitizes these cells to chemotherapeutics. In this study, we investigated the effect of NSC73306 on cells overexpressing other ABC drug transporters, including ABCG2, MRP1, MRP4, and MRP5. Our findings showed that NSC73306 is not more toxic to cells that overexpress these transporters compared with their respective parental cells, and these transporters do not confer resistance to NSC73306 either. In spite of this, we observed that NSC73306 is a transport substrate for ABCG2 that can effectively inhibit ABCG2-mediated drug transport and reverse resistance to both mitoxantrone and topotecan in ABCG2-expressing cells. Interactions between NSC73306 and the ABCG2 drug-binding site(s) were confirmed by its stimulatory effect on ATPase activity (140-150 nmol/L concentration required for 50% stimulation) and by inhibition of [(125)I]iodoarylazidoprazosin photolabeling (50% inhibition at 250-400 nmol/L) of the substrate-binding site(s). Overall, NSC73306 seems to be a potent modulator of ABCG2 that does not interact with MRP1, MRP4, or MRP5. Collectively, these data suggest that NSC73306 can potentially be used, due to its dual mode of action, as an effective agent to overcome drug resistance by eliminating P-glycoprotein-overexpressing cells and by acting as a potent modulator that resensitizes ABCG2-expressing cancer cells to chemotherapeutics.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1158/1535-7163.MCT-07-2005DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2268031PMC
December 2007

ABC drug transporters as molecular targets for the prevention of multidrug resistance and drug-drug interactions.

Curr Drug Deliv 2007 Oct;4(4):324-33

Laboratory of Cell Biology, Center for Cancer Research, National Cancer Institute, NIH/DHHS, Bethesda, MD 20892, USA.

ABC transporters play an important role in mediating the cytoplasmic concentration of endogenous and xenobiotic substances. They therefore influence the pharmacokinetic profile of a variety of drugs. By virtue of their localization to plasma membranes in the intestine, liver, blood-brain and other vital biological barriers, a majority of ABC drug transporters cause drug-drug interactions, decreased drug efficacy and multidrug resistance for chemotherapeutic agents. Thus, elucidating which drug entities are substrates for ABC drug transporters is a crucial step in the drug development and treatment process. Here, we review the current status of methodology used to categorize drug compounds as substrates or modulators for the major ABC drug transporters including ABCB1, ABCC1 and ABCG2.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.2174/156720107782151241DOI Listing
October 2007

A "silent" polymorphism in the MDR1 gene changes substrate specificity.

Science 2007 Jan 21;315(5811):525-8. Epub 2006 Dec 21.

Laboratory of Cell Biology, Center for Cancer Research, National Cancer Institute, Bethesda, MD 20892, USA.

Synonymous single-nucleotide polymorphisms (SNPs) do not produce altered coding sequences, and therefore they are not expected to change the function of the protein in which they occur. We report that a synonymous SNP in the Multidrug Resistance 1 (MDR1) gene, part of a haplotype previously linked to altered function of the MDR1 gene product P-glycoprotein (P-gp), nonetheless results in P-gp with altered drug and inhibitor interactions. Similar mRNA and protein levels, but altered conformations, were found for wild-type and polymorphic P-gp. We hypothesize that the presence of a rare codon, marked by the synonymous polymorphism, affects the timing of cotranslational folding and insertion of P-gp into the membrane, thereby altering the structure of substrate and inhibitor interaction sites.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1126/science.1135308DOI Listing
January 2007

Role of the ABCG2 drug transporter in the resistance and oral bioavailability of a potent cyclin-dependent kinase/Aurora kinase inhibitor.

Mol Cancer Ther 2006 Oct;5(10):2459-67

Cancer Therapeutics Research, Johnson & Johnson Pharmaceutical Research and Development, LLC, Raritan, NJ 08560, USA.

Cell cycle kinase inhibitors have advanced into clinical trials in oncology. One such molecule, JNJ-7706621, is a broad-spectrum inhibitor of the cyclin-dependent kinases and Aurora kinases that mediate G(2)-M arrest and inhibits tumor growth in xenograft models. To determine the putative mechanisms of resistance to JNJ-7706621 that might be encountered in the clinic, the human epithelial cervical carcinoma cell line (HeLa) was exposed to incrementally increasing concentrations of JNJ-7706621. The resulting resistant cell population, designated HeLa-6621, was 16-fold resistant to JNJ-7706621, cross-resistant to mitoxantrone (15-fold) and topotecan (6-fold), and exhibited reduced intracellular drug accumulation of JNJ-7706621. ABCG2 was highly overexpressed at both the mRNA ( approximately 163-fold) and protein levels. The functional role of ABCG2 in mediating resistance to JNJ-7706621 was consistent with the following findings: (a) an ABCG2 inhibitor, fumitremorgin C, restored the sensitivity of HeLa-6621 cells to JNJ-7706621 and to mitoxantrone; (b) human embryonic kidney-293 cells transfected with ABCG2 were resistant to both JNJ-7706621 and mitoxantrone; and (c) resistant cells that were removed from the drug for 12 weeks and reverted to susceptibility to JNJ-7706621 showed near-normal ABCG2 RNA levels. ABCG2 is likely to limit the bioavailability of JNJ-7706621 because oral administration of JNJ-7706621 to Bcrp (the murine homologue of ABCG2) knockout mice resulted in an increase in the plasma concentration of JNJ-7706621 compared with wild-type mice. These findings indicate that ABCG2 mediates the resistance to JNJ-7706621 and alters the absorption of the compound following administration.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1158/1535-7163.MCT-06-0339DOI Listing
October 2006

Plasma membrane calcium ATPase (PMCA4): a housekeeper for RT-PCR relative quantification of polytopic membrane proteins.

BMC Mol Biol 2006 Sep 17;7:29. Epub 2006 Sep 17.

Laboratory of Cell Biology, Center for Cancer Research, National Cancer Institute, NIH, DHHS, Bethesda, MD 20892-42546, USA.

Background: Although relative quantification of real-time RT-PCR data can provide valuable information, one limitation remains the selection of an appropriate reference gene. No one gene has emerged as a universal reference gene and much debate surrounds some of the more commonly used reference genes, such as glyceraldehyde-3-phosphate dehydrogenase (GAPDH). At this time, no gene encoding for a plasma membrane protein serves as a reference gene, and relative quantification of plasma membrane proteins is performed with genes encoding soluble proteins, which differ greatly in quantity and in targeting and trafficking from plasma membrane proteins. In this work, our aim was to identify a housekeeping gene, ideally one that codes for a plasma membrane protein, whose expression remains the same regardless of drug treatment and across a wide range of tissues to be used for relative quantification of real-time RT-PCR data for ATP binding cassette (ABC) plasma membrane transporters.

Results: In studies evaluating the expression levels of two commonly used reference genes coding for soluble proteins and two genes coding for membrane proteins, one plasma membrane protein, plasma membrane calcium-ATPase 4 (PMCA4), was comparable to the two reference genes already in use. In addition, PMCA4 expression shows little variation across eight drug-treated cell lines and was found to be superior to GAPDH and HPRT1, commonly used reference genes. Finally, we show PMCA4 used as a reference gene for normalizing ABC transporter expression in a drug-resistant lung carcinoma cell line.

Conclusion: We have found that PMCA4 is a good housekeeping gene for normalization of gene expression for polytopic membrane proteins including transporters and receptors.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1186/1471-2199-7-29DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC1586022PMC
September 2006

Comparison of drug transporter levels in normal colon, colon cancer, and Caco-2 cells: impact on drug disposition and discovery.

Mol Pharm 2006 Jan-Feb;3(1):87-93

Laboratory of Cell Biology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, Maryland 20892, USA.

A critical step in early phase drug development is the determination of oral bioavailability. In part, the ability to predict whether a drug will be effectively transported across the gastrointestinal mucosa can be estimated from the physicochemical properties of the compound. Although advancements through rational drug design have more correctly predicted bioavailability, considerable variability remains to be explained. Transporter expression throughout the gastrointestinal tract may explain much of this variation. ATP-binding cassette (ABC) transporters were the first family of transporters identified to modify bioavailability. More recently, the solute carrier family has also been shown to alter the pharmacokinetic profile of drugs. Currently, the Caco-2 human colon carcinoma cell line is often used by the pharmaceutical industry to evaluate intestinal absorption of drugs; however, in vivo/in vitro permeabilities with carrier mediated drugs do not correlate well, suggesting that Caco-2 transporter expression varies from that of the small intestine. With this is mind, we integrated U133A GeneChip expression data from the NCBIs Gene Expression Omnibus (GEO) collection and then compared the expression pattern of Caco-2 cells to normal colon to determine if the Caco-2 cell line is a reliable model for colonic delivery. Furthermore, transporter expression of Caco-2 cells was compared to that of human colon tumors to assess whether this cell line could be useful to predict drug absorption for colon cancer. Our analysis shows that the expression pattern for Caco-2 cells closely resembles the gene expression profile of transporters within the normal colon, suggesting that this cell line may serve as an in vitro model of colonic drug adsorption. However, the molecular "fingerprint" of Caco-2 was distinctly different from tumor samples, indicating that the Caco-2 model would unlikely predict accurate drug absorption for colon cancer sites.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1021/mp050090kDOI Listing
June 2006

Modulatory effects of plant phenols on human multidrug-resistance proteins 1, 4 and 5 (ABCC1, 4 and 5).

FEBS J 2005 Sep;272(18):4725-40

Department of Pharmacology, University of Cambridge, UK.

Plant flavonoids are polyphenolic compounds, commonly found in vegetables, fruits and many food sources that form a significant portion of our diet. These compounds have been shown to interact with several ATP-binding cassette transporters that are linked with anticancer and antiviral drug resistance and, as such, may be beneficial in modulating drug resistance. This study investigates the interactions of six common polyphenols; quercetin, silymarin, resveratrol, naringenin, daidzein and hesperetin with the multidrug-resistance-associated proteins, MRP1, MRP4 and MRP5. At nontoxic concentrations, several of the polyphenols were able to modulate MRP1-, MRP4- and MRP5-mediated drug resistance, though to varying extents. The polyphenols also reversed resistance to NSC251820, a compound that appears to be a good substrate for MRP4, as predicted by data-mining studies. Furthermore, most of the polyphenols showed direct inhibition of MRP1-mediated [3H]dinitrophenyl S-glutathione and MRP4-mediated [3H]cGMP transport in inside-out vesicles prepared from human erythrocytes. Also, both quercetin and silymarin were found to inhibit MRP1-, MRP4- and MRP5-mediated transport from intact cells with high affinity. They also had significant effects on the ATPase activity of MRP1 and MRP4 without having any effect on [32P]8-azidoATP[alphaP] binding to these proteins. This suggests that these flavonoids most likely interact at the transporter's substrate-binding sites. Collectively, these results suggest that dietary flavonoids such as quercetin and silymarin can modulate transport activities of MRP1, -4 and -5. Such interactions could influence bioavailability of anticancer and antiviral drugs in vivo and thus, should be considered for increasing efficacy in drug therapies.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1111/j.1742-4658.2005.04888.xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC1350838PMC
September 2005

Modulation of cell adhesion molecules in various epithelial cell lines after treatment with PP2.

Mol Pharm 2005 May-Jun;2(3):170-84

Department of Pharmaceutical Chemistry, The University of Kansas, Lawrence, Kansas 66047, USA.

Regulation and expression of E-cadherin and other adhesion molecules were evaluated after exposure to a selective inhibitor of the Src family of tyrosine kinases and inducer of E-cadherin, PP2. E-cadherin is located within the intercellular junction, and it is involved in the management of paracellular permeability of various epithelial barriers in the body. Epithelial cell lines HCT-116, HT29, Caco-2, LS174T, and ARPE-19 were examined for morphological, functional, protein, and mRNA changes following 20 microM PP2 treatment. PP2 treatment caused cell clustering in Caco-2, HT29, and HCT-116 cells. E-cadherin also redistributed to the points of cell contact in Caco-2 cells. These changes suggest increased E-cadherin-dependent cell adhesion. Studies evaluating transepithelial electrical resistance, an established measurement of paracellular permeability, displayed increases in resistance for the Caco-2 cells following PP2 treatment, which correlates with our microscopy data. In addition, E-cadherin protein levels increased for all cells except HCT-116. ARPE-19 cells did not express E-cadherin at the protein or mRNA level. Expression of adhesion molecules varied for the cell lines, and only Claudin 3 mRNA expression was significantly increased in the three intestinal cell lines treated with PP2. Overall, our data suggest that E-cadherin is positively regulated by inhibition of Src tyrosine kinases at the functional and protein expression levels within these epithelial cell lines.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1021/mp0499003DOI Listing
August 2005

Effects of an E-cadherin-derived peptide on the gene expression of Caco-2 cells.

Pharm Res 2004 Nov;21(11):2085-94

Department of Pharmaceutical Chemistry, The University of Kansas, Lawrence, Kansas 66047, USA.

Purpose: The goal of this study was to determine the effects of exposure to an HAV peptide (Ac-SHAVSS-NH2) on the protein and gene expression in Caco-2 cells, a model for the intestinal mucosa.

Methods: Caco-2 cells were incubated with either 100 or 500 microM of the hexapeptide then evaluated over a 48-h time period.

Results: Cell detachment from the monolayer was seen only after 48 h of exposure to the peptide, with the greatest effects occurring with a peptide concentration of 500 microM. Total protein expression of E-cadherin showed a decrease of nearly 20% at the 24-h time point for each concentration examined, whereas no significant changes were detected at the other time points studied. Short term exposure to a 500 microM solution of Ac-SHAVSS-NH2 caused few changes in gene expression as determined by Affymetrix GeneChip microarrays; however, longer exposure periods produced numerous changes in the treated cells. The variations in mRNA expression indicate that this HAV peptide has an effect in the E-cadherin signaling pathways. The greatest increases in mRNA expression were found in genes regulating excretion or degradation of the peptide.

Conclusions: This work suggests that this HAV peptide produces effects that reach beyond modulation of adhesion.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1023/b:pham.0000048201.00143.72DOI Listing
November 2004
-->