Methods in Molecular Biology

Our medical research library holds 10 studies published in Methods in Molecular Biology (2006–2019). Each entry links to the study's record on PubMed and, where there is one, to the publisher's page and a free full text. 10 have an abstract, the authors' own summary as PubMed records it: open "Abstract" to read it.

About the journal

• Publisher: Humana Press
• Published: 1984 to the present, United States
• Language: English
• ISSN: 1064-3745 (print), 1940-6029 (electronic)
• Its record in the NLM Catalog

2010s

• Autophagy in 3D In Vitro and Ex Vivo Cancer Models. C. Follo, D. Barbone, W.G. Richards, et al. 2019;1880:491-510. PubMed · Publisher

  Abstract

  Three-dimensional (3D) models are acquiring importance in cancer research due to their ability to mimic multiple features of the tumor microenvironment more accurately than standard monolayer two-dimensional (2D) cultures. Several groups, including our laboratory, are now accumulating evidence that autophagy in solid tumors is also better represented in 3D than in 2D. Here we detail how we generate 3D models, both in vitro multicellular spheroids generated from cell lines and ex vivo tumor fragment spheroids generated from tumor samples, and how autophagy can be measured in 3D cultures.

• Isolation of Chemoresistant Cell Subpopulations. C. Canino, M. Cioce. 2016;1379:139-50. PubMed · Publisher

  Abstract

  Chemoresistance is a major challenge for cancer therapy and drives tumor relapse. The emergence, within the treated tumor mass, of specific cancer cell subpopulations endowed with high tolerance to the microenvironment stress induced by therapy is being growingly recognized as a mechanism of tumor progression. To obtain detailed information with regard to the pathways underlying survival, expansion, and microenvironmental cross talk of such chemoresistant cell subpopulations may be instrumental for cancer chemoprevention. Additionally, the obtained cell subpopulations may be used for direct screening of cancer chemopreventive compounds, in appropriate experimental settings. Here we report detailed experimental procedures that we and others have setup in order to obtain cell cultures enriched for chemoresistant cells from both malignant pleural mesothelioma specimens and primary cell cultures. We provide indications for the purification and characterization of those chemoresistant cell populations and to generally validate the obtained enriched cell populations for their chemoresistance.

• microRNAs in Cancer Chemoprevention: Method to Isolate Them from Fresh Tissues. F. Ganci, G. Blandino. 2016;1379:21-9. PubMed · Publisher

  Abstract

  microRNAs are 22-nucleotide-long double-strand small RNAs, able to modulate gene expression at posttranscriptional level, degrading mRNA and/or impairing translation. They have been shown to regulate mRNA and protein abundance and to participate in many regulatory circuits controlling developmental timing, cell proliferation and differentiation, apoptosis and stress response. Notably, microRNA activity has been correlated to the pathogenesis of cancer; they are aberrantly expressed in solid and hematological tumors, suggesting that they could function as oncogenes or tumor suppressors. The emerging role of miRNAs in the carcinogenesis and tumor progression has provided opportunities for their clinical application in the capacity of cancer detection, diagnosis, and prognosis prediction. Here, we describe the experimental protocol used to isolate microRNAs from human tissues coming from head and neck, mesothelioma, and thymoma tumors in order to perform microarray and RT-qPCR experiments.

• Clinical Evaluation of ErbB-Targeted CAR T-Cells, Following Intracavity Delivery in Patients with ErbB-Expressing Solid Tumors. S. Papa, M. van Schalkwyk, J. Maher. 2015;1317:365-82. PubMed · Publisher

  Abstract

  Adoptive cell therapy using gene-modified T-cells has achieved impressive results in the treatment of B-cell malignancies. However, the development of similar strategies to treat solid tumors raises challenges with respect to tumor antigen selection and the achievement of efficient T-cell homing, survival and sustained effector function within the tumor microenvironment. To address these challenges, we have developed a gene-modified cellular therapy called T4 immunotherapy. To generate T4 immunotherapy, autologous T-cells are engineered by retroviral transduction to co-express two transgenes: (1) a chimeric antigen receptor (CAR), T1E28z, targeted against a range of ErbB homodimers and heterodimers and (2) a chimeric cytokine receptor, 4αβ, that allows the selective ex vivo expansion of engineered cells using interleukin-4. Targeting of the extended ErbB network using CAR T-cells is supported by prevalence of ErbB dysregulation in diverse solid tumors and the clinical impact of monoclonal antibody therapy directed against members of this family. However, the key obstacle to effective clinical translation is risk of on-target toxicity owing to the lower level expression of ErbB family members in many healthy tissues. To de-risk T4 immunotherapy in man, we are undertaking a trial in patients with locally advanced or recurrent head and neck squamous cell carcinoma. In that setting, engineered T-cells are injected directly into the tumor without prior lymphodepletion, an approach that we believe will minimize risk of toxicity. This chapter outlines how we plan to advance the development of T4 immunotherapy thereafter in Phase II clinical testing. In that setting, regional (intracavitary) approaches will be used to administer this therapy to patients with epithelial ovarian cancer and malignant pleural mesothelioma.

• Methylation profile landscape in mesothelioma: possible implications in early detection, disease progression, and therapeutic options. X. Zhang, N. Tang, A.K. Rishi, et al. 2015;1238:235-47. Review PubMed · Publisher

  Abstract

  Malignant Pleural Mesothelioma (MPM) is an aggressive malignancy of the pleura associated with asbestos exposure. Incidence of MPM is expected to increase over the course of next decade in both Europe and the developing countries. Although significant progress has been made in terms of etiology and pathogenesis of this disease, currently available therapeutic options have not significantly improved the survival outcome of patients on standard chemotherapeutic regimens. Integrity of the cellular DNA is often altered in many cancers. Understanding of the molecular mechanisms that regulate cellular DNA alterations to facilitate cancer initiation and development has potential to allow better design of cancer cell inhibitory strategies. In this context, there is a need to explore the gamut of "omics" strategies to provide a comprehensive epigenetics profile for MPM. This chapter discusses the functional genomics and epigenetic patterns observed by various investigators studying MPM patient populations on global fronts, and attempts to present a holistic approach in combating this insidious disease. Here we provide investigators in this field with novel insights and methodologies used in other types of cancers that might have profound impact in the early detection, prognosis and potential therapeutic strategies for MPM.

• Electron microscopy: a brief history and review of current clinical application. R.E. Gordon. 2014;1180:119-35. PubMed · Publisher

  Abstract

  This chapter describes the historic development of techniques that has made it possible to use electron microscopy, principally transmission electron microscopy, for diagnostic purposes. It was described how the standard techniques for preparing tissue for light microscopy had been modified to make it possible to view the ultrastructural components of a cell, tissue, or organ that cannot be resolved with a light microscope. There is a discussion of the types of tissues and cells that were and are currently observed by electron microscopy for diagnostic purposes. All of the materials that are used in tissue preparation and the general protocols for processing the tissues are also included. There are also notes which describe steps that can be changed or modified and why depending on conditions and anticipated outcome.

• Applications of array-CGH for lung cancer. K.J. Craddock, W.L. Lam, M.S. Tsao. 2013;973:297-324. Review PubMed · Publisher

  Abstract

  This chapter summarizes the current knowledge on gene copy number changes found in lung tumors, and their application in the diagnosis, prognostication, and prediction of response to chemotherapy. Examples of the identification of specific "driver" oncogenes within amplified DNA segments are described. A model of how array-CGH could be integrated clinically into the routine workup of lung cancers in clinical laboratory is proposed.

2000s

• Environmental and occupational risk factors for lung cancer. I. Brüske-Hohlfeld. 2009;472:3-23. Review PubMed · Publisher

  Abstract

  Lung cancer is the world's leading cause of cancer death. It is primarily due to the inhalation of carcinogens and highly accessible to prevention by diminishing exposures to lung carcinogens. Most important will be the complete cessation of exposure to cigarette smoke (first and second hand) and to asbestos. Two environmental exposures--radon in homes and arsenic in drinking water--cannot be totally avoided, but people in certain geographical regions would greatly benefit from a reduction in exposure magnitude. And last but not least, workers all over the world deserve that preventive measures at the workplace are observed with regard to exposures, such as arsenic, beryllium, bis-chloromethyl ether (BCME), cadmium, chromium, polycyclic aromatic hydrocarbons (PAHs), and nickel.

• Analysis of asbestos-induced gene expression changes in bronchiolar epithelial cells using laser capture microdissection and quantitative reverse transcriptase-polymerase chain reaction. C.B. Manning, B.T. Mossman, D.J. Taatjes. 2006;319:231-6. PubMed · Publisher

  Abstract

  Laser capture microdissection (LCM) enables the removal of discrete microstructures or cell types from properly prepared histological sections. Extraction of RNA from microdissected tissue followed by quantitative reverse transcriptase-polymerase chain (QRT-PCR) reaction permits the analysis of cell-type or microstructure-specific gene expression changes that occur in response to various stimuli in the environment. In our lab, the combination of LCM and QRT-PCR has proven very useful in the determination of the in vivo gene expression changes that occur in bronchiolar epithelium in response to inhalation of crocidolite asbestos. A detailed description of the preparation of cDNA from bronchiolar epithelial cells obtained by LCM is described in this work.

• Multifluorescence labeling techniques and confocal laser scanning microscopy on lung tissue. M. Stern, D.J. Taatjes, B.T. Mossman. 2006;319:67-76. PubMed · Publisher

  Abstract

  Lung tissue consists of more than 40 individual cell types that might interact to produce adverse pathologies. After injury, a number of signaling proteins expressed in various epithelial and other cell types have been linked to the advent of apoptosis, compensatory proliferation, and adaptation to stress. We describe here the use of immunochemistry and multifluorescence approaches using confocal laser scanning microscopy to define the signaling pathways (protein kinases C and mitogen-activated protein kinases) activated by asbestos fibers after inhalation. Using these approaches, we are able to localize signaling events in distinct cell types of the lung and determine their status in the cell cycle (resting or nonresting). Moreover, we are able to determine whether various signaling proteins colocalize in cells and the sites affected by asbestos fibers.

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