TY - CHAP
T1 - Combination of Tissue Microarray Profiling and Multiplexed IHC Approaches to Investigate Transport Mechanism of Nucleoside Analog Drug Resistance
AU - Nayak, Debasis
AU - Weadick, Brenna
AU - Govindarajan, Rajgopal
N1 - Funding Information:
This work is supported by a grant award of National Institute of Health (R01GM143217 and R03CA262490) and a Research Scholars Grant (RSG-15-036-01-DDC) from the American Cancer Society awarded to R.G. IHC images presented in this article were generated and analyzed using the instruments and services at the Campus Microscopy and Imaging Facility (CMIF) at The Ohio State University. This facility is partly supported by grant P30 CA016058 from National Cancer Institute, Bethesda, MD. We are grateful to Dr. Terence M. Williams, Department of Radiation Oncology, The Ohio State University Comprehensive Cancer Center and Dr. Wei Chen, Department of Pathology, The Ohio State University Wexner Medical Center for generating the TMA and help in analyzing the IHC images. We acknowledge Pelotonia Fellowship Program of The Ohio State University Comprehensive Cancer Center for supporting D.N.
Publisher Copyright:
© 2023, The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature.
PY - 2023
Y1 - 2023
N2 - Nucleoside analogs (NAs) are an established class of anticancer agents being used clinically for the treatment of diverse cancers, either as monotherapy or in combination with other established anticancer or pharmacological agents. To date, nearly a dozen anticancer NAs are approved by the FDA, and several novel NAs are being tested in preclinical and clinical trials for future applications. However, improper delivery of NAs into tumor cells because of alterations in expression of one or more drug carrier proteins (e.g., solute carrier (SLC) transporters) within tumor cells or cells surrounding the tumor microenvironment stands as one of the primary reasons for therapeutic drug resistance. The combination of tissue microarray (TMA) and multiplexed immunohistochemistry (IHC) is an advanced, high-throughput approach over conventional IHC that enables researchers to effectively investigate alterations to numerous such chemosensitivity determinants simultaneously in hundreds of tumor tissues derived from patients. In this chapter, taking an example of a TMA from pancreatic cancer patients treated with gemcitabine (a NA chemotherapeutic agent), we describe the step-by-step procedure of performing multiplexed IHC, imaging of TMA slides, and quantification of expression of some relevant markers in these tissue sections as optimized in our laboratory and discuss considerations while designing and carrying out this experiment.
AB - Nucleoside analogs (NAs) are an established class of anticancer agents being used clinically for the treatment of diverse cancers, either as monotherapy or in combination with other established anticancer or pharmacological agents. To date, nearly a dozen anticancer NAs are approved by the FDA, and several novel NAs are being tested in preclinical and clinical trials for future applications. However, improper delivery of NAs into tumor cells because of alterations in expression of one or more drug carrier proteins (e.g., solute carrier (SLC) transporters) within tumor cells or cells surrounding the tumor microenvironment stands as one of the primary reasons for therapeutic drug resistance. The combination of tissue microarray (TMA) and multiplexed immunohistochemistry (IHC) is an advanced, high-throughput approach over conventional IHC that enables researchers to effectively investigate alterations to numerous such chemosensitivity determinants simultaneously in hundreds of tumor tissues derived from patients. In this chapter, taking an example of a TMA from pancreatic cancer patients treated with gemcitabine (a NA chemotherapeutic agent), we describe the step-by-step procedure of performing multiplexed IHC, imaging of TMA slides, and quantification of expression of some relevant markers in these tissue sections as optimized in our laboratory and discuss considerations while designing and carrying out this experiment.
KW - Equilibrative nucleoside transporter (ENT), Concentrative nucleoside transporters (CNT)
KW - Heat-induced antigen retrieval (HIER)
KW - Immunohistochemistry, Multiplexing, Solute carrier (SLC) transporters
KW - Nucleoside analogs
KW - Nucleoside transporter (NT)
KW - Pancreatic cancer
KW - Tissue microarray
UR - http://www.scopus.com/inward/record.url?scp=85159740163&partnerID=8YFLogxK
U2 - 10.1007/978-1-0716-3163-8_8
DO - 10.1007/978-1-0716-3163-8_8
M3 - Chapter
C2 - 37191793
AN - SCOPUS:85159740163
T3 - Methods in Molecular Biology
SP - 95
EP - 121
BT - Methods in Molecular Biology
PB - Humana Press Inc.
ER -