Biomaterials for 3D Tumor Modeling

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BIOMATERIALS FOR 3D TUMOR
MODELING
 BIOMATERIALS
FOR 3D TUMOR
MODELING
Edited by

SUBHAS C. KUNDU
3B’s Research Group, I3Bs Research Institute on Biomaterials, Biodegradables and Biomimetics, University of
Minho, Headquarters of the European Institute of Excellence on Tissue Engineering and Regenerative Medicine,
Guimarães, Portugal
ICVS/3B’s PT Government Associate Laboratory, Braga/Guimarães, Portugal

RUI L. REIS
3B’s Research Group, I3Bs Research Institute on Biomaterials, Biodegradables and Biomimetics, University of
Minho, Headquarters of the European Institute of Excellence on Tissue Engineering and Regenerative Medicine,
Guimarães, Portugal
ICVS/3B’s PT Government Associate Laboratory, Braga/Guimarães, Portugal
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 Contents

List of Contributors xi 2.3 3D In vitro tumor models—bridging the gap

Preface xvii from 2D flat cultures to in vivo 45
2.4 Classes of 3D multicellular tumor models 47
2.5 Conclusions 61
I References 62

Engineering biomaterials for 3D 3. Biofabrication of 3D tumor models in

cancer modelling cancer research 67
M.A. Grimaudo, A. Herreros-Pomares, M. Alonso, S. Calabuig-
Fariñas, E. Jantus-Lewintre and Maria de la Fuente
1. Trends in biomaterials for three-
dimensional cancer modeling 3 3.1 Current challenges in oncology 67
David Caballero, Rui L. Reis and Subhas C. Kundu 3.2 The tumor microenvironment 69
3.3 Development of the cancer therapeutics
Abbreviations 3 field 71
1.1 A historical introduction 4 3.4 3D tumor models in cancer research 73
1.2 The three-dimensional tumor 3.5 Evaluation of anticancer therapeutics in 3D
microenvironment 6 tumor models 78
1.3 Engineering the native tumor 3.6 Implementation of 3D tumor models in a
microenvironment using custom-designed clinical setting 82
three-dimensional biomaterials 15 3.7 Final remarks 86
1.4 Advanced models of the three-dimensional Acknowledgments 86
tumor microenvironment 22 References 86
1.5 Applications of three-dimensional tumor
models in cancer therapeutics 30 4. Biomatrices that mimic the
1.6 Limitations of biomaterials-based three- cancer extracellular environment 91
dimensional tumor models 33 Sara Amorim, Rui L. Reis and Ricardo A. Pires
1.7 Future of three-dimensional biomaterials for
cancer research 34 4.1 Introduction 91
1.8 Final remarks and conclusions 35 4.2 The three-dimensional in vitro models 92
Acknowledgments 36 4.3 Conclusions and future remarks 102
References 36 References 102

5. 3D neuroblastoma in vitro models using

2. Bioinspired biomaterials to develop
engineered cell-derived matrices 107
cell-rich spherical microtissues for 3D Enrico Almici, David Caballero, Joan Montero and Josep Samitier
in vitro tumor modeling 43
Maria V. Monteiro, Vı́tor M. Gaspar and João F. Mano 5.1 Introduction 107
5.2 Neuroblastoma 108
2.1 Introduction 43 5.3 Cell-derived matrices in tumor modeling 112
2.2 Human Tumor microenvironment—key 5.4 Engineering cell-derived matrix
hallmarks to mimic in vitro 44 deposition 115

v
vi Contents

5.5 Cell-derived matrices and cell morphodynamic 8.5 Microvessels on a chip 184
characterization 123 8.6 Future perspectives 186
5.6 Cell-derived matrix capture relevant processes Acknowledgments 186
involved in neuroblastoma malignancy 125 References 187
5.7 Conclusions 126
Acknowledgments 126 9. Metastasis in three-dimensional
References 126 biomaterials 191
Banani Kundu, Rui L. Reis and Subhas C. Kundu
6. 3D culture systems as models for solid
tumors and cancer metabolism 131 9.1 Why biomaterial is needed in cancer
Sónia Pires Celeiro, Fátima Baltazar and Marta Viana-Pereira modeling? 191
9.2 Biomaterials employed in tumor ECM
Abbreviations 131 modeling 193
6.1 Introduction 132 9.3 Properties of cell surrounding matrix/niche
6.2 Solid tumors: tumor microenvironment and contribute to tumor cell migration 197
tumorigenesis 133 9.4 Biomaterial-based stepwise modeling of cancer
6.3 Cancer metabolism: influence in tumor metastasis in vitro 199
microenvironment 135 9.5 Biomaterial-based in vitro models of cancer
6.4 Solid tumors in vitro models 139 dormancy and reactivation 208
6.5 3D cell culture systems in cancer research 143 9.6 Concluding remarks 209
6.6 3D cell culture systems for study cancer Acknowledgments 210
metabolism 146 References 210
6.7 Conclusions 148
Acknowledgments 149 10. 3D cancer spheroids and
Conflict of interest 149 microtissues 217
References 149
Virginia Brancato, Rui L. Reis and Subhas C. Kundu

7. Biomaterials as ECM-like matrices for Abbreviations 217

3D in vitro tumor models 157 10.1 Introduction 217
Lara Pierantoni, Joana Silva-Correia, Antonella Motta, Rui L. Reis 10.2 Biomaterials advances tumor cell culture to
and Joaquim M. Oliveira
the third dimension 219
Abbreviations 157 10.3 Recapitulating the tumor stroma crosstalk in
7.1 Introduction 158 spheroid and microtissue models 223
7.2 Biomaterials as ECM-like matrices for cancer 10.4 Vascularized microtumor models 224
3D in vitro models 158 10.5 The contribution of immune system cells in
7.3 Conclusion and future trends 170 microtumors 226
Acknowledgments 170 10.6 Spheroids as screening platform for drug
References 170 testing 228
10.7 Conclusion and future trends 230
8. Three-dimensional in vitro models of Acknowledgments 231
angiogenesis 175 References 231
Laura di Blasio, Marianela Vara-Messler and Luca Primo
11. Biomaterial-based in vitro models for
8.1 Vessels formation and tumor angiogenesis 175 pancreatic cancer 235
8.2 Vascular extracellular matrix 176 Eirini Velliou, Priyanka Gupta, Claudio Ricci and Serena Danti
8.3 Endothelial cells-based 3D angiogenesis
models 178 11.1 Introduction 235
8.4 Vascular explant-based 3D angiogenesis 11.2 In vitro 3D models for pancreatic
models 182 cancer 237
 Contents vii
11.3 Using 3D models for disease
understanding 241
II
11.4 Using 3D models for therapeutic Advanced models for cancer
screening 246
11.5 Conclusions and future trends 247
research
References 247
15. Microfluidic systems in cancer
12. In vitro three-dimensional modeling research 331
for prostate cancer 251 David Caballero, Maria Angélica Luque-González, Rui L. Reis and
Subhas C. Kundu
Eleonora Dondossola and Claudia Paindelli
15.1 Introduction 331
12.1 Introduction 251 15.2 Fundamentals of microfluidics: fluid mechanics
12.2 Modeling primary tumors 254 in miniaturized devices 336
12.3 Modeling early stages of prostate cancer 15.3 Fabrication principles of microfluidic
progression 267 devices 339
12.4 Modeling advanced stages of prostate cancer 15.4 Mimicking the tumor microenvironment using
progression 273 microfluidics 343
12.5 Conclusion 280 15.5 Microfluidic models of cancer 354
References 281 15.6 Future perspectives 366
15.7 Conclusions 370
13. 3D in vitro cutaneous melanoma Acknowledgments 370
models 287 Conflicts of interest 370
Ana I. Soares, Rui L. Reis and Alexandra P. Marques References 370

Abbreviations 287 16. Perfusion-based 3D tumor-on-chip

13.1 Introduction 288 devices for anticancer drug testing 379
13.2 Types of melanoma 289 Nandini Dhiman and Subha Narayan Rath
13.3 Risk factors for melanoma 291
13.4 Cutaneous melanoma development 293 Abbreviations 379
13.5 Cutaneous melanoma treatment 293 16.1 Introduction 379
13.6 In vitro models 296 16.2 Disadvantages of 2D in vitro, 3D in vitro, and
References 300 animal models 380
16.3 Microfluidic devices for tumor modeling 382
16.4 Tumor components and their inclusion in
14. 3D scaffold materials for skin cancer tumor-on-chip 383
modeling 305 16.5 Types of perfusion methods 387
Amir Zarebkohan, Roghayeh Sheervalilou, Roya Ghods, Subhas C. 16.6 Benefits of perfusion and specific
Kundu and Mazaher Gholipourmalekabadi applications 388
16.7 Specific designs for enhancing perfusion 393
14.1 Introduction 305 16.8 Conclusion 394
14.2 Effective factors in cell culture; 2D and 3D Acknowledgments 394
models 308 References 394
14.3 Skin cancers 313
14.4 Modeling of skin cancer 315 17. Engineering breast cancer models
14.5 Conclusion and future prospective 321 in vitro with 3D bioprinting 399
Acknowledgment 322 Brad A. Krajina and Daniela F. Duarte Campos
Conflict of interest 322
References 322 17.1 Breast cancer microenvironment in vivo 399
viii Contents

17.2 Biomaterial-based breast cancer in vitro 19.5 Future outlook 472

models 401 References 473
17.3 Biomaterials design for in vitro breast cancer
models 407
20. Three-dimensional tumor model and
17.4 3D bioprinting methods and their suitability
for breast cancer in vitro engineering 413 their implication in drug screening for
17.5 Discussion and outlook 417 tackling chemoresistance 481
References 419 Manashi Priyadarshini, Sibasish Mohanty, Tanushree Mahapatra,
Pallavi Mohapatra and Rupesh Dash
18. A predictive oncology framework—
Abbreviations 481
modeling tumor proliferation using a FEM
20.1 Chemoresistance in cancer 482
platform 427 20.2 3D tumor culture: an advanced model
Gianpaolo Ruocco, Paolo Caccavale and Maria Valeria De Bonis
preferred over 2D culture 483
Chapter points 427 20.3 3D culture and chemoresistance 486
18.1 Introduction 427 20.4 Methods of generating 3D culture
system 487
18.2 A perspective framework of predictive
oncology 430 20.5 3D culture and biomaterials 490
18.3 Detailed model formulation using level 1 20.6 Drug screening in 3D culture 494
modeling 436 20.7 Future aspects of the 3D tumor organoid
18.4 A sensitivity analysis of hallmark parameters: model: biobanks for tumor tissues 497
results 439 20.8 Limitations of 3D culture technology 498
18.5 POEM as a tool to empower the clinical 20.9 Conclusion 498
decisions 448 Acknowledgment 498
References 498
18.6 Conclusions 448
Acknowledgments 449
Glossary 449 21. Co-culture and 3D tumor models for
References 449 drug/gene therapy testing 505
Lara S. Costard, Harumi Ramanayake and Caroline M. Curtin

III 21.1 Introduction 505

21.2 Lung cancer 506
Tumor models for drug discovery 21.3 Breast cancer 515
and therapeutics 21.4 Prostate cancer 523
21.5 Future outlook 527
19. Tissue-engineered 3D cancer References 527
microenvironment for screening
therapeutics 453 22. Newly emerged engineering of in vitro
Nancy T. Li, Ileana L. Co, Natalie Landon-Brace, Simon Latour
and Alison P. McGuigan
3D tumor models using biomaterials for
chemotherapy 533
19.1 Introduction 453 Bo Cai, Qiying Lv, Zheng Wang and Lin Wang
19.2 Tumor microenvironment 454
19.3 Current strategies for creating cell and matrix 22.1 Introduction 533
organization to mimic 22.2 Constitution of artificially engineered tumor
microenvironment 458 models 534
19.4 Modeling important aspects of the tumor 22.3 Newly emerged engineering of in vitro 3D
microenvironment 462 tumors for chemotherapy 536
 Contents ix
22.4 Summary 545 26. Biodetection and sensing for cancer
References 546 diagnostics 643
23. Marine-derived biomaterials for cancer Rita Rebelo, Ana I. Barbosa, Subhas C. Kundu, Rui L. Reis and
Vitor M. Correlo
treatment 551
Catarina Oliveira, Ana C. Carvalho, Rui L. Reis, Nuno N. Neves, 26.1 Introduction 643
Albino Martins and Tiago H. Silva 26.2 Biomarkers for cancer detection 644
26.3 Cancer biosensors 648
23.1 Introduction 551 26.4 Commercialization and clinical trials of cancer
23.2 Marine biopolymers as bioactive agents 553 biosensors 654
23.3 Drug-delivery systems 556 26.5 Conclusions 656
23.4 Three-dimensional in vitro models of Acknowledgments 657
cancer 562 References 657
23.5 Conclusions 569
23.6 Acknowledgments 570
References 570
27. Understanding the impact of
controlled oxygen delivery to 3D cancer
24. Mesoporous silica nanoparticles for cell culture 661
cancer theranostic applications 577 David Grosh, William J. Wulftange, Robert W. Robey, Thomas J.
Pohida, Nicole Y. Morgan and Michael M. Gottesman
Mohammad Eltohamy

27.1 Introduction 661

24.1 Introduction 577
27.2 What is known about physiological oxygen
24.2 MSNs chemistry 579
levels? 662
24.3 Biological effects of MSNs 580
27.3 Importance of oxygen levels in various stages
24.4 3D modeling of MSN for cancer therapy 583
of cancer progression 664
24.5 Medical applications of MSNs 584
27.4 Techniques for measuring oxygenation 668
24.6 Diagnostic application of MSNs 594
27.5 Traditional/current strategies for controlling
24.7 Theranostics application of MSNs 596
oxygen concentration in vitro 674
24.8 Conclusions and outlook 597
27.6 Characterizing the effects of oxygenation on
References 597
cells and tissues 684
27.7 Conclusions and future prospects 688
Acknowledgments 689
IV Disclaimer 689
References 689
Point-of-care applications
25. Causes of cancer: physical, chemical, 28. Tissue engineering strategies for the
biological carcinogens, and viruses 607 treatment of skeletal maxillofacial defects
Subhayan Das, Moumita Kundu, Bikash Chandra Jena and resulting from neoplasms resections 697
Mahitosh Mandal J.P. Ribeiro, Esteban A. Astudillo-Ortiz, Pedro S. Babo and
Manuela E. Gomes
Abbreviations 607
25.1 Introduction 608 28.1 Background 697
25.2 Physical carcinogens 611 28.2 Tissue engineering for reconstruction of
25.3 Chemical carcinogens 617 ablated skeletal maxillofacial tissues 705
25.4 Biological carcinogens and viruses 621 28.3 Future perspectives and unmet challenges 720
25.5 Conclusion 629 References 722
Acknowledgments 631
References 631 Index 731
 List of Contributors

Enrico Almici Nanobioengineering Group, Fátima Baltazar Life and Health Science
Institute for Bioengineering of Catalonia Research Institute (ICVS), School of
(IBEC), The Barcelona Institute of Science and Medicine, University of Minho, Braga,
Technology (BIST), Barcelona, Spain; Portugal; ICVS/3B’s PT Government
Department of Electronics and Biomedical Associate Laboratory, Braga/Guimarães,
Engineering, University of Barcelona, Portugal
Barcelona, Spain; Networking Biomedical Ana I. Barbosa 3B’s Research Group, I3Bs
Investigation Center for Bioengineering, Research Institute on Biomaterials,
Biomaterials and Nanomedicine (CIBER- Biodegradables and Biomimetics, University of
BBN), Madrid, Spain Minho, Headquarters of the European Institute
M. Alonso Nano-Oncology and Translational of Excellence on Tissue Engineering and
Therapeutics Unit, Health Research Institute Regenerative Medicine, Guimarães, Portugal;
of Santiago de Compostela (IDIS), SERGAS, ICVS/3B’s PT Government Associate
Santiago de Compostela, Spain; Cancer Laboratory, Braga/Guimarães, Portugal
Research Network (CIBERONC), Spain
Virginia Brancato 3B’s Research Group, I3Bs
Sara Amorim 3B’s Research Group, I3Bs Research Institute on Biomaterials,
Research Institute on Biomaterials, Biodegradables and Biomimetics, University of
Biodegradables and Biomimetics, University Minho, Headquarters of the European Institute
of Minho, Headquarters of the European of Excellence on Tissue Engineering and
Institute of Excellence on Tissue Engineering Regenerative Medicine, Guimarães, Portugal;
and Regenerative Medicine, Guimarães, ICVS/3B’s PT Government Associate
Portugal; ICVS/3B’s - PT Government Laboratory, Braga/Guimarães, Portugal
Associate Laboratory, Braga/Guimarães,
David Caballero 3B’s Research Group, I3Bs
Portugal
Research Institute on Biomaterials,
Esteban A. Astudillo-Ortiz 3B’s Research Biodegradables and Biomimetics, University
Group, I3Bs Research Institute on of Minho, Headquarters of the European
Biomaterials, Biodegradables and Institute of Excellence on Tissue Engineering
Biomimetics, University of Minho, and Regenerative Medicine, Guimarães,
Guimarães, Portugal; ICVS/3B’s PT Portugal; ICVS/3B’s PT Government
Government Associate Laboratory, Braga/ Associate Laboratory, Braga/Guimarães,
Guimarães, Portugal; GIRO Research Group Portugal
in Oral Rehabilitation, University of Cuenca,
Cuenca, Ecuado Paolo Caccavale University of Basilicata,
Pedro S. Babo 3B’s Research Group, I3Bs College of Engineering, Modeling and
Research Institute on Biomaterials, Prototyping Laboratory—ModProLab,
Biodegradables and Biomimetics, University of Potenza, Italy
Minho, Headquarters of the European Institute Bo Cai Research Center for Tissue Engineering
of Excellence on Tissue Engineering and and Regenerative Medicine, Union Hospital,
Regenerative Medicine, Guimarães, Portugal; Tongji Medical College, Huazhong
ICVS/3B’s PT Government Associate University of Science and Technology,
Laboratory, Braga/Guimarães, Portugal Wuhan, P.R. China

xi
xii List of Contributors

S. Calabuig-Fariñas Molecular Oncology Materials and Bioengineering Research

Laboratory, General University Hospital of Centre (AMBER), RCSI and TCD, Dublin,
Valencia Research Foundation, Valencia, Ireland.
Spain; Cancer Research Network Serena Danti Department of Civil and
(CIBERONC), Spain; Department of Industrial Engineering, University of Pisa,
Pathology, Universitat de València, Valencia, Pisa, Italy
Spain
Subhayan Das School of Medical Science and
Ana C. Carvalho 3B’s Research Group, I3Bs Technology, Indian Institute of Technology
Research Institute on Biomaterials, Kharagpur, Kharagpur, India
Biodegradables and Biomimetics, University
Rupesh Dash Institute of Life Sciences,
of Minho, Headquarters of the European
Bhubaneswar, India
Institute of Excellence on Tissue Engineering
and Regenerative Medicine, Guimarães, Maria Valeria De Bonis University of
Portugal; ICVS/3B’s, PT Government Basilicata, College of Engineering, Modeling
Associate Laboratory, Braga/Guimarães, and Prototyping Laboratory—ModProLab,
Portugal Potenza, Italy
Sónia Pires Celeiro Life and Health Science Maria de la Fuente Nano-Oncology and
Research Institute (ICVS), School of Translational Therapeutics Unit, Health
Medicine, University of Minho, Braga, Research Institute of Santiago de Compostela
Portugal; ICVS/3B’s PT Government (IDIS), SERGAS, Santiago de Compostela,
Associate Laboratory, Braga/Guimarães, Spain; Cancer Research Network
Portugal (CIBERONC), Spain
Ileana L. Co Department of Chemical Nandini Dhiman Department of Biomedical
Engineering and Applied Chemistry, Engineering, Indian Institute of Technology
Institute for Biomaterials and Biomedical Hyderabad, Kandi, India; Department of
Engineering, University of Toronto, Toronto, Chemistry and Biotechnology, Faculty of
ON, Canada Science, Engineering, and Technology,
Swinburne University of Technology,
Vitor M. Correlo 3B’s Research Group, I3Bs Hawthorn, VIC, Australia
Research Institute on Biomaterials,
Laura di Blasio Department of Oncology,
Biodegradables and Biomimetics, University
University of Torino, Turin, Italy; Candiolo
of Minho, Headquarters of the European
Cancer Institute FPO-IRCCS, Candiolo, Italy
Institute of Excellence on Tissue Engineering
and Regenerative Medicine, Guimarães, Eleonora Dondossola David H. Koch Center
Portugal; ICVS/3B’s PT Government for Applied Research of Genitourinary
Associate Laboratory, Braga/Guimarães, Cancers, The University of Texas MD
Portugal Anderson Cancer Center, Houston, TX,
United States
Lara S. Costard Tissue Engineering Research
Group (TERG), Department of Anatomy and Daniela F. Duarte Campos Department of
Regenerative Medicine, Royal College of Materials Science & Engineering, Stanford
Surgeons in Ireland (RCSI), Dublin, Ireland University, CA, United States
Caroline M. Curtin Tissue Engineering Mohammad Eltohamy Glass Research
Research Group (TERG), Department of Department, National Research Centre, Giza,
Anatomy and Regenerative Medicine, Royal Egypt
College of Surgeons in Ireland (RCSI), Vı́tor M. Gaspar Department of Chemistry,
Dublin, Ireland; Trinity Centre of CICECO—Aveiro Institute of Materials,
Bioengineering (TCBE), Trinity College University of Aveiro, Campus Universitário
Dublin (TCD), Dublin, Ireland; Advanced de Santiago, Aveiro, Portugal
 List of Contributors xiii
Roya Ghods Oncopathology Research Center, Valencia Research Foundation, Valencia,
Iran University of Medical Sciences, Tehran, Spain; Cancer Research Network
Iran; Department of Molecular Medicine, (CIBERONC), Spain
Faculty of Advanced Technologies in E. Jantus-Lewintre Molecular Oncology
Medicine, Iran University of Medicine Laboratory, General University Hospital of
Sciences, Tehran, Iran Valencia Research Foundation, Valencia,
Mazaher Gholipourmalekabadi Department Spain; Cancer Research Network
of Molecular Medicine, Faculty of Advanced (CIBERONC), Spain; Department of
Technologies in Medicine, Iran University of Biotechnology, Universitat Politècnica de
Medicine Sciences, Tehran, Iran; Cellular and València, Valencia, Spain
Molecular Research Center, Iran University
Bikash Chandra Jena School of Medical
of Medical Sciences, Tehran, Iran;
Science and Technology, Indian Institute of
Department of Tissue Engineering &
Technology Kharagpur, Kharagpur, India
Regenerative Medicine, Faculty of Advanced
Technologies in Medicine, Iran University of Brad A. Krajina Department of Materials
Medical Sciences, Tehran, Iran Science & Engineering, Stanford University,
CA, United States
Manuela E. Gomes 3B’s Research Group, I3Bs
Research Institute on Biomaterials, Banani Kundu 3B’s Research Group, I3Bs—
Biodegradables and Biomimetics, University Research Institute on Biomaterials,
of Minho, Headquarters of the European Biodegradables and Biomimetics, University
Institute of Excellence on Tissue Engineering of Minho, Headquarters of the European
and Regenerative Medicine, Guimarães, Institute of Excellence on Tissue Engineering
Portugal; ICVS/3B’s PT Government and Regenerative Medicine, Guimarães,
Associate Laboratory, Braga/Guimarães, Portugal; ICVS/3B’s—PT Government
Portugal Associate Laboratory, Braga/Guimarães,
Michael M. Gottesman Laboratory of Cell Portugal
Biology (LCB), Center for Cancer Research, Moumita Kundu School of Medical Science
National Cancer Institute (NCI), National and Technology, Indian Institute of
Institutes of Health, Bethesda, MD, United Technology Kharagpur, Kharagpur, India
States
Subhas C. Kundu 3B’s Research Group,
M.A. Grimaudo Nano-Oncology and I3Bs—Research Institute on Biomaterials,
Translational Therapeutics Unit, Health Biodegradables and Biomimetics, University
Research Institute of Santiago de Compostela of Minho, Headquarters of the European
(IDIS), SERGAS, Santiago de Compostela, Institute of Excellence on Tissue Engineering
Spain and Regenerative Medicine, Guimarães,
David Grosh Trans-NIH Shared Resource on Portugal; ICVS/3B’s—PT Government
Biomedical Engineering and Physical Science Associate Laboratory, Braga/Guimarães,
(BEPS), National Institute of Biomedical Portugal
Imaging and Bioengineering (NIBIB), Natalie Landon-Brace Department of
National Institutes of Health, Bethesda, MD, Chemical Engineering and Applied
United States Chemistry, Institute for Biomaterials and
Priyanka Gupta Bioprocess and Biochemical Biomedical Engineering, University of
Engineering Group (BioProChem), Toronto, Toronto, ON, Canada
Department of Chemical and Process Simon Latour Department of Chemical
Engineering, University of Surrey, Surrey, Engineering and Applied Chemistry,
United Kingdom Institute for Biomaterials and Biomedical
A. Herreros-Pomares Molecular Oncology Engineering, University of Toronto, Toronto,
Laboratory, General University Hospital of ON, Canada
xiv List of Contributors

Nancy T. Li Department of Chemical Alison P. McGuigan Department of Chemical

Engineering and Applied Chemistry, Engineering and Applied Chemistry,
Institute for Biomaterials and Biomedical Institute for Biomaterials and Biomedical
Engineering, University of Toronto, Toronto, Engineering, University of Toronto, Toronto,
ON, Canada ON, Canada
Maria Angélica Luque-González 3B’s Sibasish Mohanty Institute of Life Sciences,
Research Group, I3Bs Research Institute on Bhubaneswar, India; Regional Center for
Biomaterials, Biodegradables and Biotechnology, Faridabad, India
Biomimetics, University of Minho, Pallavi Mohapatra Institute of Life Sciences,
Headquarters of the European Institute of Bhubaneswar, India; Regional Center for
Excellence on Tissue Engineering and Biotechnology, Faridabad, India
Regenerative Medicine, Guimarães, Portugal; Maria V. Monteiro Department of Chemistry,
ICVS/3B’s PT Government Associate CICECO—Aveiro Institute of Materials,
Laboratory, Braga/Guimarães, Portugal University of Aveiro, Campus Universitário
Qiying Lv Research Center for Tissue de Santiago, Aveiro, Portugal
Engineering and Regenerative Medicine, Joan Montero Nanobioengineering Group,
Union Hospital, Tongji Medical College, Institute for Bioengineering of Catalonia
Huazhong University of Science and (IBEC), The Barcelona Institute of Science and
Technology, Wuhan, P.R. China Technology (BIST), Barcelona, Spain
Tanushree Mahapatra Institute of Life Nicole Y. Morgan Trans-NIH Shared Resource
Sciences, Bhubaneswar, India on Biomedical Engineering and Physical
Science (BEPS), National Institute of
Mahitosh Mandal School of Medical Science
Biomedical Imaging and Bioengineering
and Technology, Indian Institute of
(NIBIB), National Institutes of Health,
Technology Kharagpur, Kharagpur, India
Bethesda, MD, United States
João F. Mano Department of Chemistry,
Antonella Motta BIOtech Research Center,
CICECO—Aveiro Institute of Materials,
Department of Industrial Engineering and
University of Aveiro, Campus Universitário
European Institute of Excellence on Tissue
de Santiago, Aveiro, Portugal
Engineering and Regenerative Medicine,
Alexandra P. Marques 3B’s Research Group, University of Trento, Trento, Italy
I3Bs—Research Institute on Biomaterials, Nuno N. Neves 3B’s Research Group, I3Bs
Biodegradables and Biomimetics, University Research Institute on Biomaterials,
of Minho, Headquarters of the European Biodegradables and Biomimetics, University of
Institute of Excellence on Tissue Engineering Minho, Headquarters of the European Institute
and Regenerative Medicine, Braga/ of Excellence on Tissue Engineering and
Guimarães, Portugal; ICVS/3B’s—PT Regenerative Medicine, Guimarães, Portugal;
Government Associate Laboratory, Braga/ ICVS/3B’s, PT Government Associate
Guimarães, Portugal Laboratory, Braga/Guimarães, Portugal
Albino Martins 3B’s Research Group, I3Bs Catarina Oliveira 3B’s Research Group, I3Bs
Research Institute on Biomaterials, Research Institute on Biomaterials,
Biodegradables and Biomimetics, University Biodegradables and Biomimetics, University
of Minho, Headquarters of the European of Minho, Headquarters of the European
Institute of Excellence on Tissue Engineering Institute of Excellence on Tissue Engineering
and Regenerative Medicine, Guimarães, and Regenerative Medicine, Guimarães,
Portugal; ICVS/3B’s, PT Government Portugal; ICVS/3B’s, PT Government
Associate Laboratory, Braga/Guimarães, Associate Laboratory, Braga/Guimarães,
Portugal Portugal
 List of Contributors xv
Joaquim M. Oliveira 3B’s Research Group, Harumi Ramanayake Tissue Engineering
I3Bs—Research Institute on Biomaterials, Research Group (TERG), Department of
Biodegradables and Biomimetics, University Anatomy and Regenerative Medicine, Royal
of Minho, Headquarters of the European College of Surgeons in Ireland (RCSI),
Institute of Excellence on Tissue Engineering Dublin, Ireland
and Regenerative Medicine, Guimarães, Subha Narayan Rath Department of
Portugal; ICVS/3B’s—PT Government Biomedical Engineering, Indian Institute of
Associated Laboratory, Braga/Guimarães, Technology Hyderabad, Kandi, India
Portugal
Rita Rebelo 3B’s Research Group, I3Bs
Claudia Paindelli David H. Koch Center for Research Institute on Biomaterials,
Applied Research of Genitourinary Cancers, Biodegradables and Biomimetics, University
The University of Texas MD Anderson of Minho, Headquarters of the European
Cancer Center, Houston, TX, United States; Institute of Excellence on Tissue Engineering
Department of Cell Biology, Radboud and Regenerative Medicine, Guimarães,
Institute for Molecular Life Sciences, Portugal; ICVS/3B’s PT Government
Radboud University Medical Center, Associate Laboratory, Braga/Guimarães,
Nijmegen, The Netherlands Portugal
Lara Pierantoni 3B’s Research Group, I3Bs— Rui L. Reis 3B’s Research Group, I3Bs—
Research Institute on Biomaterials, Research Institute on Biomaterials,
Biodegradables and Biomimetics, University Biodegradables and Biomimetics, University
of Minho, Headquarters of the European of Minho, Headquarters of the European
Institute of Excellence on Tissue Engineering Institute of Excellence on Tissue Engineering
and Regenerative Medicine, Guimarães, and Regenerative Medicine, Guimarães,
Portugal; ICVS/3B’s—PT Government Portugal; ICVS/3B’s—PT Government
Associated Laboratory, Braga/Guimarães, Associated Laboratory, Braga/Guimarães,
Portugal Portugal
Ricardo A. Pires 3B’s Research Group, I3Bs J.P. Ribeiro 3B’s Research Group, I3Bs
Research Institute on Biomaterials, Research Institute on Biomaterials,
Biodegradables and Biomimetics, University Biodegradables and Biomimetics, University
of Minho, Headquarters of the European of Minho, Headquarters of the European
Institute of Excellence on Tissue Engineering Institute of Excellence on Tissue Engineering
and Regenerative Medicine, Guimarães, and Regenerative Medicine, Guimarães,
Portugal; ICVS/3B’s - PT Government Portugal; ICVS/3B’s PT Government
Associate Laboratory, Braga/Guimarães, Associate Laboratory, Braga/Guimarães,
Portugal Portugal
Thomas J. Pohida Signal Processing and Claudio Ricci National Interuniversity
Instrumentation Section (SPIS), Consortium of Materials Science and
Computational Bioscience and Engineering Technology (INSTM), Florence, Italy;
Laboratory, Center for Information Department of Civil and Industrial
Technology, National Institutes of Health, Engineering, University of Pisa, Pisa, Italy
Bethesda, MD, United States Robert W. Robey Laboratory of Cell Biology
Luca Primo Department of Oncology, (LCB), Center for Cancer Research, National
University of Torino, Turin, Italy; Candiolo Cancer Institute (NCI), National Institutes of
Cancer Institute FPO-IRCCS, Candiolo, Italy Health, Bethesda, MD, United States
Manashi Priyadarshini Institute of Life Gianpaolo Ruocco University of Basilicata,
Sciences, Bhubaneswar, India; KIIT School of College of Engineering, Modeling and
Biotechnology, KIIT University, Prototyping Laboratory—ModProLab,
Bhubaneswar, India Potenza, Italy
xvi List of Contributors

Josep Samitier Nanobioengineering Group, Eirini Velliou Bioprocess and Biochemical

Institute for Bioengineering of Catalonia Engineering Group (BioProChem),
(IBEC), The Barcelona Institute of Science and Department of Chemical and Process
Technology (BIST), Barcelona, Spain; Engineering, University of Surrey, Surrey,
Department of Electronics and Biomedical United Kingdom
Engineering, University of Barcelona, Marta Viana-Pereira Life and Health Science
Barcelona, Spain; Networking Biomedical Research Institute (ICVS), School of Medicine,
Investigation Center for Bioengineering, University of Minho, Braga, Portugal; ICVS/
Biomaterials and Nanomedicine (CIBER- 3B’s PT Government Associate Laboratory,
BBN), Madrid, Spain Braga/Guimarães, Portugal
Roghayeh Sheervalilou Cellular and Lin Wang Research Center for Tissue
Molecular Research Center, Resistant Engineering and Regenerative Medicine,
Tuberculosis Institute, Zahedan University of Union Hospital, Tongji Medical College,
Medical Sciences, Zahedan, Iran Huazhong University of Science and
Tiago H. Silva 3B’s Research Group, I3Bs Technology, Wuhan, P.R. China; Department
Research Institute on Biomaterials, of Clinical Laboratory, Union Hospital,
Biodegradables and Biomimetics, University Tongji Medical College, Huazhong
of Minho, Headquarters of the European University of Science and Technology,
Institute of Excellence on Tissue Engineering Wuhan, P.R. China
and Regenerative Medicine, Guimarães, Zheng Wang Research Center for Tissue
Portugal; ICVS/3B’s, PT Government Engineering and Regenerative Medicine,
Associate Laboratory, Braga/Guimarães, Union Hospital, Tongji Medical College,
Portugal Huazhong University of Science and
Joana Silva-Correia 3B’s Research Group, Technology, Wuhan, P.R. China; Department
I3Bs—Research Institute on Biomaterials, of Gastrointestinal Surgery, Union Hospital,
Biodegradables and Biomimetics of Tongji Medical College, Huazhong
University of Minho, Headquarters of the University of Science and Technology,
European Institute of Excellence on Tissue Wuhan, P.R. China.
Engineering and Regenerative Medicine,
William J. Wulftange Trans-NIH Shared
Guimarães, Portugal; ICVS/3B’s—PT
Resource on Biomedical Engineering and
Government Associated Laboratory, Braga/
Physical Science (BEPS), National Institute of
Guimarães, Portugal
Biomedical Imaging and Bioengineering
Ana I. Soares 3B’s Research Group, I3Bs— (NIBIB), National Institutes of Health,
Research Institute on Biomaterials, Bethesda, MD, United States; Laboratory of
Biodegradables and Biomimetics, University Cell Biology (LCB), Center for Cancer
of Minho, Headquarters of the European Research, National Cancer Institute (NCI),
Institute of Excellence on Tissue Engineering National Institutes of Health, Bethesda, MD,
and Regenerative Medicine, Braga/ United States
Guimarães, Portugal; ICVS/3B’s—PT
Amir Zarebkohan Department of Medical
Government Associate Laboratory, Braga/
Nanotechnology, Faculty of Advanced
Guimarães, Portugal
Medical Sciences, Tabriz University of
Marianela Vara-Messler Department of Medical Sciences, Tabriz, Iran; Drug Applied
Oncology, University of Torino, Turin, Italy; Research Center, Tabriz University of
Candiolo Cancer Institute FPO-IRCCS, Medical Sciences, Tabriz, Iran
Candiolo, Italy
 Preface
Traditionally, in vitro approaches
employed to model tumor progression
involve the use of tissue culture flasks and
a single cell population. This oversimplified
scenario is in contrast with the dynamic
tumor microenvironment that is observed
in vivo, which is three-dimensional (3D)
and contains multiple cell types. During
the recent years, a new generation of pre-
dictive tumor models has been emerging.
This new paradigm in tumor models com-
bines the advantages of 3D biomaterials
with the most recent advancements in the
fields of tissue engineering, nanotechnol-
ogy, cell biology, biochemistry, 3D bioprint-
ing, and computer simulation. As a result,
more realistic and predictive platform(s)
have been created, defining the roadmap
for their future use as preclinical drug/
therapy screening platforms in cancer
research.
The editors feel very proud to have col-
lected a diverse variety of relevant chapters
from recognized international leaders,
which include their personal and forward-
looking vision in the fields. The chapters
report further on the latest trends of
xvii
biomaterials for 3D cancer modeling, and
in particular, about their uses as physiolog-
ically relevant platforms for drug discovery
and screening as well as for mechanistic
investigations. The editors believe that this
book will be an additional support and sti-
muli for designing new experiments in dif-
ferent research areas, such as biomaterials,
tissue engineering, regenerative medicine,
biomedical devices, pharmacology, and
preclinical experimentations. Furthermore,
this book will be in general helpful to the
material science, biology, and medical sci-
entific communities working in the field of
cancer-related research. Overall, this new
paradigm in 3D in vitro tumor modeling
will univocally improve the field of cancer
diagnosis and treatment.
Finally, we are grateful to all the authors
and co-authors for their contributions in
this book. We are also thankful to the entire
Elsevier team for their continuous support
and help during the edition of this book.
We sincerely acknowledge the financial
support from the project Forefront
Research in 3D disease cancer models as
in vitro screening technologies (FoReCaST),
xviii Preface

which is supported by the European Union The Editors,

Framework Program for Research and
Innovation Horizon 2020 under grant Subhas C. Kundu Rui L. Reis
agreement no. 668983. This ERA-Chairs
widening spreading of excellence project
aims to develop a new generation of 3D
in vitro tumor models for drug screening
applications and has, therefore, motivated
us to edit this unique book. We also sin-
cerely thank all the FoReCaST collaborators
and team-members, namely Drs. Joaquim
M. Oliveira, Vitor M. Correlo, David
Caballero, Banani Kundu, Virginia
Brancato, and Ricardo Pires for their
encouragement during the entire course of August 1st
action for editing this book. 3B’s Research Group, I3Bs-Research
Institute for Biomaterials, Biodegradables,
and Biomimetics, University of Minho,
Guimarães, Portugal
 C H A P T E R

1
Trends in biomaterials for
three-dimensional cancer modeling
David Caballero1,2, Rui L. Reis1,2 and Subhas C. Kundu1,2
1
3B’s Research Group, I3Bs Research Institute on Biomaterials, Biodegradables and
Biomimetics, University of Minho, Headquarters of the European Institute of Excellence on
Tissue Engineering and Regenerative Medicine, Guimarães, Portugal 2ICVS/3B’s PT
Government Associate Laboratory, Braga/Guimarães, Portugal

Abbreviations
2D Two-dimensions
3D Three-dimensions
4D Four-dimensions
ADMET Adsorption, distribution, metabolism, excretion, and toxicity
CAFs Cancer-associated fibroblasts
CDMs Cell-derived matrices
CTCs Circulating tumor cells
ECM Extracellular matrix
EMEA European Medicines Agency
EMT Epithelial-to-mesenchymal transition
EPR Enhanced permeability and retention
FDA Federal drug agency
HA Hyaluronic acid
HUVEC Human umbilical vein endothelial cells
MMPs Matrix metalloproteases
PDMS Polydimethylsiloxane
PEG Poly(ethylene glycol)
TAMs Tumor-associated macrophages
TME Tumor microenvironment
ToC Tumor-on-a-chip
VEGF Vascular endothelial growth factor

Biomaterials for 3D Tumor Modeling

DOI: https://doi.org/10.1016/B978-0-12-818128-7.00001-0 3 © 2020 Elsevier Inc. All rights reserved.
4 1. Trends in biomaterials for three-?dimensional cancer modeling

1.1 A historical introduction

1.1.1 In vitro and in vivo models: an overview
During the last decade, the type of biological assays that are used for extracting informa-
tion about the efficiency of drugs (including cancer-related compounds) have dramatically
changed. The reason is that a large amount of these drugs fail when they are tested in pre-
clinical assays. This is because most pre-clinical drug evaluations rely on simplified in vitro
assays based on flat two-dimensional (2D) surfaces. This type of assay poorly correlates with
the human disease state. Therein, the cells display artificial phenotypes and perturbed gene
expressions. In general, the drugs respond differently than in vivo. Ex vivo (e.g., biopsies)
and in vivo (e.g., animal) models are also employed for drug evaluation. In cancer research,
these models display certain advantages over 2D surfaces, such as a greater biological com-
plexity. This makes the drugs to produce native-like responses. However, ex vivo models
typically lack perfusion and are not representative of the heterogeneity of the tumor. In con-
trast, in vivo (animal) models are highly dynamic systems, but they are very costly, lack the
human immune system, and are ethically controversial. In addition, regardless of the type of
animal model, it is extremely difficult to investigate cellular and physiological interactions on
this type of models. More advanced tumor models are patient-derived xenografts, where a
surgically resected tumor sample of a patient is engrafted into an immunodeficient mice.
However, these models are extremely expensive and time-consuming, they are associated
with ethical concerns, and individual parameters cannot be isolated [1].

1.1.2 A paradigm shift

To resolve the abovementioned issues, a paradigm shift has occurred since the late
1990s about how the cells are studied [2,3]. Indeed, there is a huge difference between cells
cultured on flat surfaces and/or on three-dimensional (3D) environments. Indeed, seminal
works using 3D in vitro models have demonstrated the important differences in the
phenotypes and activities between cells grown in 2D surfaces and 3D cultures. The latter
recapitulate the important interactions between—healthy and malignant—cells and the
surrounding 3D environment [4,5]. As a result, the cancer research community, including
academic researchers, pharma/biotech industries, and clinicians, have started to move
toward the third dimension. In this regard, 2D assays in the form of traditional tissue cul-
ture flasks or Petri dishes, are being replaced by more sophisticated 3D tissue-engineered
cell culture microenvironments. Therein, the cells display several similarities with the
native scenario, and therefore, are considered as physiologically relevant environments
that bridge the gap between obsolete flat materials and the native scenario. In particular,
the early events of cancer progression can closely be reproduced in 3D culture platforms,
where the cells display phenotypes, morphodynamics, and gene expression patterns much
closer to the in vivo physiological microenvironment. In addition, as previously men-
tioned, this is of upmost importance during the development of drugs, where the cells
respond to compounds as they do in vivo.
Finally, the combination of cutting-edge nanotechnologies with advances in materials sci-
ence and tissue engineering tools, have created a powerful toolbox in the field of biomedical

I. Engineering biomaterials for 3D cancer modelling

Another random document with
no related content on Scribd:
 Other Historic Sites in the Park
Cades Cove and
Oconaluftee are the
primary locations of
historic structures in the
national park, but
elsewhere there are a
few interesting buildings
to see.
From Gatlinburg head
south on Airport Road,
which runs into
Cherokee Orchard
Road in the park. Soon
you come to Noah On the way to and from
“Bud” Ogle’s place. Sugarlands you can take side
Ogle and his wife, trips to (below) Mingus Mill, Little
Cindy, started farming Greenbrier School, and Bud
here on 160 hectares Ogle’s place at Roaring Fork.
(400 acres) in 1879. Plan on devoting nearly a full day
Here you can see a log to visit isolated Cataloochee,
house, log barn, and where you can see (right) the
restored tub mill. Caldwell home, schoolhouse,
Palmer Chapel, and several other
South of the Ogle place
structures.
you come to Roaring
Fork Auto Tour. On this
one-way 8-kilometer (5-mile) tour you can see that nature has
reclaimed most of the Roaring Fork community. Among the few
remaining buildings are Jim Bales’ corncrib and barn, plus a log
house that was moved here.
Home for Ephraim
Bales, his wife, and nine
children consisted of
two joined log cabins.
The smaller one was
the kitchen, and in front
of its hearth is a “tater
hole.” Family members
could lift up a floor
board, remove some
potatoes from storage,
and toss them on the
fire to bake. Other
structures here include National Park Service
a corncrib and barn.
A log house and mill are
the only structures that
remain of the many that
belonged to Alfred
Reagan, one of Roaring
Fork’s more talented
residents. He was a
farmer, blacksmith,
preacher, miller,
storekeeper, and
carpenter. His house
was more refined than
most in the Smokies.
The Roaring Fork Auto
Tour road is open from mid-April to mid-November.
In the Oconaluftee Valley just north of the Pioneer Farmstead is
Mingus Mill, built for Abraham Mingus in the 1870s by Sion Thomas
Early. This gristmill, the finest and most advanced in the Smokies,
has a water-powered turbine beneath it. Water flows down a millrace
and flume to the mill, and, when the flume gate is raised, fills the
penstock to power the turbine. The mill has two sets of grinding
stones, one for corn and one for wheat. The mill was in operation
until 1936, reopened for a few months in 1940, and reconditioned by
the Great Smoky Mountains Natural History Association in 1968. It is
open daily from May through October with a miller usually on duty to
explain its workings.
North of Mingus Mill is Smokemont. All that
remains of this small community is the
Oconaluftee Baptist Church, a frame structure
that sits high on a bluff.
Just off Little River Road between Sugarlands
and Tremont is Little Greenbrier School (see
pages 85 and 144). In the summer an interpreter
often is on hand to help children, and adults,
understand what going to school was like in the
Smokies. The road to the school is narrow and
unpaved and not the easiest to negotiate in
inclement weather, so you may want to walk in.
Several buildings are still standing in the isolated
Cataloochee area on the North Carolina side of
the park. They include Palmer Chapel, Beech
Grove School, and the Jarvis Palmer, Hiram
Caldwell, and Steve Woody homes. Most of the
buildings are open, and a ranger is on duty to
answer your questions. The fields are mowed to
maintain the cove effect from early settlement
days. Reaching Cataloochee from the north
means a lengthy trip on unpaved road; from the south it’s a bit
easier. If you have the time, visiting Cataloochee is worth the extra
effort.
 Related Nearby Sites
A number of nearby sites are related in one way or the other to the
history of the Great Smoky Mountains. Here are a few that you might
visit while vacationing in the Smokies:
The arts, crafts, and lifeways of the Cherokees are portrayed by the
tribe at the Qualla Reservation, adjacent to the North Carolina side
of the park. The Museum of the Cherokee Indian displays a
collection of artifacts, and the Oconaluftee Living Indian Village
shows typical early Cherokee life in log structures. The play “Unto
These Hills” tells the story of the Cherokees and their encounters
with Europeans settling in the Smokies and of the forced removal of
most of the tribe to Oklahoma in 1838. About 4,000 Cherokees live
on the Qualla Reservation today.
The Arrowmont School of Arts and Crafts in Gatlinburg has done
much to perpetuate the pottery, weaving, and other skills indicative
of the Smokies people. The school displays and sells objects created
by local artisans.
The Museum of Appalachia in Norris, Tennessee, just north of
Knoxville, has 30 restored pioneer log structures, a representative
farmstead, and more than 200,000 artifacts of mountain life.
The Blue Ridge Parkway, administered by the National Park Service,
has several log houses, a gristmill, a reconstructed farm, and other
early American buildings. Much of the 755-kilometer (469-mile),
parkway, which adjoins Great Smoky Mountains National Park near
Oconaluftee and runs north into Virginia, is quite far from the park,
but some of the historic points of interest are in the southern portion.
The Folk Art Center, at milepost 382, displays traditional crafts of the
Southern Highlands.
 Armchair Explorations
General histories of the Great Smoky Mountains:
Elizabeth Skaggs Bowman, Land of High Horizons, 1938
Carlos C. Campbell, Birth of a National Park, 1960
Michael Frome, Strangers in High Places, 1980
Horace Kephart, Our Southern Highlanders, 1922
Horace Kephart, Journals at Western Carolina University
Robert Lindsay Mason, The Lure of the Great Smokies,
1927
Roderick Peattie, ed., The Great Smokies and the Blue
Ridge, 1943
Laura Thornborough, The Great Smoky Mountains, 1937
Cherokee history:
James Adair, The History of the American Indians, 1775
William Bartram, Travels, 1792
John P. Brown, Old Frontiers, 1938
William H. Gilbert, The Eastern Cherokees, 1943
Henry T. Malone, Cherokees of the Old South, 1956
James Mooney, Myths of the Cherokees, 1900
Charles C. Royce, The Cherokee Nation of Indians, 1887
William L. Smith, The Story of the Cherokees, 1927
Henry Timberlake, Memoirs, 1765
Grace Steele Woodward, The Cherokees, 1963
Other historical works:
W. C. Allen, The Annals of Haywood County, 1935
John Preston Arthur, Western North Carolina, 1914
John C. Campbell, The Southern Highlander and His
Homeland, 1921
Wilma Dykeman, The French Broad, 1955
Allen H. Eaton, Handicrafts of the Southern Highlands,
1937
Paul Fink, “Early Explorers in the Great Smokies,” East
Tennessee Historical Society Bulletin, 1933
Joseph S. Hall, Smoky Mountain Folks and Their Lore,
1960
Joseph S. Hall, Yarns and Tales from the Great Smokies,
1978
Archibald Henderson, The Conquest of the Old
Southwest, 1920
Charles Lanman, Letters from the Alleghany Mountains,
1849
Ruth W. O’Dell, Over the Misty Blue Hills: The Story of
Cocke County, Tennessee, 1950
John Parris, articles in The Asheville Citizen-Times
Randolph Shields, “Cades Cove,” Tennessee Historical
Quarterly, 1965
Randolph Shields, The Cades Cove Story, 1977
Foster A. Sondley, A History of Buncombe County, 1930
Wilbur Zeigler and Ben Grosscup, The Heart of the
Alleghanies, 1883
Robert Woody, “Life on Little Cataloochee,” South Atlantic
Quarterly, 1950
 Index
Numbers in italics refer to photographs, illustrations, or maps.

Abbott, Rhodie, 11, 94-95

Adair, James, 39
American Revolution, 45, 47
Animals, 18, 91-92, 98
Appalachian National Park Association, 114
Appalachian Trail, 122
Arrowmont School of Arts and Crafts, 130, 131, 156
Arthur, John Preston, 83, 84, 86
Attakullakulla, 42, 44
Ayunini (Swimmer), 40-41, 46

Bartram, William, 42, 75

Beck, John, 49, 56, 57
Bell, Hazel, 78
Big Greenbrier Cove, 50, 86
Blount, John Gray, 19
Bohanan, Dave, 135
Boone, Daniel, 45, 58
Boudinot, Elias, 63, 65, 66, 67, 68
Bradley family, 49
Brainerd Mission, 63
Bryson City, 109
Buckley, S. B., 75
Burns, John, 134
Burrell, Elsie, 144

Cable Mill, 152, 153

Cades Cove, 51, 52, 56, 136, 137, 141, 148, 152-53
Caldwell family, George H., 22-23
Caldwell family, Levi, 20, 21
Caldwell family, Lush, 21
Caldwell home, 155
Cameron, Alexander, 45
Cammerer, Arno B., 116-17, 118
Campbell, Aunt Sophie, 58
Cardwell, Columbus “Clum”, 123
Carver, Aden, 6, 140
Cataloochee, 17, 18-29, 155
Champion Coated Paper (Fibre) Company, 98, 119
Chapman, David C., 114, 116, 117, 118
Charlie’s Bunion, 105
Cherokee Indians:
alphabet, 63-64;
Civil War, 74, 79;
community and homelife, 40-41, 43, 63, 150;
Eastern Band (Qualla Reservation), 71, 127-30, 156;
government, 37, 39, 65;
photos, 34, 38, 44, 46, 62, 64, 65, 66, 67, 69, 71, 128;
removal, 65-70;
rituals and religion, 39, 42, 43, 57, 63, 127-29;
 settlement, 17, 19, 35-37, 49-52, 70;
treaties, 45, 47, 68
Chickamauga Indians, 47
Chota, 37, 47
Churches, 26, 56-57, 63, 86-87, 126, 152-53, 154, 155
Civil War, 77-79, 82-83
Civilian Conservation Corps, 114, 121-22, 123, 140
Clark, Dick, 27
Clingman, Thomas Lanier, 76
Clingmans Dome, 18, 76, 103, 105
Collins family, 49
Community life, 26, 27, 29
Conner family, 49
Cove Creek Gap, 118
Crestmont, 98, 102
Crockett, Davy, 67
Cuming, Alexander, 44

Davis, John, 150

Davis, Willis P., 114, 116, 118
Deep Creek, 49-50
DeSoto, Hernando, 36
Dowdle family, Alfred, 126
Dragging Canoe, 47
Duckett, Kate, 81
Dwight Mission, 65

Eakin, J. Ross, 106, 121, 122

Economy, 77, 98-99
Education, 65, 84-87, 85, 130-31, 140-41, 145, 155. See also
Little Greenbrier School House
Elkmont, 17, 98, 99
Enloe family, Abraham, 49, 56, 57
Enloe, Mrs. Clem, 83
Environmental Education Center, 140-41, 145

Farming, 52, 54, 151

Fences, 27, 30-33, 91
Floyd family, 49
Folk Art Center, 156
Folk culture, 57. See also Homelife
Fontana, 98
Forge, iron, 51-52
Forney Creek, 49, 105
Foute, David, 51
French and Indian War, 44-45

Gatlin, Radford, 78-79

Gatlinburg, 50, 78, 123, 130, 148
Geology, 17-18
Gibson, Wiley, 60, 61
Ginatiyun tihi (Stephen Tehee), 69
Gold, 66
Granny’s College, 86
Great Smoky Mountain Conservation Area, 116
Great Smoky Mountains National Park:
 accommodations, 149;
founding, 18, 29, 114, 116-17, 119, 123, 126;
map, 14-15;
nearby sites, 156;
officials, 106, 107, 118, 119;
safety and health, 149;
site, 148;
size, 17;
visitor centers, 148, 149
Great Smoky Mountains Natural History Association, 155
Greenbrier, 17
Gregg-Cable house, 152, 153
Gregory, Russell, 52
Gregory Bald, 52
Guyot, Arnold, 76-77
Guyot, Mount, 102, 105

Handicrafts, 94-95, 131-35, 156

Hearon, Tom and Jerry, 134-35
Higinbotham, Phyllis, 131
Hill, Martha, 130
Homelife, 52, 53-55, 78, 79, 80-81, 83-84, 87, 88-89, 90, 94-95,
150-51
Housing, 4-5, 12-13, 16, 21, 40-41, 91, 136, 150, 152, 153, 154-
55
Huff, Andy, 97, 105
Hughes family, Ralph, 49, 56
Huskey, Claude, 134

Jackson, Andrew, 66-67

Jefferson, Thomas, 65
Job Corps, 140
Jones, John, 134

Kephart, Horace, 107-110, 112, 114, 119, 138;

photo, 108
Kituwah, 36, 37

Lamon, George, 51
Language, 92-93
Lanman, Charles, 75
Little Greenbrier Cove, 123, 125
Little Greenbrier School House, 84, 85, 144, 148, 154
Little River Lumber Company, 97, 99, 117, 140
Love, Robert, 20
Lumber industry, 29, 97-105, 117;
photos, 96, 100-101, 103, 104. See also Little River Lumber
Company
Lyttleton, William Henry, 44

Mabry mill, 156

McCarter, Mack, 132-33, 134
McFalls, Neddy, 27
Marshall, John, 67
Maps, 14-15, 36, 70
Matthews, Herman, 85
Medlin, 108
Meigs, Return Jonathan, 65
Messer family, E.J., 21
Messer, Milas, 90-91, 150
Messer, Will, 27
Mingus, Abraham, 31
Mingus family, John Jacob, 49, 56
Mingus mill, 148, 154-55
Mitchell, Elisha, 76
Mitchell, Mount, 36, 76
Music, 87, 91, 115
Myers, Dan, 26
Myers, Sherman, 31

Newman, Grace, 115

Oakley family, Wiley, 111

Oakley, Becky, 94
Oconaluftee, 49, 56, 148, 150-51
Ogle family, Martha Huskey, 50
Ogle, Mollie McCarter, 48
Ogle family, 88-89
Ogle home, Noah “Bud”, 154
Oliver family, John and Lurena, 33, 51, 52, 136, 152-53
Ownby, Celia, 80
Ownby, Giles and Lenard, 19
Ownby, Humphry John, 86
Ownby, Lillie Whaley, 86
Ownby, Matt, 60
Ownby, Mrs. Matt, 132
Ownby family, Tilman, 142-43

Palmer family, George, 20, 21, 27, 28

Palmer, Lafayette, 21, 27
Palmer, Jesse, 21
Parson’s Bald, 52
Payne, John Howard, 64, 68
Plants, 18, 75; medicinal, 57-58, 84
Proctor, 98
Proffitt, Jim, 115

Qualla Arts and Crafts Mutual, 130

Ravensford, 98
Reagan family, Richard, 50, 78
Ridge, John, 63, 67
Ridge, Major, 67, 68
Rifle, long, 58, 59-61
Roads, 21, 26, 27, 56, 59, 79, 148
Robertson, James, 45
Rockefeller, Jr., John D., 117
Roosevelt, Franklin D., 119, 121
Roosevelt, Theodore, 97, 116
Rose, Aquilla, 110, 112, 113
Ross, John, 65, 66, 68

Schermerhorn, J.D., 68
Scott, Winfield, 68, 71
Sequoyah (George Gist), 63-65;
 portrait, 64
Settlers, white, 42-52, 152
Sherrill family, Samuel, 49, 56
Shields family, Robert, 52
Shults, George Washington, 104
Siler’s Bald, 105
Smokemont, 17, 98
Swaniger, Aaron, 72

Thomas, William, 79
Thomason, W.W., 111, 112
Tipton, William, 51
Tremont, 137-38
Tryon, William, 45
Tsali, 70, 71
Tsiskwa-kaluya (Bird Chopper), 71

Van Buren, Martin, 68

Vance, Zebulon B., 77

Walker, William “Black Bill”, 137-38, 139

Walker, John, 81, 120, 123
Walker, Nancy, 137, 138
Walker sisters, 123-127;
photos, 81, 124
Walini, 62
Welsh, Samson, 128
Whaley family, 50
 Whitepath, 66, 69
Whisky, 91, 111, 112-14
Wiggins family, Abraham, 50
Worcester, Samuel Austin, 63, 65, 67
Work, Hubert, 116

✩ GPO: 1984—421-611/10001
 Handbook 125
The cover photograph was taken by Ed Cooper. The rest of the color
photography, unless otherwise credited, was taken by William A.
Bake of Boone, North Carolina. Nearly all of the black-and-white
photographs come from the files of Great Smoky Mountains National
Park. About half of them were taken in the 1930s for historic
recording purposes by Edouard E. Exline and Charles S. Grossman
on behalf of the National Park Service. Exline was a landscape
architect with the Civilian Conservation Corps and a photographer by
avocation. Grossman was a structural architect for the park who was
in charge of the cultural preservation program. The other
photographers who have been identified are Laura Thornborough,
who resided in the Smokies and wrote the book The Great Smoky
Mountains; Joseph S. Hall, who has studied and written about
linguistics of the Smokies since the 1930s; Harry M. Jennison, a
research botanist from the University of Tennessee who worked in
the park from 1935 to 1940; H.C. Wilburn, a CCC history technician
who collected and purchased artifacts of mountain life; Maurice
Sullivan, a CCC wildlife technician who subsequently became a Park
Service naturalist; Alden Stevens, a museum specialist for the Park
Service; Jim Shelton, husband of one of the Walker sisters, Sarah
Caroline; George Masa, who established the Asheville Photo Service
shortly after World War I; Burton Wolcott; and National Park Service
photographers George A. Grant, Alan Rinehart, Fred R. Bell, M.
Woodbridge Williams, and Clair Burket.
Many of the logging photographs were donated to the park by the
Little River Lumber Company. Most of the photographs of Cherokees
come from the National Anthropological Archives at the Smithsonian
Institution; many of them were taken by James Mooney in the
Smokies area in 1888.
National Park Service
U.S. Department of the Interior
As the Nation’s principal conservation agency, the Department of the
Interior has responsibility for most of our nationally owned public
lands and natural resources. This includes fostering the wisest use
of our land and water resources, protecting our fish and wildlife,
preserving the environmental and cultural values of our national
parks and historical places, and providing for the enjoyment of life
through outdoor recreation. The Department assesses our energy
and mineral resources and works to assure that their development is
in the best interest of all our people. The Department also has a
major responsibility for American Indian reservation communities
and for people who live in Island Territories under U.S.
administration.
 At Home
In the Smokies
ISBN 0—912627-22-0
 Transcriber’s note
In the original captions for photographs/drawings
were often on a separate page from its image. In
these versions, they have been placed under the
image.
Directional words for the phototgraphs, i.e., below,
right, etc. are linked in this version and point to the
correct photographs.
Minor punctuation errors have been changed
without notice. Inconsistencies in hyphenation have
been standardized. Spelling has been retained as
published.
Index entries that were out of alphabetical order has
been corrected. Also, in this version page numbers
have been changed to reflect any captions that have
been moved.
The name Jim Proffitt/Proffit was spelled once each
way in the text. Both spellings have been retained.
The following Printer errors have been changed.
CHANGED FROM TO
“particularly “particularly
Page 17:
signifcant” significant”
“Humphy John “Humphry John
Page 86:
Ownby” Ownby”
“fibers with a “fibers with a
Page 95:
spining” spinning”
Page 158: “Park Assocation” “Park Association”