title: Flow-based pipeline for systematic modulation and analysis of 3D tumor microenvironments.
abstract: The cancer microenvironment, which incorporates interactions with stromal cells, extracellular matrix (ECM), and other tumor cells in a 3-dimensional (3D) context, has been implicated in every stage of cancer development, including growth of the primary tumor, metastatic spread, and response to treatment. Our understanding of the tumor microenvironment and our ability to develop new therapies would greatly benefit from tools that allow us to systematically probe microenvironmental cues within a 3D context. Here, we leveraged recent advances in microfluidic technology to develop a platform for high-throughput fabrication of tunable cellular microniches ("microtissues") that allow us to probe tumor cell response to a range of microenvironmental cues, including ECM, soluble factors, and stromal cells, all in 3D. We further combine this tunable microniche platform with rapid, flow-based population level analysis (n > 500), which permits analysis and sorting of microtissue populations both pre- and post-culture by a range of parameters, including proliferation and homotypic or heterotypic cell density. We used this platform to demonstrate differential responses of lung adenocarcinoma cells to a selection of ECM molecules and soluble factors. The cells exhibited enhanced or reduced proliferation when encapsulated in fibronectin- or collagen-1-containing microtissues, respectively, and they showed reduced proliferation in the presence of TGF-β, an effect that we did not observe in monolayer culture. We also measured tumor cell response to a panel of drug targets and found, in contrast to monolayer culture, specific sensitivity of tumor cells to TGFβR2 inhibitors, implying that TGF-β has an anti-proliferative affect that is unique to the 3D context and that this effect is mediated by TGFβR2. These findings highlight the importance of the microenvironmental context in therapeutic development and that the platform we present here allows the high-throughput study of tumor response to drugs as well as basic tumor biology in well-defined microenvironmental niches.
authors.affiliation: Chemical Engineering, Massachusetts Institute of Technology, Cambridge, Massachusetts 02139, United States.
authors.forename: Cheri Y
authors.initials: CY
authors.lastname: Li
authors.forename: David K
authors.initials: DK
authors.lastname: Wood
authors.forename: Joanne H
authors.initials: JH
authors.lastname: Huang
authors.forename: Sangeeta N
authors.initials: SN
authors.lastname: Bhatia
mesh_terms: D000818:Animals; D018929:Cell Culture Techniques / Q000295:instrumentation / Q000379:methods*; D045744:Cell Line, Tumor; D003094:Collagen / Q000494:pharmacology; D005109:Extracellular Matrix / Q000378:metabolism; D005353:Fibronectins / Q000494:pharmacology; D006801:Humans; D051379:Mice; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D017346:Protein Serine-Threonine Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism; D051381:Rats; D000077294:Receptor, Transforming Growth Factor-beta Type II; D018125:Receptors, Transforming Growth Factor beta / Q000037:antagonists & inhibitors / Q000378:metabolism; D017154:Stromal Cells / Q000166:cytology / Q000187:drug effects; D016212:Transforming Growth Factor beta / Q000494:pharmacology; D059016:Tumor Microenvironment
chemical_list: D005353:Fibronectins; D047428:Protein Kinase Inhibitors; D018125:Receptors, Transforming Growth Factor beta; D016212:Transforming Growth Factor beta; D003094:Collagen; D017346:Protein Serine-Threonine Kinases; D000077294:Receptor, Transforming Growth Factor-beta Type II
references.citation: Nat Commun. 2012;3:1122
references.pmid: 23047680
references.citation: Lab Chip. 2010 Sep 21;10(18):2424-32
references.pmid: 20694216
references.citation: Br J Cancer. 2005 Nov 14;93(10):1157-67
references.pmid: 16251876
references.citation: Lab Chip. 2012 Sep 21;12(18):3277-80
references.pmid: 22824954
references.citation: Lab Chip. 2008 Aug;8(8):1262-4
references.pmid: 18651066
references.citation: Cancer Metastasis Rev. 1994 Jun;13(2):121-38
references.pmid: 7923546
references.citation: Nat Rev Cancer. 2009 Apr;9(4):285-93
references.pmid: 19308068
references.citation: Lab Chip. 2008 Jul;8(7):1056-61
references.pmid: 18584079
references.citation: Proc Natl Acad Sci U S A. 1998 Dec 8;95(25):14821-6
references.pmid: 9843973
references.citation: Curr Opin Cell Biol. 2003 Oct;15(5):565-71
references.pmid: 14519391
references.citation: Lab Chip. 2009 Sep 21;9(18):2628-31
references.pmid: 19704976
references.citation: Science. 1993 May 28;260(5112):1335-8
references.pmid: 8388126
references.citation: Annu Rev Pathol. 2006;1:119-50
references.pmid: 18039110
references.citation: Biotechnol J. 2008 Oct;3(9-10):1172-84
references.pmid: 18566957
references.citation: Adv Drug Deliv Rev. 2004 Sep 22;56(11):1635-47
references.pmid: 15350293
references.citation: Nature. 2012 Jul 26;487(7408):500-4
references.pmid: 22763439
references.citation: Acta Biomater. 2011 Sep;7(9):3267-76
references.pmid: 21704198
references.citation: Mol Cell Biol. 1995 Jun;15(6):3273-81
references.pmid: 7760823
references.citation: Trends Cell Biol. 2010 Dec;20(12):705-14
references.pmid: 20965727
references.citation: Proc Natl Acad Sci U S A. 2009 Jan 13;106(2):399-404
references.pmid: 19126683
references.citation: Nature. 2011 May 5;473(7345):101-4
references.pmid: 21471965
references.citation: Cell. 2003 Jul 11;114(1):33-45
references.pmid: 12859896
references.citation: Cell. 2011 Nov 23;147(5):992-1009
references.pmid: 22118458
references.citation: Cancer Lett. 1998 Sep 11;131(1):29-34
references.pmid: 9839617
references.citation: Proc Natl Acad Sci U S A. 1994 Dec 20;91(26):12378-82
references.pmid: 7528920
references.citation: Annu Int Conf IEEE Eng Med Biol Soc. 2009;2009:7037-40
references.pmid: 19964195
references.citation: Cell. 2006 Jan 27;124(2):263-6
references.pmid: 16439202
references.citation: Nat Rev Cancer. 2006 Jul;6(7):506-20
references.pmid: 16794634
references.citation: Cell. 2011 Oct 14;147(2):275-92
references.pmid: 22000009
references.citation: Clin Exp Metastasis. 2002;19(2):161-8
references.pmid: 11964080
references.citation: Biomaterials. 1998 Jul;19(14):1287-94
references.pmid: 9720892
references.citation: Nat Methods. 2007 Oct;4(10):855-60
references.pmid: 17767164
references.citation: Cell Res. 2009 Jan;19(1):89-102
references.pmid: 19050696
references.citation: Nat Rev Cancer. 2009 Apr;9(4):239-52
references.pmid: 19279573
references.citation: Lab Chip. 2011 Sep 7;11(17):2967-75
references.pmid: 21776518
references.citation: Biomaterials. 2009 Feb;30(6):1113-20
references.pmid: 19046762
references.citation: Biochim Biophys Acta. 1997 Oct 24;1333(2):F105-50
references.pmid: 9395284
references.citation: Am J Respir Cell Mol Biol. 2008 Jan;38(1):95-104
references.pmid: 17673689
references.citation: Biomaterials. 2006 Nov;27(31):5391-8
references.pmid: 16828863
references.citation: Nat Rev Drug Discov. 2004 Dec;3(12):1011-22
references.pmid: 15573100
references.citation: Integr Biol (Camb). 2011 Jun;3(6):653-62
references.pmid: 21526262
references.citation: Integr Biol (Camb). 2010 Oct;2(10):517-27
references.pmid: 20820630
references.citation: Endocr Rev. 1996 Jun;17(3):207-20
references.pmid: 8771356
references.citation: Curr Opin Biotechnol. 2011 Oct;22(5):655-60
references.pmid: 21306888
references.citation: Biomaterials. 2007 Jan;28(2):256-70
references.pmid: 16979755
references.citation: Lung Cancer. 2003 Jul;41(1):65-70
references.pmid: 12826314
references.citation: Proc Natl Acad Sci U S A. 2007 Dec 26;104(52):20938-43
references.pmid: 18087037
references.citation: Cancer Metastasis Rev. 1995 Sep;14(3):173-89
references.pmid: 8548867