Cancer progression is known to be accompanied by changes in tissue stiffness. Previous studies have primarily employed immortalized cell lines and 2D hydrogel substrates, which do not recapitulate the 3D tumor niche. How matrix stiffness affects patient-derived cancer cell fate in 3D remains unclear. In this study, we report a matrix metalloproteinase-degradable poly(ethylene-glycol)-based hydrogel platform with brain-mimicking biochemical cues and tunable stiffness (40–26,600 Pa) for 3D culture of patient-derived glioblastoma xenograft (PDTX GBM) cells. Our results demonstrate that decreasing hydrogel stiffness enhanced PDTX GBM cell proliferation, and hydrogels with stiffness 240 Pa and below supported robust PDTX GBM cell spreading in 3D. PDTX GBM cells encapsulated in hydrogels demonstrated higher drug resistance than 2D control, and increasing hydrogel stiffness further enhanced drug resistance. Such 3D hydrogel platforms may provide a valuable tool for mechanistic studies of the role of niche cues in modulating cancer progression for different cancer types.
Impact statement
Cancer progression has been demonstrated to be accompanied by changes in tissue stiffness; however, how matrix stiffness affects patient-derived glioblastoma xenograft glioblastoma (PDTX GBM) cells in 3D remains elusive. By employing a biomimetic hydrogel platform with brain-mimicking biochemical cues and tunable stiffness (40–26,600 Pa), we demonstrated the effect of varying hydrogel stiffness on PDTX GBM cell proliferation, spreading, and drug resistance in 3D, which cannot be recapitulated using 2D culture. Such 3D hydrogel platforms may provide a valuable tool for mechanistic studies or drug discovery and screening using patient-derived GBM cells.
Get full access to this article
View all access options for this article.
References
1.
WellsR.G.The role of matrix stiffness in regulating cell behavior. Hepatology, 47, 1394, 2008.
2.
YehY.T., HurS.S., ChangJ., et al.Matrix stiffness regulates endothelial cell proliferation through septin 9. PLoS One, 7, e46889, 2012.
3.
LoC.M., WangH.B., DemboM., WangY.L., and WangY.L.Cell movement is guided by the rigidity of the substrate. Biophys J, 79, 144, 2000.
4.
YeungT., GeorgesP.C., FlanaganL.A., et al.Effects of substrate stiffness on cell morphology, cytoskeletal structure, and adhesion. Cell Motil Cytoskeleton, 60, 24, 2005.
PolacheckW.J., ZervantonakisI.K., and KammR.D.Tumor cell migration in complex microenvironments. Cell Mol Life Sci, 70, 1335, 2013.
7.
NettiP.A., BerkD.A., SwartzM.A., GrodzinskyA.J., and JainR.K.Role of extracellular matrix assembly in interstitial transport in solid tumors. Cancer Res, 60, 2497, 2000.
8.
KuroiwaT., UekiM., IchikiH., et al. Time Course of Tissue Elasticity and Fluidity in Vasogenic Brain Edema. In: James H.E., Marshall, L.F., Reulen, H.J., et al. eds. Brain Edema X. Acta Neurochirurgica Supplements. Vienna, Springer-Verlag Wien, 1997, p. 87.
9.
UlrichT.A., de Juan PardoE.M., and KumarS.The mechanical rigidity of the extracellular matrix regulates the structure, motility, and proliferation of glioma cells. Cancer Res, 69, 4167, 2009.
AnanthanarayananB., KimY., and KumarS.Elucidating the mechanobiology of malignant brain tumors using a brain matrix-mimetic hyaluronic acid hydrogel platform. Biomaterials, 32, 7913, 2011.
12.
GrundyT.J., De LeonE., GriffinK.R., et al.Differential response of patient-derived primary glioblastoma cells to environmental stiffness. Sci Rep, 6, 23353, 2016.
13.
KaufmanL.J., BrangwynneC.P., KaszaK.E., et al.Glioma expansion in collagen I matrices: analyzing collagen concentration-dependent growth and motility patterns. Biophys J, 89, 635, 2005.
14.
PedronS., and HarleyB.A.Impact of the biophysical features of a 3D gelatin microenvironment on glioblastoma malignancy. J Biomed Mater Res A, 101, 3404, 2013.
15.
HughesC.S., PostovitL.M., and LajoieG.A.Matrigel: a complex protein mixture required for optimal growth of cell culture. Proteomics, 10, 1886, 2010.
16.
LutolfM.P., and HubbellJ.A.Synthetic biomaterials as instructive extracellular microenvironments for morphogenesis in tissue engineering. Nat Biotechnol, 23, 47, 2005.
17.
NemirS., and WestJ.L.Synthetic materials in the study of cell response to substrate rigidity. Ann Biomed Eng, 38, 2, 2010.
18.
MiroshnikovaY.A., JorgensD.M., SpirioL., AuerM., Sarang-SieminskiA.L., and WeaverV.M.Engineering strategies to recapitulate epithelial morphogenesis within synthetic three-dimensional extracellular matrix with tunable mechanical properties. Phys Biol, 8, 026013, 2011.
19.
WangC., TongX., and YangF.Bioengineered 3D brain tumor model to elucidate the effects of matrix stiffness on glioblastoma cell behavior using PEG-based hydrogels. Mol Pharm, 11, 2115, 2014.
20.
WangC., TongX., JiangX., and YangF.Effect of matrix metalloproteinase-mediated matrix degradation on glioblastoma cell behavior in 3D PEG-based hydrogels. J Biomed Mater Res A, 105, 770, 2017.
21.
LoessnerD., StokK.S., LutolfM.P., HutmacherD.W., ClementsJ.A., and RizziS.C.Bioengineered 3D platform to explore cell-ECM interactions and drug resistance of epithelial ovarian cancer cells. Biomaterials, 31, 8494, 2010.
22.
SinghS.P., SchwartzM.P., TokudaE.Y., et al.A synthetic modular approach for modeling the role of the 3D microenvironment in tumor progression. Sci Rep, 5, 17814, 2015.
23.
HuszthyP.C., DaphuI., NiclouS.P., et al.In vivo models of primary brain tumors: pitfalls and perspectives. Neuro-Oncology, 14, 979, 2012.
24.
LiA., WallingJ., KotliarovY., et al.Genomic changes and gene expression profiles reveal that established glioma cell lines are poorly representative of primary human gliomas. Mol Cancer Res, 6, 21, 2008.
25.
RidkyT.W., ChowJ.M., WongD.J., and KhavariP.A.Invasive three-dimensional organotypic neoplasia from multiple normal human epithelia. Nat Med, 16, 1450, 2010.
26.
MahesparanR., ReadT.A., Lund-JohansenM., SkaftnesmoK.O., BjerkvigR., and EngebraatenO.Expression of extracellular matrix components in a highly infiltrative in vivo glioma model. Acta Neuropathol, 105, 49, 2003.
27.
BignerS.H., ScholdS.C., FriedmanH.S., MarkJ., and BignerD.D.Chromosomal composition of malignant human gliomas through serial subcutaneous transplantation in athymic mice. Cancer Genet Cytogenet, 40, 111, 1989.
28.
JooK.M., KimJ., JinJ., et al.Patient-specific orthotopic glioblastoma xenograft models recapitulate the histopathology and biology of human glioblastomas in situ. Cell Rep, 3, 260, 2013.
29.
HeffernanJ.M., and SirianniR.W.Modeling Microenvironmental Regulation of Glioblastoma Stem Cells: a biomaterials perspective. Front Mater, 5, 7, 2018.
30.
BahadurS., SahuA.K., BaghelP., and SahaS.Current promising treatment strategy for glioblastoma multiform: a review. Oncol Rev, 13, 417, 2019.
31.
FairbanksB.D., SchwartzM.P., HaleviA.E., NuttelmanC.R., BowmanC.N., and AnsethK.S.A versatile synthetic extracellular matrix mimic via thiol-norbornene photopolymerization. Adv Mater Weinheim, 21, 5005, 2009.
AndersonS.B., LinC.C., KuntzlerD.V., and AnsethK.S.The performance of human mesenchymal stem cells encapsulated in cell-degradable polymer-peptide hydrogels. Biomaterials, 32, 3564, 2011.
34.
DelpechB., MaingonnatC., GirardN., et al.Hyaluronan and hyaluronectin in the extracellular matrix of human brain tumour stroma. Eur J Cancer, 29A, 1012, 1993.
35.
BignerS.H., HumphreyP.A., WongA.J., et al.Characterization of the epidermal growth factor receptor in human glioma cell lines and xenografts. Cancer Res, 50, 8017, 1990.
36.
WongA.J., RuppertJ.M., BignerS.H., et al.Structural alterations of the epidermal growth factor receptor gene in human gliomas. Proc Natl Acad Sci USA, 89, 2965, 1992.
37.
WangC., LiJ., SinhaS., PetersonA., GrantG.A., and YangF.Mimicking brain tumor-vasculature microanatomical architecture via co-culture of brain tumor and endothelial cells in 3D hydrogels. Biomaterials, 202, 35, 2019.
38.
RaoS.S., DejesusJ., ShortA.R., OteroJ.J., SarkarA., and WinterJ.O.Glioblastoma behaviors in three-dimensional collagen-hyaluronan composite hydrogels. ACS Appl Mater Interfaces, 5, 9276, 2013.
39.
ConradB., HanL.H., and YangF.Gelatin-Based Microribbon Hydrogels Accelerate Cartilage Formation by Mesenchymal Stem Cells in Three Dimensions. Tissue Eng Part A, 24, 1631, 2018.
40.
LeeS., TongX., HanL.H., BehnA., and YangF.Winner of the Young Investigator Award of the Society for Biomaterials (USA) for 2016, 10th World Biomaterials Congress, May 17–22, 2016, Montreal QC,Canada: aligned microribbon-like hydrogels for guiding three-dimensional smooth muscle tissue regeneration. J Biomed Mater Res A 104, 1064, 2016.
41.
WangT., LaiJ.H., and YangF.Effects of hydrogel stiffness and extracellular compositions on modulating cartilage regeneration by mixed populations of stem cells and chondrocytes in vivo. Tissue Eng Part A, 22, 1348, 2016.
42.
Jiguet JiglaireC., Baeza-KalleeN., DenicolaïE., et al.Ex vivo cultures of glioblastoma in three-dimensional hydrogel maintain the original tumor growth behavior and are suitable for preclinical drug and radiation sensitivity screening. Exp Cell Res, 321, 99, 2014.
43.
WuC.X., LinG.S., LinZ.X., ZhangJ.D., LiuS.Y., and ZhouC.F.Peritumoral edema shown by MRI predicts poor clinical outcome in glioblastoma. World J Surg Oncol, 13, 97, 2015.
44.
RapeA., AnanthanarayananB., and KumarS.Engineering strategies to mimic the glioblastoma microenvironment. Adv Drug Deliv Rev, 79–80, 172, 2014.
45.
CaliariS.R., and BurdickJ.A.A practical guide to hydrogels for cell culture. Nat Methods, 13, 405, 2016.
46.
BerryR., JowittT.A., FerrandJ., et al.Role of dimerization and substrate exclusion in the regulation of bone morphogenetic protein-1 and mammalian tolloid. Proc Natl Acad Sci USA, 106, 8561, 2009.
47.
da SilvaR., UnoM., MarieS.K., and Oba-ShinjoS.M.LOX expression and functional analysis in astrocytomas and impact of IDH1 mutation. PLoS One, 10, e0119781, 2015.
48.
PiretJ.P., MottetD., RaesM., and MichielsC.Is HIF-1alpha a pro- or an anti-apoptotic protein. Biochem Pharmacol, 64, 889, 2002.
49.
HuangW.J., ChenW.W., and ZhangX.Glioblastoma multiforme: effect of hypoxia and hypoxia inducible factors on therapeutic approaches. Oncol Lett, 12, 2283, 2016.
50.
HardeeM.E., and ZagzagD.Mechanisms of glioma-associated neovascularization. Am J Pathol, 181, 1126, 2012.
51.
CuddapahV.A., RobelS., WatkinsS., and SontheimerH.A neurocentric perspective on glioma invasion. Nat Rev Neurosci, 15, 455, 2014.
52.
PedronS., BeckaE., and HarleyB.A.Spatially gradated hydrogel platform as a 3D engineered tumor microenvironment. Adv Mater Weinheim, 27, 1567, 2015.
53.
SinghS.P., SchwartzM.P., LeeJ.Y., FairbanksB.D., and AnsethK.S.A peptide functionalized poly(ethylene glycol) (PEG) hydrogel for investigating the influence of biochemical and biophysical matrix properties on tumor cell migration. Biomater Sci, 2, 1024, 2014.
54.
OsswaldM., JungE., SahmF., et al.Brain tumour cells interconnect to a functional and resistant network. Nature, 528, 93, 2015.
55.
CoxT.R., and ErlerJ.T.Remodeling and homeostasis of the extracellular matrix: implications for fibrotic diseases and cancer. Dis Model Mech, 4, 165, 2011.
56.
Viapiano,. M.S., and Lawler, S.E. Glioma Invasion: Mechanisms and Therapeutic Challenges. In: MeirE., ed. CNS Cancer: Models, Markers, Prognostic Factors, Targets, and Therapeutic Approaches. New York City, Humana Press, 2009, pp. 1219–1252.
57.
RaoJ.S.Molecular mechanisms of glioma invasiveness: the role of proteases. Nat Rev Cancer, 3, 489, 2003.
58.
GladsonC.L.The extracellular matrix of gliomas: modulation of cell function. J Neuropathol Exp Neurol, 58, 1029, 1999.
59.
FriedmanH.S., KerbyT., and CalvertH.Temozolomide and treatment of malignant glioma. Clin Cancer Res, 6, 2585, 2000.
60.
RoosW.P., BatistaL.F., NaumannS.C., et al.Apoptosis in malignant glioma cells triggered by the temozolomide-induced DNA lesion O6-methylguanine. Oncogene, 26, 186, 2007.
61.
AgarwalaS.S., and KirkwoodJ.M.Temozolomide, a novel alkylating agent with activity in the central nervous system, may improve the treatment of advanced metastatic melanoma. Oncologist, 5, 144, 2000.
62.
EdmondsonR., BroglieJ.J., AdcockA.F., and YangL.Three-dimensional cell culture systems and their applications in drug discovery and cell-based biosensors. Assay Drug Dev Technol, 12, 207, 2014.
63.
HeW., KuangY., XingX., et al.Proteomic comparison of 3D and 2D glioma models reveals increased HLA-E expression in 3D models is associated with resistance to NK cell-mediated cytotoxicity. J Proteome Res, 13, 2272, 2014.
64.
ChaudhuriO., GuL., KlumpersD., et al.Hydrogels with tunable stress relaxation regulate stem cell fate and activity. Nat Mater, 15, 326, 2016.
65.
WisdomK.M., AdebowaleK., ChangJ., et al.Matrix mechanical plasticity regulates cancer cell migration through confining microenvironments. Nat Commun, 9, 4144. 2018.
