E-Cadherin and EpCAM expression by NSCLC tumour cells associate with normal fibroblast activation through a pathway initiated by integrin αvβ6 and maintained through TGFβ signalling.
Fibroblasts in the tumour stroma (cancer-associated fibroblasts) influence tumour progression and response to therapeutics; little is known about the mechanisms through which the tumour cell co-opts a normal fibroblast. To study the activation of fibroblasts by tumour cells, a panel of non-small cell lung cancer (NSCLC) cell lines and normal human dermal fibroblasts were co-cultured. A subset of the NSCLC cells induced an activated cancer-associated fibroblast-like fibroblast phenotype defined by induction of fibroblast α-smooth muscle actin expression. Tumour cells that activated fibroblasts were associated with E-Cadherin and EpCAM expression and expression of integrin αvβ6. Co-culture of activating tumour cells with fibroblasts resulted in induction of transcripts associated with tumour cell invasion and growth, TGFβ1 and TGFBR1, SERPINE-1, BMP6, SPHK1 and MMP9. Fibroblast activation was inhibited by an αvβ6/8 integrin blocking antibody (264RAD) and a small molecule inhibitor of the TGF-beta type I receptor activin-like kinase (ALK5) (SB431542), demonstrating that transactivation of the TGFβ pathway initiates fibroblast activation. Both integrin and ALK5 antagonists inhibited initiation. Only ALK5 was effective when added after 3 days of co-culture. This suggests that although activation is αvβ6-dependent, once fibroblasts are activated alternative TGFβ pathway regulators maintain an activation loop. In co-culture activating cells had reduced sensitivity to selumetinib, AZD8931 and afatinib compared with mono-culture. In contrast, non-activating cells were insensitive to selumetinib and AZD8931 in both mono-culture and co-culture. In conclusion NSCLC cell lines, positive for E-Cadherin, EpCAM and αvβ6 expression, activate normal fibroblasts through avβ6/TGFβ signalling in vitro, and influence both gene expression and response to therapeutic agents.
Oncology iMED, AstraZeneca, Alderley Park, Macclesfield, UK.
C
C
Eberlein
Oncology iMED, AstraZeneca, Alderley Park, Macclesfield, UK.
C
C
Rooney
Oncology iMED, AstraZeneca, Alderley Park, Macclesfield, UK.
S J
SJ
Ross
Oncology iMED, AstraZeneca, Alderley Park, Macclesfield, UK.
M
M
Farren
Oncology iMED, AstraZeneca, Alderley Park, Macclesfield, UK.
H M
HM
Weir
Oncology iMED, AstraZeneca, Alderley Park, Macclesfield, UK.
S T
ST
Barry
D000077716:Afatinib; D000951:Antigens, Neoplasm / Q000096:biosynthesis* / Q000235:genetics*; D015820:Cadherins / Q000096:biosynthesis* / Q000235:genetics; D002289:Carcinoma, Non-Small-Cell Lung / Q000188:drug therapy / Q000235:genetics* / Q000473:pathology; D015815:Cell Adhesion Molecules / Q000096:biosynthesis* / Q000235:genetics; D018920:Coculture Techniques; D000071858:Epithelial Cell Adhesion Molecule; D005347:Fibroblasts / Q000378:metabolism; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects; D006801:Humans; D016023:Integrins / Q000096:biosynthesis / Q000235:genetics*; D009363:Neoplasm Proteins / Q000096:biosynthesis; D011799:Quinazolines / Q000008:administration & dosage; D015398:Signal Transduction / Q000187:drug effects; D017154:Stromal Cells / Q000378:metabolism / Q000473:pathology; D016212:Transforming Growth Factor beta / Q000235:genetics* / Q000378:metabolism
C548875:AZD 8931; D000951:Antigens, Neoplasm; D015820:Cadherins; D015815:Cell Adhesion Molecules; C499379:EPCAM protein, human; D000071858:Epithelial Cell Adhesion Molecule; D016023:Integrins; D009363:Neoplasm Proteins; D011799:Quinazolines; D016212:Transforming Growth Factor beta; C073878:integrin alphavbeta6; D000077716:Afatinib