KEYWORDS: Photodynamic therapy, Receptors, Cell death, Proteins, Simulation of CCA and DLA aggregates, Cancer, Modulation, In vitro testing, Resistance, Confocal microscopy
EGFR, a member of the ERBB family, plays a pivotal role in carcinogenesis. EGFR overexpression is
implicated in DNA repair and synergistic interactions between EGFR-targeting drugs and conventional
chemo/radiotherapy have been reported in preclinical studies for different cancers but not
cholangiocarcinoma (CCA). To date there are no in vitro data available on the cellular response and
effect of either photodynamic therapy (PDT) or EGFR-targeting drugs on CCA. Therefore, we aimed
to study the: (i) response to Verteporfin PDT and to EGFR-targeting drugs, as single agents; (ii) effect
of PDT on ERBBs expression, phosporylation status and activation of its signaling pathways; (iii)
response to combination of PDT and EGFR-targeting agents.
We showed that two cholangiocarcinoma cell lines (HuCCT1 and TFK1 cells, intra- and extrahepatic,
respectively) differentially respond to verteporfin-PDT treatment and are resistant to EGFR-targeting
agents. A constitutive activation of EGFR in both cell lines was also observed, which could partly
account for the observed resistance to EGFR-targeting drugs. In addition, verteporfin-PDT induced
further phosphorylation of both EGFR and other Receptor Tyrosine Kinases. Mitochondria-independent
apoptosis was induced by PDT in both CCA cell lines; in particular, PDT modulated the
expression of members of the Inhibitor of Apoptosis (IAP) family of proteins. Interestingly, there was a
PDT-induced EGFR nuclear translocation in both cell lines; co-treatment with either an EGFR-inhibitor
(Cetuximab) or a nuclear import blocking agent (Wheat Germ Agglutinin) had an additive
effect on PDT cell killing, thus implying a role of EGFR in repairing the potential PDT-induced DNA
damage.
KEYWORDS: Simulation of CCA and DLA aggregates, Photodynamic therapy, Receptors, Cancer, Tumors, In vitro testing, Proteins, Solids, Therapeutic agents, Oncology
Accumulating preclinical and clinical evidence supports a pro-oncogenic function for Notch signaling in several solid
tumors. Therefore, Notch inhibitory agents, such as gamma-secretase inhibitors (GSI), are being investigated as cancer
therapeutic agents and a potential adjuvant to conventional chemo/radiotherapy. To date, no in vitro data are available on
the cellular response and effect of either photodynamic therapy (PDT) or GSI on human cholangiocarcinoma (CCA).
Consequently, we aimed to study the: (i) constitutive expression of Notch signaling pathway in CCA cell lines; (ii)
response to Verteporfin-PDT and to GSI, as single agents on CCA cell lines; (iii) effect of Verteporfin-PDT on Notch
signaling pathway expression.
Expression of Notch signaling components was studied in two cholangiocarcinoma cell lines, HuCCT1 and
TFK-1 (intra- and extrahepatic, respectively). No difference in basal expression of Notch1, 2 and Jagged1 was observed
in either cell line. In contrast, Notch3 was found to be weakly and highly expressed in HuCCT1 and TFK-1 cells,
respectively - supporting our recent microarray data which showed Notch3 overexpression in biliary brushings from
patients with extrahepatic CCA. HuCCT1 and TFK-1 differentially responded to Verteporfin-PDT treatment; preliminary
data showed no clear effect of GSI on proliferation/apoptosis in either cell line following short exposure (6 and 24h).
Following Verteporfin-PDT, Notch1, 2 and Jagged-1 expression was down-regulated in both cell lines, while Notch3
expression was unaffected in HuCCT1 cells and down-regulated in TFK-1 cells. The Notch signaling pathway could
represent a potential target for combination therapy in CCA treatment.
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