SRPK Inhibition in Cholangiocarcinoma Cells Induces LAMB1-Associated Adaptive Responses and Enhances Growth Suppression by the MEK Inhibitor U0126 via the LAMB1/ERK/c-JUN Axis

Background: Cholangiocarcinoma (CCA) is an aggressive bile duct malignancy with a high incidence and mortality rate in Southeast Asia, particularly in Thailand. However, current treatments offer only limited clinical benefit. Aberrant mRNA splicing, driven by hyperactivation of serine/arginine-rich splicing factors (SRSFs) and their upstream kinases SRPK1 and SRPK2, contributes to the progression of CCA. Although SRPK inhibitors suppress tumor growth, their effects remain incomplete, highlighting the need for rational combination strategies. Methods: Liquid chromatography–tandem mass spectrometry (LC-MS/MS)-based proteomic profiling was performed on CCA cell lines (KKU-213A and KKU-055) treated with the SRPK inhibitors SRPIN340 and SPHINX31. Subsequently, commonly upregulated differentially expressed proteins (UpDEPs) were identified and cross-validated with data from the Cancer Genome Atlas (TCGA) using GEPIA, followed by reverse transcription-quantitative polymerase chain reaction (qRT-qPCR) validation. The functional effects of combined SRPK and mitogen-activated protein kinase kinase (MEK) inhibition (with U0126) were evaluated using cell viability and colony formation assays, and by analyzing signaling pathways. Results: A total of 34 common UpDEPs were identified following SRPK inhibition. Laminin subunit beta 1 (LAMB1) was prioritized as a candidate combination target due to robust upregulation, concordant overexpression in TCGA datasets, and increased mRNA levels after SRPK inhibitor treatment. Given prior evidence linking MEK signaling to LAMB1 regulation, we also evaluated the MEK inhibitor U0126. Combined treatment with SRPK inhibitors (SRPIN340 or SPHINX31) and U0126 produced greater suppression of viability and clonogenic growth in KKU-213A cells than either treatment alone. These effects were accompanied by reduced ERK phosphorylation and cellular Jun proto-oncogene (c-JUN) expression. Conclusions: LAMB1 represents a potential adaptive response target following SRPK inhibition in CCA. Dual targeting of SRPK and MEK signaling pathways exerts greater antitumor effects than either treatment alone and may provide a promising therapeutic strategy for CCA management.