The role of DDR1 ectodomain shedding in glioblastoma radiation resistance

Introduction: Glioblastoma (GBM) is the most common malignant brain tumor in adults, and resistance to radiation therapy remains a major therapeutic obstacle. Although genetic alterations contribute to GBM progression, they do not fully explain the emergence of radiation resistance, suggesting contributions from non-genetic mechanisms. Notably, radiation-resistant GBM exhibits elevated collagen deposition. Discoidin domain receptor tyrosine kinase 1 (DDR1), a collagen-binding receptor regulated by ectodomain shedding, is highly expressed in GBM. Here, we demonstrate that radiation-resistant GBM alters DDR1 activity and shedding in response to collagen to promote tumor cell proliferation. Methods and Results: To investigate DDR1 regulation, we utilized a GBM patient-derived xenoline pair, JX39P (radiation-sensitive) and JX39P-RT (radiation-resistant). JX39P-RT displayed significantly higher DDR1 mRNA and protein expression. When cells were embedded in collagen I to mimic the extracellular environment, DDR1 phosphorylation at Y792 increased more rapidly and to a greater extent in JX39P-RT than in JX39P. Collagen embedment also led to the accumulation of a short DDR1 fragment in JX39P-RT, indicating enhanced ectodomain shedding. Western blotting of conditioned media confirmed increased release of the DDR1 N-terminal fragment and confirmed that ectodomain shedding occurred only in JX39P-RT in response to collagen. Treatment of JX39P-RT cells with marimastat, a broad-spectrum MMP inhibitor, diminished both the N-terminal fragment shed in the media and the short C-terminal fragment in cell lysates. Consistent with elevated DDR1 expression and activation, JX39P-RT exhibited increased phospho-ERK1/2 levels when embedded in collagen I, indicating enhanced downstream signaling pathways. Furthermore, cell proliferation assays and Ki67 staining showed that JX39P cells proliferate more rapidly than JX39P-RT in liquid culture; however, when embedded in collagen I, JX39P showed no significant change in Ki67 staining, whereas JX39P-RT demonstrated a time-dependent increase, indicating that radiation-resistant GBM cells proliferate in response to collagen- and DDR1-mediated signaling. Conclusions: Together, our results identify DDR1 upregulation and ectodomain shedding as key adaptive mechanisms that promote proliferation and survival of radiation-resistant GBM cells. These regulatory processes enable resistant cells to sustain growth under therapeutic stress, providing a potential mechanism of therapeutic evasion. Future studies will investigate how DDR1 shedding and downstream signaling integrate to maintain radiation resistance and explore strategies to sensitize GBM to radiation therapy.