Classical Hodgkin's lymphoma (cHL) is a B cell lymphoma characterized by a minority of malignant Hodgkin Reed-Sternberg (HRS) cells that are surrounded by a vast majority of reactive cells. These reactive cells provide a microenvironment that supports survival and growth of the HRS cells. The interaction between cancer cells and microenvironment includes aberrant activity of multiple receptor tyrosine kinases (RTK). The insulin-like growth factor-1 receptor (IGF1R) is a RTK that is involved in many malignancies and plays a crucial role in proliferation, transformation, apoptosis protection and chemotherapy-resistant. In this study we evaluated the expression, prognostic significance and function of IGF1R in cHL.
Immunohistochemistry was applied on 80 cHL cases. IGF1R expression was correlated with clinical data in a cHL patient cohort using Cox-regression, Chi-square and one-way ANOVA statistics. Western blot was used to determine IGF1R expression in four cHL cell lines (L428, L1236, KMH2 and UHO-1) and the IGF1 production was measured by ELISA. The Alamar blue assay was performed to determine the effect of IGF1 and cyclolignan picropodophyllin (PPP, a highly selective IGF1R inhibitor) treatment on cell growth. Flowcytometry was performed to analyze cell cycle distribution after PPP treatment.
Expression of IGF1R was observed in all or the vast majority of HRS cells in positive cases, whereas in negative cases almost all HRS cells were negative. Expression of IGF1R was detected in 55% (44/80) of the cHL patients. There was a significant difference in IGF1R expression with subtype (p=0.0157), with a high percentage of positive cases in the nodular-sclerosis cases (38/61) and a low percentage of positive cases in mixed-cellularity cases (1/9). There was no association between IGF1R expression and age, EBV status or Ann Arbor Stage. The 5-year progression-free survival (PFS) was 95% in IGF1R positive patients, whereas in IGF1R negative patients the 5-years PFS was 77% (p=0.018).
All four cHL cell lines showed expression of IGFR1, with the highest levels observed for L428 cells. The IGF1 production level was similar for all four cell lines. IGF1 treatment had the most prominent effect (31% increase) on the proliferation of L428 cells, whereas only a small effect was seen in the other three cell lines. Thus L428 cells were most sensitive to IGF1 treatment, consistent with its high IGF1R expression level. After 72 hours of PPP treatment (2μM) both the viability and proliferation decreased in all 4 cHL cell lines. The strongest effects were observed for L428 cells and UHO-1 cells with a reduction in cell growth of 48% and 43% respectively. The reduction in cell growth was 30% in KM-H2 cells and 26% in L1236. Treatment with PPP induced a G2/M cell cycle arrest in all 4 cell lines. After 24 hours incubation with PPP (2μM), the G2/M cell population increased from 8% to 45% in L428, from 10% to 64% in UHO-1, from 4% to 48% in KMH2 and from 3% to 38% in L1236.
Expression of IGF1R in the HRS cells was observed at a high frequency in the nodular sclerosis subtype and correlates with a favorable 5-years PFS. Functional studies in cHL showed a marked decrease in cell growth and an increase in the percentage of cells in the G2/M phase in all four cHL cell lines upon inhibition of IGF1R.
No relevant conflicts of interest to declare.
Asterisk with author names denotes non-ASH members.