Summary
Microarray studies have revealed the differential expression of several genes in mantle cell lymphoma (MCL), but it is unknown which of these differences are dependent on the transformed MCL cell itself or on the tumour microenvironment. To investigate which genes and signalling pathways are aberrantly expressed in MCL cells we used oligonucleotide microarrays to perform gene expression profiling of both purified leukaemic MCL cells and their normal counterparts, the naive B cells. A total of 106 genes were differentially expressed at least threefold in MCL cells compared with naive B cells; 63 upregulated and 43 downregulated. To validate the microarray results in a larger set of samples, we selected 10 differentially expressed genes and quantified their expression by real‐time polymerase chain reaction in peripheral blood of MCL patients (n = 21), purified MCL cells (n = 6) and naive B cells (n = 4), obtaining fully concordant results. A computer‐assisted approach was used to procure specific molecular signalling pathways that were aberrantly expressed in MCL cells. Several genes related to apoptosis and to the PI3K/AKT, WNT and tumour growth factor β signalling pathways were altered in MCL cells when compared with naive B cells. These pathways may play a significant role in the pathogenesis of MCL and deserve further investigation as candidates for new therapeutic targets.
Purpose
Proteasome inhibition disrupts protein homeostasis and induces apoptosis. Up to 50% of patients with relapsed mantle cell lymphoma (MCL) respond to bortezomib. We used gene expression profiling to investigate the connection between proteasome inhibition, cellular response, and clinical efficacy.
Experimental Design
We assessed transcriptional changes in primary tumor cells from five patients during treatment with bortezomib in vivo, and in 10 MCL cell lines exposed to bortezomib in vitro, on Affymetrix microarrays. Key findings were confirmed by Western blotting.
Results
MCL cell lines exposed to bortezomib in vitro showed upregulation of endoplasmic reticulum and oxidative stress response pathways. Gene expression changes were strongest in bortezomib sensitive cells and these cells were also more sensitive to oxidative stress induced by H2O2. Purified tumor cells obtained at several timepoints during bortezomib treatment in five previously untreated patients with leukemic MCL showed strong activation of the antioxidant response controlled by NRF2. Unexpectedly, activation of this homeostatic program was significantly stronger in tumors with the best clinical response. Consistent with its pro-apoptotic function we found upregulation of NOXA in circulating tumor cells of responding patients. In resistant cells gene expression changes in response to bortezomib were limited and upregulation of NOXA was absent. Interestingly, at baseline, bortezomib resistant cells displayed a relatively higher expression of the NRF2 gene expression signature than sensitive cells (P<0.001).
Conclusion
Bortezomib triggers an oxidative stress response in vitro and in vivo. High cellular antioxidant capacity contributes to bortezomib resistance.
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