Bortezomib-Induced Apoptosis with Limited Clinical Response Is Accompanied by Inhibition of Canonical but not Alternative Nuclear Factor-κB Subunits in Head and Neck Cancer

Abstract

Purpose: Nuclear factor-nB (NF-nB)/REL transcription factors promote cancer cell survival and progression. The canonical (NF-nB1/RELA or cREL) and alternate (NF-nB2/RELB) pathways require the proteasome for cytoplasmic-nuclear translocation, prompting the investigation of bortezomib for cancer therapy. However, limited clinical activity of bortezomib has been observed in many epithelial malignancies, suggesting this could result from incomplete inhibition of NF-nB/ RELs or other prosurvival signal pathways. Experimental Design:To examine these possibilities, matched biopsies from 24 h posttreatment were obtained from accessible tumors of patients who received low-dose bortezomib (0.6 mg/m 2 ) before reirradiation in a phase I trial for recurrent head and neck squamous cell carcinoma (HNSCC). Effects of bortezomib on apoptosis and proliferation by TUNEL and Ki67 staining were compared with nuclear staining for all five NF-nB subunits, phosphorylated extracellular signal-regulated kinase 1/2 (ERK1/2), and phosphorylated signal transducers and activators of transcription 3 (STAT3) in tumor biopsies, and by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTP) and DNA binding assay for the five NF-nB subunits in HNSCC cell lines. Results: HNSCC showed increased nuclear staining for all five NF-nB subunits, phosphorylated ERK1/2, and phosphorylated STAT3. Bortezomib treatment significantly enhanced apoptosis with inhibition of nuclear RELA in three of four tumors, but other NF-nB subunits, ERK1/2, and STAT3 were variably or not affected, and tumor progression was observed within 3 months. In HNSCC cell lines, 10 -8 mol/L bortezomib inhibited cell density while inhibiting tumor necrosis factor-aî nduced and partially inhibiting basal activation of NF-nB1/RELA, but not NF-nB2/RELB. Conclusions: Although low-dose bortezomib inhibits activation of subunits of the canonical pathway, it does not block nuclear activation of the noncanonical NF-nB or other prosurvival signal pathways, which may contribute to the heterogeneous responses observed in HNSCC.Nuclear factor-nB (NF-nB) is an injury, pathogen, growth factor, cytokine, and matrix signal-inducible transcription factor that normally functions to regulate cell survival and proliferation and innate and adaptive immune responses (1). NF-nB transcription factors are heterodimers or homodimers composed from two families of REL proteins. Each member of the REL family contains an NH 2 terminal REL homology domain that mediates dimerization and domains mediating nuclear localization and binding to specific nB DNA sequence motifs in target genes (1). The first class, RELA (p65), RELB, and cREL, is synthesized in their mature forms, whereas the second class, consisting of NF-nB1(p105) and NF-nB2(p100), requires proteasome-dependent proteolysis of COOH terminal ankyrin repeats to yield p50 and p52, respectively (2). In the absence of activating stimuli, NF-nB dimers are also bound by a family of ankyrin repeat-containing inhibitor-nB (InB) proteins, sequeste...

EGFR and NF-κB: partners in cancer

“…Bortezomib, a proteasome inhibitor, triggers cell apoptosis by blocking NF-kB activation through IkBa stabilization [63]. However, bortezomib showed limited clinical efficacy in HNSCC because NF-kB activation through the noncanonical pathway remained unchanged [64,65]. Moreover, bortezomib did not impact on other prosurvival pathways driven by STAT3 and ERK1/2 phosphorylation [64].…”

“…However, bortezomib showed limited clinical efficacy in HNSCC because NF-kB activation through the noncanonical pathway remained unchanged [64,65]. Moreover, bortezomib did not impact on other prosurvival pathways driven by STAT3 and ERK1/2 phosphorylation [64]. Therefore, the combination of bortezomib with EGFR inhibitors was expected to improve the clinical response.…”

The Early Marginal Zone B Cell-Initiated T-Independent Type 2 Response Resists the Proteasome Inhibitor Bortezomib

“…Hence, it remains elusive as to which stimuli or cellular properties in particular activate potential resistance mechanisms such as induction of immunoproteasome subunits and antiapoptotic UPR pathways in MZ B cells. As noncanonical NF-kB signaling is crucial for MZ B cell development (30), it is possible that activation of this pathway might contribute to the resistance toward bortezomib (31). In this respect it is noteworthy that we have preliminary evidence that noncanonical NF-kB signaling is not inhibited by therapeutic concentrations of bortezomib (D. Mielenz, V.R.…”

Fighting cancer from different signalling pathways: Effects of the proteasome inhibitor Bortezomib in combination with the polo-like-kinase-1-inhibitor BI2536 in SCCHN

“…Several in vitro and in vivo studies showed the proteasome inhibitor Bortezomib to effectively inhibit tumour growth, induce apoptosis and lower vessel density in SCCHN (7,(16)(17)(18)(19). Furthermore, Bortezomib led to a sustained but limited response rate in SCCHN patients suffering from tumour recurrence, who underwent radiation therapy combined with Bortezomib (20,21). With regard to these findings, it may be possible that optimisation of Bortezomib efficacy may yield substantial improvement in the therapy of head and neck cancer although the use of Bortezomib in SCCHN is discussed controversially in the literature (22,23).…”