Targeting anti-apoptotic Bcl-2 by AT-101 to increase radiation efficacy: data from in vitro and clinical pharmacokinetic studies in head and neck cancer

Abstract

BackgroundPro-survival Bcl-2 family members can promote cancer development and contribute to treatment resistance. Head and neck squamous cell carcinoma (HNSCC) is frequently characterized by overexpression of anti-apoptotic Bcl-2 family members. Increased levels of these anti-apoptotic proteins have been associated with radio- and chemoresistance and poor clinical outcome. Inhibition of anti-apoptotic Bcl-2 family members therefore represents an appealing strategy to overcome resistance to anti-cancer therapies. The aim of this study was to evaluate combined effects of radiation and the pan-Bcl-2 inhibitor AT-101 in HNSCC in vitro. In addition, we determined human plasma levels of AT-101 obtained from a phase I/II trial, and compared these with the effective in vitro concentrations to substantiate therapeutic opportunities.MethodsWe examined the effect of AT-101, radiation and the combination on apoptosis induction and clonogenic survival in two HNSCC cell lines that express the target proteins. Apoptosis was assessed by bis-benzimide staining to detect morphological nuclear changes and/or by propidium iodide staining and flow-cytometry analysis to quantify sub-diploid apoptotic nuclei. The type of interaction between AT-101 and radiation was evaluated by calculating the Combination Index (CI) and by performing isobolographic analysis. For the pharmacokinetic analysis, plasma AT-101 levels were measured by HPLC in blood samples collected from patients enrolled in our clinical phase I/II study. These patients with locally advanced HNSCC were treated with standard cisplatin-based chemoradiotherapy and received dose-escalating oral AT-101 in a 2-weeks daily schedule every 3 weeks.ResultsIn vitro results showed that AT-101 enhances radiation-induced apoptosis with CI’s below 1.0, indicating synergy. This effect was sequence-dependent. Clonogenic survival assays demonstrated a radiosensitizing effect with a DEF37 of 1.3 at sub-apoptotic concentrations of AT-101. Pharmacokinetic analysis of patient blood samples taken between 30 min and 24 h after intake of AT-101 showed a dose-dependent increase in plasma concentration with peak levels up to 300–700 ng/ml between 1.5 and 2.5 h after intake.ConclusionAT-101 is a competent enhancer of radiation-induced apoptosis in HNSCC in vitro. In addition, in vitro radiosensitization was observed at clinically attainable plasma levels. These finding support further evaluation of the combination of AT-101 with radiation in Bcl-2-overexpressing tumors.

Gossypol stimulates opening of a Ca2+‐ and Na+‐permeable but Ni2+‐ and Co2+‐impermeable pore in bEND.3 endothelial cells

“…Acute cytotoxicity inflicted by 10 µmol/L gossypol was attenuated by extensive washing (Figure C) and in part depended on extracellular Ca 2+ influx (Figure D). In a clinical trial, head and neck cancer patients had peak plasma gossypol concentration of around 1.35 µmol/L after oral intake of gossypol for about 2 hours . This concentration is close to the cytotoxic concentration we report here (3 µmol/L) and in a previous work showing gossypol killed HUVEC with LD 50 of 1.7 μmol/L .…”

Apoptosis signaling molecules as treatment targets in head and neck squamous cell carcinoma

“…21 The in vitro efficacy of the first-generation Bcl-2/ Bcl-xL inhibitor, ABT-737, has been examined in HNSCC cells in combination with cisplatin and etoposide 22 and also with sorafenib. 23 Other apoptosis-acting therapeutics, including AT-101, 24 obatoclax, 25 and venetoclax, 26 have been studied as single agents. Several studies prior to the advent of newer Mcl-1-directed molecular agents have examined down-regulation of Mcl-1 as an effective mechanism of action of several drugs in HNSCC.…”

Effects of microRNA-26b on proliferation and invasion of glioma cells and related mechanisms

“…The depressed apoptosis level and over-proliferation of tumors cells all elevate the probability of tumor progression and invasion or metastasis (25). The over-expression of the anti-apoptotic gene Bcl-2 leads to abnormal proliferation of tumor cells and diminished apoptosis of necrotic cells, all of which are important factors causing tumor formation and progression, making the progression of tumor uncontrollable (26). The initiation of the cell apoptotic program can inhibit tumor growth and facilitate activation of the apoptosis family of proteins, in which caspase-3 is one of the most important members.…”