Christoph W. M. Reuter

Targeting EGF-receptor-signalling in squamous cell carcinomas of the head and neck.

Despite significant advances in the use of surgery, chemotherapy and radiotherapy to treat squamous cell carcinoma of the head and neck (SCCHN), prognosis has improved little over the past 30 years. There is a clear need for novel, more effective therapies to prevent relapse, control metastases and improve overall survival. Improved understanding of SCCHN disease biology has led to the introduction of molecularly targeted treatment strategies in these cancers. The epidermal growth factor receptor (EGFR) is expressed at much higher levels in SCCHN tumours than in normal epithelial tissue, and EGFR expression correlates with poor prognosis. Therefore, much effort is currently directed toward targeting aberrant EGFR activity (e.g. cell signalling) in SCCHN. This review discusses the efficacy of novel therapies targeting the EGFR (e.g. anti-EGFR antibodies and EGFR tyrosine kinase inhibitors) that are currently tested in SCCHN patients.

Integrative prognostic risk score in acute myeloid leukemia with normal karyotype.

To integrate available clinical and molecular information for cytogenetically normal acute myeloid leukemia (CN-AML) patients into one risk score, 275 CN-AML patients from multicenter treatment trials AML SHG Hannover 0199 and 0295 and 131 patients from HOVON/SAKK protocols as external controls were evaluated for mutations/polymorphisms in NPM1, FLT3, CEBPA, MLL, NRAS, IDH1/2, and WT1. Transcript levels were quantified for BAALC, ERG, EVI1, ID1, MN1, PRAME, and WT1. Integrative prognostic risk score (IPRS) was modeled in 181 patients based on age, white blood cell count, mutation status of NPM1, FLT3-ITD, CEBPA, single nucleotide polymorphism rs16754, and expression levels of BAALC, ERG, MN1, and WT1 to represent low, intermediate, and high risk of death. Complete remission (P = .005), relapse-free survival (RFS, P < .001), and overall survival (OS, P < .001) were significantly different for the 3 risk groups. In 2 independent validation cohorts of 94 and 131 patients, the IPRS predicted different OS (P < .001) and RFS (P < .001). High-risk patients with related donors had longer OS (P = .016) and RFS (P = .026) compared with patients without related donors. In contrast, intermediate-risk group patients with related donors had shorter OS (P = .003) and RFS (P = .05). Donor availability had no impact on outcome of patients in the low-risk group. Thus, the IPRS may improve consolidation treatment stratification in CN-AML patients. Study registered at www.clinicaltrials.gov as #NCT00209833.

Circulating endothelial cells are an early predictor in renal cell carcinoma for tumor response to sunitinib

BackgroundTyrosine kinase inhibitors (TKI) have enriched the therapeutic options in patients with renal cell carcinoma (RCC), which frequently induce morphological changes in tumors. However, only little is known about the biological activity of TKI. Circulating endothelial cells (CEC) have been associated with endothelial damage and, hence, may serve as a putative marker for the biological activity of TKI. The main objective of our study was to evaluate the predictive value of CEC, monocytes, and soluble vascular endothelial growth factor receptor (sVEGFR)-2 in RCC patients receiving sunitinib treatment.MethodsAnalyses of CEC, monocytes, and sVEGFR-2 were accomplished for twenty-six consecutive patients with metastatic RCC who received treatment with sunitinib (50 mg, 4 wks on 2 wks off schedule) at our institution in 2005 and 2006.ResultsIn RCC patients CEC are elevated to 49 ± 44/ml (control 8 ± 8/ml; P = 0.0001). Treatment with sunitinib is associated with an increase in CEC within 28 days of treatment in patients with a Progression free survival (PFS) above the median to 111 ± 61 (P = 0.0109), whereas changes in patients with a PFS below the median remain insignificant 69 ± 61/ml (P = 0.1848). Monocytes and sVEGFR2 are frequently altered upon sunitinib treatment, but fail to correlate with clinical response, defined by PFS above or below the median.ConclusionsSunitinib treatment is associated with an early increase of CEC in responding patients, suggesting superior endothelial cell damage in these patients as a putative predictive biomarker.

A Tagging-via-substrate Approach to Detect the Farnesylated Proteome Using Two-dimensional Electrophoresis Coupled with Western Blotting

Prenylation is a post-translational modification critical for the proper function of multiple physiologically important proteins, including small G-proteins, such as Ras. Methods allowing rapid and selective detection of protein farnesylation and geranylgeranylation are fundamental for the understanding of prenylated protein function and for monitoring efficacy of drugs such as farnesyltransferase inhibitors (FTIs). Although the natural substrates for prenyltransferases are farnesyl pyrophosphate and geranylgeranyl pyrophosphate, farnesyltransferase has been shown to incorporate isoprenoid analogues into protein substrates. In this study, protein prenyltransferase targets were labeled using anilinogeraniol, the alcohol precursor to the unnatural farnesyl pyrophosphate analogue 8-anilinogeranyl diphosphate in a tagging-via-substrate approach. Antibodies specific for the anilinogeranyl moiety were used to detect the anilinogeranyl-modified proteins. Coupling this highly effective labeling/detection method with two-dimensional electrophoresis and subsequent Western blotting allowed simple, rapid analysis of the complex farnesylated proteome. For example, this method elucidated the differential effects induced by two chemically distinct FTIs, BMS-214,662 and L-778,123. Although both FTIs strongly inhibited farnesylation of many proteins such as Lamins, NAP1L1, N-Ras, and H-Ras, only the dual prenylation inhibitor L-778,123 blocked prenylation of Pex19, RhoB, K-Ras, Cdc42, and Rap1. This snapshot approach has significant advantages over traditional techniques, including radiolabeling, anti-farnesyl antibodies, or mass spectroscopy, and enables dynamic analysis of the farnesylated proteome.

Combining prenylation inhibitors causes synergistic cytotoxicity, apoptosis and disruption of RAS‐to‐MAP kinase signalling in multiple myeloma cells

The high incidence of activating RAS mutations, coupled with accumulating evidence linking RAS to multiple myeloma (MM) pathogenesis, indicate that novel therapies utilising inhibitors of RAS prenylation and signalling may be successful in the management of this disease. While preclinical studies investigating prenylation inhibitors, such as lovastatin, farnesyltransferase inhibitors (FTI) and geranylgeranyltransferase inhibitors (GGTI), have been promising, recent phase I/II clinical trials with FTI R115777 were disappointing, suggesting resistance to FTI monotherapy. To address this issue, the effects of FTI, GGTI and lovastatin alone and in combination were analysed in MM cell lines and primary cells. FTI treatment blocked H‐RAS processing, but was ineffective at inhibiting K‐ and N‐RAS prenylation because of alternative geranylgeranylation of these isoforms. However, combinations of FTI and GGTI or lovastatin were found to synergistically inhibit MM cell proliferation, migration, K‐ and N‐RAS processing, RAS‐to‐mitogen‐activated protein kinase signalling and to induce apoptosis. In contrast to FTI, lovastatin and some GGTI were found to cause intracellular accumulation of Rho proteins. Our results suggest that clinical efficacy of prenylation inhibitors in MM are limited by alternative prenylation of several small G‐proteins, such as RhoB, K‐ and N‐RAS. Furthermore, strategies combining FTI with GGTI or statins may provide greater efficacy in MM treatment.

Efficacy of sunitinib re-exposure after failure of an mTOR inhibitor in patients with metastatic RCC.

Background: The sequential use of tyrosine kinase inhibitors (TKI) followed by mTOR inhibitors (mTORi) has been recently established for the systemic treatment of metastatic renal cell carcinoma (mRCC). However, subsequent treatment in mTORi-refractory disease remains undetermined. We analyzed the efficacy of sunitinib re-challenge after failure of an mTORi at 2 German centers. Patients and Methods: Thirteen patients who failed both sunitinib and an mTORi were analyzed, and all patients were re-exposed to sunitinib. Tumor assessment was performed every 2nd cycle of sunitinib or every 3 months. Tumor response was assessed according to RECIST criteria. Results: Initial treatment with sunitinib was associated with a median progression free survival (PFS) of 21 months. Objective response consisted of 2 (15%) complete remissions and 7 (54%) partial remissions (PR) as best response. At the time of re-exposure, 12 of 13 (92%) patients again showed clinical benefit which was associated with a median PFS of 6.9 months and consisted of 2 (15%) PR and 10 (77%) disease stabilizations. Conclusions: In sunitinib-responsive patients, re-challenge with sunitinib has been successfully introduced after mTORi-refractory disease, underscoring the at least partially transient nature of TKI resistance in mRCC.

Molecularly targeted therapies in myelodysplastic syndromes and acute myeloid leukemias

Although there has been significant progress in acute myeloid leukemia (AML) treatment in younger adults during the last decade, standard induction therapy still fails to induce remission in up to 40% of AML patients. Additionally, relapses are common in 50–70% of patients who achieve a complete remission, and only 20–30% of patients enjoy long-term disease-free survival. The natural history of myelodysplastic syndrome (MDS) is variable, with about half of the patients dying from cytopenic complications, and an additional 20–30% transforming to AML. The advanced age of the majority of MDS patients limits the therapeutic strategies often to supportive care. To address these shortcomings, much effort has been directed toward the development of novel treatment strategies that target the evolution and proliferation of malignant clones. Presented here is an overview of molecularly targeted therapies currently being tested in AML and MDS patients, with a focus on FMS-like tyrosine kinase 3 inhibitors, farnesyltransferase inhibitors, antiangiogenesis agents, DNA hypomethylation agents, and histone deacetylase inhibitors.

Synergistic cytotoxic effects in myeloid leukemia cells upon cotreatment with farnesyltransferase and geranylgeranyl transferase-I inhibitors

As deregulation of RAS signaling is important in the pathogenesis of myeloid leukemias, molecular targeting of RAS signaling may be a promising therapeutic strategy. Farnesyl transferase inhibitors (FTIs) are the most promising class of these new cancer therapeutics. Several FTIs have entered phase II clinical trials in acute myeloid leukemia (AML). Since geranylgeranylation of K-RAS and N-RAS in the presence of FTIs may represent an important mechanism of FTI resistance, 6 geranylgeranyl transferase-I inhibitors (GGTIs) were screened alone and in combination with FTI for growth inhibition of myeloid leukemia cells. Significant growth inhibition (>70%) in myeloid cell lines was observed for GGTI-286 (9/19), GGTI-298 (14/19), GGTI-2147 (16/19) and FTI L-744,832 (17/17). GGTI treatment of NB-4 cells resulted in an accumulation of cells in G0/G1, whereas FTI L-744,832 primarily caused an increase in G2/M. FTI and GGTIs both induced apoptosis. In all cases, FTI/GGTI cotreatment led to synergistic cytotoxic effects in both myeloid cell lines (5/5) and primary AML cells (6/6). This synergy coincided with increased apoptosis. FTI/GGTI cotreatment caused an accumulation of unprocessed N-RAS and inactive N-RAS–RAF complexes. Our results suggest that alternative geranylgeranylation of N-RAS may represent an important mechanism of resistance to FTI monotherapy in myeloid leukemia cells.

Therapeutic efficacy of prenylation inhibitors in the treatment of myeloid leukemia

Farnesyltransferase inhibitors (FTIs) represent a new class of anticancer agents that specifically target post-translational farnesylation of various proteins that mediate several cellular processes such as signal transduction, growth, differentiation, angiogenesis and apoptosis. These compounds were originally designed to block oncogenic RAS-induced tumor growth by impeding RAS localization to the membrane, but it is now evident that FTIs also affect processing of several other proteins. The need for novel therapies in myeloid leukemia is underscored by the high rate of treatment failure due to high incidences of relapse- and treatment-related toxicities. As RAS deregulation is important in the pathogenesis of myeloid leukemias, targeting of RAS signaling may provide a new therapeutic strategy. Several FTIs (eg BMS-214662, L-778,123, R-115777 and SCH66336) have entered phase I and phase II clinical trials in myeloid leukemias. This review discusses recent clinical results, potential combination therapies, mechanisms of resistance and the clinical challenges of toxicities associated with prenylation inhibitors.

A Randomised Phase 2 Study Combining LY2181308 Sodium (Survivin Antisense Oligonucleotide) with First-line Docetaxel/Prednisone in Patients with Castration-resistant Prostate Cancer

Castration-resistant prostate cancer (CRPC) is partially characterised by overexpression of antiapoptotic proteins, such as survivin. In this phase 2 study, patients with metastatic CRPC (n=154) were randomly assigned (1:2 ratio) to receive standard first-line docetaxel/prednisone (control arm) or the combination of LY2181308 with docetaxel/prednisone (experimental arm). The primary objective was to estimate progression-free survival (PFS) for LY2181308 plus docetaxel. Secondary efficacy measures included overall survival (OS), several predefined prostate-specific antigen (PSA)-derived end points, and Brief Pain Inventory (BPI) and Functional Assessment of Cancer Therapy-Prostate (FACT-P) scores. The median PFS of treated patients for the experimental arm (n=98) was 8.64 mo (90% confidence interval [CI], 7.39-10.45) versus 9.00 mo (90% CI, 7.00-10.09) in the control arm (n=51; p=0.755). The median OS for the experimental arm was 27.04 mo (90% CI, 19.94-33.41) compared with 29.04 mo (90% CI, 20.11-39.26; p=0.838). The PSA responses (≥ 50% PSA reduction), BPI, and FACT-P scores were similar in both arms. In the experimental arm, patients had a numerically higher incidence of grades 3-4 neutropenia, anaemia, thrombocytopenia, and sensory neuropathy. In conclusion, this study failed to detect a difference in efficacy between the two treatment groups.