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8007 Background: Objectives of this study included assessing safety and activity of AMG 420/BI 836909, which binds BCMA (B-Cell Maturation Antigen) on MM cells and CD3 on T cells, in relapsed and/or refractory (R/R) MM. Methods: In this FIH study, 6-week cycles of AMG 420 were given for ≤5 cycles or until disease progression (PD), toxicity, or consent withdrawal; 5 more cycles could be given for benefit. Eligible patients had progression after ≥2 lines (incl PI and IMiD). Excluded were PC leukemia, extramedullary relapse, CNS involvement, or prior allo-SCT. MRD was defined as 4 bone marrow cells per flow cytometry. Results: As of Dec 10, 2018, 42 patients received AMG 420 (0.2-800 µg/d). Patients D/C for PD (n=24), adverse events (AE, n=7, incl 3 DLTs), death (4), completed 10 cycles (2), and consent (1). Median age was 65 y, median MM duration 5.2 y, and median # prior therapies 4. Patients were treated for a mean (SD) of 2.5 (2.6) cycles. There were 2 deaths from AEs (acute respiratory distress from flu / aspergillosis; fulminant hepatitis related to adenovirus infection); neither treatment related. Of those with serious AEs (SAEs, n=21, 50%), 18 required hospitalization. SAEs occurring in >1 patient were infections (n=12) and polyneuropathy (PN, n=2). Treatment-related SAEs included 2 grade 3 PNs and 1 edema. Grade 2-3 CRS was seen in 3 patients. No anti-AMG 420 Ab were detected. In this study, 800 µg/d was determined to not be tolerable as 2/3 patients had DLTs, 1 case of grade 3 CRS and 1 case of grade 3 PN; both required hospitalization and subsequently resolved. At 400 µg/d, there were 5 minimal residual disease (MRD)-negative sCRs, 1 VGPR, and 1 PR, for a response rate of 7/10 (70%); at Dec datacut, responses lasted for 5.6-10.4 months with 4 patients ongoing on treatment. As of Feb 2019, some responses lasted >1 year. Overall, there were 13/42 responders (6 sCRs, 3 CRs, 2 VGPRs, 2 PRs). Median time to any response was 1 month. Conclusions: In this FIH study of AMG 420, a BiTE vs BCMA, in R/R MM, there was a 70% response rate (7/10) with 5 out of 7 responders achieving a sCR at 400 µg/d, a recommended dose for further investigation. Clinical trial information: NCT02514239. more...

Background: Polo like kinase 1 (PLK1) is frequently upregulated in tumors and is thus viewed as a promising therapeutic target in various cancers. Several PLK1 inhibitors have recently been developed and clinically tested in solid cancers, albeit with limited success. So far, no predictive biomarkers for PLK1 inhibitors have been established. To this end, we conducted a post-hoc biomarker analysis of tumor samples from non-small cell lung cancer (NSCLC) patients treated with the PLK1 inhibitor BI2536 in a phase II study. Methods: We analyzed formalin-fixed paraffin-embedded surplus tumor tissue from 47 study patients using immunohistochemistry (IHC) and DNA sequencing of KRAS, EGFR, BRAF, and PIK3CA. Results:KRAS-mutated patients showed numerically prolonged progression-free survival, but statistical significance was not established. Interestingly, when pathways rather than single genes were analyzed, a positive correlation between IHC staining of activated ERK (p-ERK) and mutated KRAS was detected, whereas KRAS mutation status was found to be negatively correlated with activated AKT (p-AKT). Conclusion: With this hypothesis-generating study in BI2531-treated patients, we could not establish a correlation between KRAS mutations and relevant clinical endpoints. Future clinical trials with concomitant systematic biosampling and comprehensive molecular analyses are required to identify biomarkers predictive for response to PLK1 inhibitors. more...

Objectives This phase II, open-label study was designed to evaluate the response rate to the polo-like kinase 1 (Plk-1) inhibitor BI 2536 in patients with sensitive-relapsed small cell lung cancer (SCLC). Secondary endpoints included progression-free survival (PFS), overall survival (OS), duration of response, and safety. Materials and methods Patients were treated with the recommended phase II dose of 200 mg of BI 2536 intravenously every 21 days. This was a two-stage design with an early stopping rule in place if responses were not seen in at least 2 of the first 18 enrolled patients. Results and conclusion Twenty-three patients were enrolled in the study and 21 patients were evaluable for response. No responses were observed and all 23 patients have progressed. The median PFS was 1.4 months. Treatment was generally well tolerated and the most frequent adverse events were neutropenia, fatigue, nausea, vomiting, and constipation. BI 2536 is not effective in the treatment of sensitive relapsed SCLC. The criteria for expanding the trial to the second stage were not achieved, and the study was terminated for a lack of efficacy. more...

Polo-like kinase 1 (Plk1) is expressed during mitosis and overexpressed in multiple cancers, including non-Hodgkin lymphoma (NHL). This phase I study determined the maximum tolerated dose (MTD) of BI 2536, a Plk1 inhibitor, as a 1 h infusion once every 3 weeks in post-transplant relapsed (n = 17) and transplant-naive (n = 24) patients with relapsed/refractory NHL. Median treatment cycles were 2 and 1.5, respectively. MTD was 175 mg for both populations; dose-limiting toxicities were grade 4 thrombocytopenia and neutropenia. Most treatment-related adverse events were grade 1/2; drug-related grade 3/4 events included thrombocytopenia and neutropenia. Four patients achieved responses (three complete and one partial at doses ≥ 150 mg, all post-transplant relapsed patients) for an overall response rate of 9.8%. BI 2536 exhibited multi-compartmental pharmacokinetics with a high volume of distribution. The activity and safety of BI 2536 in this pretreated patient population support Plk inhibitors as a therapeutic strategy in oncology. more...

Background: BI 2536, a novel Polo-like kinase 1 inhibitor, was assessed in patients with unresectable advanced exocrine adenocarcinoma of the pancreas. Methods: The study employed a two-stage design. Randomised first-line patients received BI 2536 200 mg on day 1 (n=43) or 60 mg on days 1-3 (n=43) every 21 days. Recruitment of second-line patients was planned for a second stage dependent on an interim analysis demonstrating ≥2 responses in the first 18 evaluable patients following 12 weeks of treatment and/or tumour control ≥12 weeks in 5 patients per schedule. Primary end point was objective response rate (ORR). Results: By independent review, ORR was 2.3% (all partial) and 24.4% had stable disease as confirmed best response. The second stage was not initiated. Median overall and progression-free survivals were 149 (95% confidence interval (CI), 91-307) and 46 days (95% CI, 44-56). Most common drug-related adverse events were neutropenia (37.2%), leukopenia (29.1%), fatigue (29.1%) and nausea (22.1%); most common grade 3/4-related events were neutropenia (36.0%), leukopenia (27.9%) and thrombocytopenia (8.1%). Conclusion: Given the low ORR and poor survival, further development of BI 2536 monotherapy is not warranted in this population. © 2012 Cancer Research UK All rights reserved. more...

Background: Of all mastectomy patients, 90% will use an external prosthesis where the standard of care uses a stock prosthesis that is purchased “off the shelf.” Our objectives were to determine patient demand for and perceived value of a custom breast prosthesis. The information obtained will influence future research and program direction. Methods: We asked 65 women who had undergone lumpectomy or mastectomy to participate before exploring rehabilitation options. The quantitative outcome measures were the European Organisation for Research and Treatment of Cancer QLQ-C30 general and -BR23 breast cancer–specific quality of life questionnaires, and the Ambulatory Oncology Patients Satisfaction Tool. The QLQ results were analyzed using the Mann–Whitney U-test. Results of the satisfaction tool were compared using the Fisher exact and chi-square tests. A descriptive qualitative approach—involving in-depth interviews exploring the experiences of the women—was used to establish the perceived value of the services to the patients. The analysis of the interview transcripts was conducted using a standardized content method to describe the experiences of the women. Results: All the women had had previous experiences with a conventional prosthesis, and they reported that wearing a custom prosthesis was more satisfying for them. They reported comfort and ease in wearing it, coupled with a sense of feeling less like a victim. Comparison of the qlq and patient satisfaction scores showed no significant difference between the women wearing the conventional prosthesis and those wearing the custom prosthesis. Conclusions: The willingness of women to pay for a prosthesis and the qualitative results from the present study demonstrate that there is demand for a custom approach to treatment. However, if a mixed-methods approach had not been applied during this initial exploration of women’s experiences with custom breast prostheses, the essence of the perceived value of the custom prosthesis would have been lost. Quantitative measures suggest that there is no difference between custom and conventional breast prostheses, but the qualitative data captured in the study provide a sense of aspects of care that a standardized outcome measure cannot capture. Further research with a larger sample size is needed to determine if real differences from a quantitative perspective are possible. Suggestions for improvements in the device and in program operations were gathered and will influence the future development and implementation of a breast prosthesis service, but financial assistance will most likely be needed to make such a service universally accessible. more...

Background Volasertib (BI 6727) is a potent and selective cell-cycle kinase inhibitor that induces mitotic arrest and apoptosis by targeting Polo-like kinase (Plk). This phase I dose-escalation study evaluated the maximum tolerated dose (MTD) of volasertib, safety and efficacy, and pharmacokinetic (PK) parameters. Methods This trial followed an open-label, toxicity-guided dose-titration design. Patients with progressive advanced or metastatic solid tumours received a single 1-h infusion of volasertib every 3 weeks. A total of 65 patients were treated at doses of 12–450 mg. Results Reversible haematological toxicity was the main side-effect; thrombocytopenia, neutropenia, and febrile neutropenia constituting the main dose-limiting events. Anaemia (all grades 22%; grade 3: 8%), neutropenia (15%; grade 3/4: 14%), fatigue (15%; grade 3: 2%), and thrombocytopenia (14%; grade 3/4: 14%) were the most frequent drug-related adverse events. The MTD was 400 mg; however, 300 mg was the recommended dose for further development based on overall tolerability. Three patients achieved confirmed partial response. Stable disease as best response was reported in 40% of patients. Two patients remained progression free for >1 year. PK analysis showed no indication of deviation from ‘dose-linear PK’ behaviour, a large volume of distribution (>4000 l), moderate clearance and a long half-life (∼111 h). Conclusion This first-in-man trial demonstrated a favourable PK profile of volasertib, with manageable toxicities. As expected, the most common events were haematological. Encouraging preliminary antitumour activity has been observed, supporting Plk inhibition as a therapeutic approach. Clinical development of volasertib in phase II monotherapy and combination trials is ongoing. more...

The aim of this investigation was to compare the performance of a commonly used semi-mechanistic model for drug-related neutropenia with other semi-mechanistic models published in the literature. After their implementation in NONMEM VI, five semi-mechanistic models were assessed using the pharmacokinetic and absolute neutrophil count data obtained from 95 patients with non-small cell lung cancer receiving either 200 mg on day 1 or 50 or 60 mg on days 1, 2 and 3 of a 21-day treatment course with the new Plk-1 inhibitor BI 2536. The model performance was compared by means of predictive (visual and numerical) checks, precision in the parameter estimates and objective function-based measures. Details of model parameterization, model stability and run times are also provided. The time course of the drug plasma concentrations was described by a three compartment model with a first-order elimination rate. With respect to neutropenia, all models were successfully implemented in NONMEM and provided reasonable fits for the median (although not all models described all percentiles of the data well), and in general precise parameter estimates. In the current evaluation performed in a single drug, none of the models showed superior performance compared to the most commonly used model first described by Friberg et al. (J Clin Oncol 20:4713–4721, 2002). more...

Background: BCMA, a member of the TNF receptor family, is expressed on MM and plasma cells. AMG 420, formerly BI 836909, binds BCMA on tumor cells and plasma cells and CD3 on T cells, resulting in T-cell mediated lysis of BCMA+ cells. Objectives of this study of AMG 420 in patients with R/R MM included assessing safety and tolerability and anti-tumor activity per IMWG 2006. Methods: This is a FIH phase I dose escalation study (NCT02514239) of 6-week cycles of AMG 420 (1 cycle=4 weeks continuous IV infusion, 2 weeks off). Single-patient cohorts [0.2-1.6 µg/day (d)] were followed by cohorts of 3-6 patients (3.2-800 µg/d). Eligible patients had R/R MM and progression after ≥2 prior treatment lines (including proteasome inhibitor and immunomodulators); excluded were patients with plasma cell leukemia, extramedullary relapse, known central nervous system involvement, or prior allogeneic stem cell transplant. Treatment continued for up to 5 cycles or until disease progression (PD), start of new therapy, toxicity, withdrawal of consent, or investigator decision; 5 more cycles could be given per investigator for perceived benefit. MRD response was defined for this study as Results: As of May 22, 2018, 35 patients received AMG 420 (0.2-800 µg/d). Patients discontinued for PD (n=21), adverse events [AE, n=7, including 2 dose-limiting toxicities (DLTs)], or completed 10 cycles (n=2); 5 remain on study. Mean (SD) age was 63.8 (8.7) years, median 65 years, min-max 39-79 years, and 22 (63%) were male. Median MM disease duration was 5.4 (Q1: 3.3, Q3: 7.4) years, min-max 1.3-20 years, and median # of prior therapies was 4 (Q1: 2, Q3: 5), min-max 2-13. Patients (n=35) were treated for a mean (SD) of 2.3 (2.3) cycles and a median (min-max) of 1 (1-10) cycles; responders (n=8) were treated for a mean (SD) of 5.3 (3.3) cycles and a median (min-max) of 3.5 (2-10) cycles, including those with treatment ongoing. Regarding safety, one patient in the 50 µg/d cohort died after the first cycle from acute respiratory distress due to concurrent flu and aspergillosis not considered related to treatment. Of those with serious AEs (n=17, 49%), 12 required hospitalization and another 3 had prolonged hospitalization. Serious AEs included infections (n=10, 29%, 3 device-related, 3 pneumonias, 2 catheter site, 1 aspergillus, 1 influenza, and 1 fever/infection), cytokine release syndrome (CRS, n=3), and 1 each of peripheral polyneuropathy (PPN), cardiac failure, edema, pyrexia, biliary obstruction, and renal failure. Treatment-related serious AEs included CRS (n=3, 2 grade 1 and 1 grade 3) and 1 each of PPN (grade 3), edema (grade 3), and pyrexia (grade 1). No anti-AMG 420 antibodies were detected up to 800 μg/d and no DLTs were observed up to 400 µg/d. In this study, 800 µg/d was determined to not be tolerable as 2/3 patients experienced DLTs: 1) Grade 3 CRS within 1d of initiating treatment with fever, hypertension, tachycardia, and retrograde amnesia; symptoms resolved after stopping drug, and 2) Grade 3 PPN that required hospitalization with subsequent complete recovery; after 15d of treatment, M protein decreased by 60%. Six patients had complete responses (CRs), 1 each at 6.5, 100, and 200 µg/d, and 3 at 400 µg/d; responses are ongoing for these last 3 (≥4.6 months). There also were two partial remissions, a partial response (PR) at 50 µg/d and a very good partial response at 800 µg/d. Response duration was for up to 8 cycles (1 patient had PR cycles 3-10). All patients at 400 µg/d (3/3) had MRD negative CRs. In the dose confirmation cohort enrolled after May 22, 2 of 3 patients had PRs as of cycle 1. Thus, at the dose of 400 µg/d, the objective response rate is 5/6 (83%); all 5 are still responding on treatment. Pharmacokinetic analyses show that responders had higher free-drug exposure levels than non-responders [median (range) of 3,225 (36-108,000) vs 97 (27-1,380) pg/mL]. Conclusions: In this FIH study, AMG 420, a short half-life BiTE® targeting BCMA, showed encouraging evidence of activity in patients with R/R multiple myeloma. During dose escalation, all 3 patients dosed with 400 µg/d had MRD-negative CRs, with 2 more responders in the dose confirmation cohort to date; 3 patients at lower doses also attained CRs. No major toxicities were observed up to 400 µg/d, which is a recommended dose for further investigation; DLTs at 800 µg/d were CRS and PPN. Figure Figure. Disclosures Topp: Regeneron Pharmaceuticals, Inc.: Honoraria, Research Funding; Amgen: Honoraria, Membership on an entity's Board of Directors or advisory committees, Other: Travel, Research Funding; F. Hoffmann-La Roche Ltd: Membership on an entity's Board of Directors or advisory committees, Research Funding; Boehringer Ingelheim: Research Funding. Zugmaier:Amgen Inc.: Consultancy, Employment, Patents & Royalties: 20170327581, 9688760, 20170122947, 9486475, 20160208001, 9192665, 20150071928, 8840888, 20140227272, 20140228316, 20130323247, 20130287774, 20130287778, 20110262440, 20100112603, 7700299, 20070037228. Moreau:Abbvie: Honoraria, Membership on an entity's Board of Directors or advisory committees; Takeda: Honoraria, Membership on an entity's Board of Directors or advisory committees; Celgene: Honoraria, Membership on an entity's Board of Directors or advisory committees; Amgen: Honoraria, Membership on an entity's Board of Directors or advisory committees; Janssen: Honoraria, Membership on an entity's Board of Directors or advisory committees. Facon:Takeda: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Janssen: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Amgen: Membership on an entity's Board of Directors or advisory committees; Karyopharm: Membership on an entity's Board of Directors or advisory committees; Sanofi: Membership on an entity's Board of Directors or advisory committees; Celgene: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Oncopeptides: Membership on an entity's Board of Directors or advisory committees. Munzert:Boehringer Ingelheim: Employment, Patents & Royalties: and other intellectual property . more...

This study investigated the feasibility of predicting the neutropenia-related effects of a therapy that combines the investigational drug BI 2536 (inhibitor of Polo-like kinase 1) and pemetrexed, an approved anticancer drug. Predictions were arrived at using the pharmacokinetic/pharmacodynamic (PK/PD) parameters of each of the drugs obtained from monotherapy studies and assuming that the neutropenic effect is additive when the drugs are administered as a combination therapy. Subsequently, a PK/PD model was developed to determine whether this assumption of additive effect was reasonable in relation to these two drugs. All analyses and simulations were performed using the population approach in NONMEM, version VI. Clinical Pharmacology & Therapeutics (2010) 88 5, 660–667. doi: 10.1038/clpt.2010.148 more...

Objective To investigate the efficacy, safety, and pharmacokinetics of two dosing schedules of BI 2536, a novel polo-like kinase-1 inhibitor, in patients with relapsed stage IIIB/IV non-small cell lung cancer. Methods Ninety-five patients were randomized to intravenous BI 2536 on day 1 (200 mg) or days 1 to 3 (50 or 60 mg) of a 21-day treatment course. BI 2536 doses were escalated beyond course 2 if well tolerated. The primary objective was response, and the secondary objectives were progression-free survival (PFS) and overall survival (OS), quality of life, safety, and pharmacokinetics. Primary statistical aim was to demonstrate the difference in objective response rate to historical placebo for both treatment groups. Results Four patients (4.2%) had a partial response; two were confirmed by independent review. Median PFS was 8.3 weeks (58 days 95% confidence interval [CI]: 48–85) and 7 weeks (49 days 95% CI: 46–70) assessed by investigator and independent review, respectively. Median OS was 28.7 weeks (201 days 95% CI: 180–305). No statistically significant difference was observed between the two treatment schedules regarding clinical benefit, PFS, or OS. Grade 4 neutropenia occurred in 37% of patients; common nonhematologic adverse events were fatigue (31%) and nausea (27%). Two deaths (pulmonary hemorrhage and sepsis) were considered drug related. There was a trend in favor of the days 1 to 3 dosing schedule in quality of life. BI 2536 displayed moderate interpatient variability. Conclusions BI 2536 monotherapy has modest efficacy and favorable safety in relapsed non-small cell lung cancer. The findings support the further development of polo-like kinase-1 inhibitors within this indication. more...

(1) To describe the neutropenic response of BI 2536 a polo-like kinase 1 inhibitor in patients with cancer using a semi-mechanistic model. (2) To explore by simulations (a) the neutropenic effects for the maximum tolerated dose (MTD) and the dose at which dose-limiting toxicity occurred, (b) the possibility to reduce the cycle duration without increasing neutropenia substantially, and (c) the impact of the initial absolute neutrophil count (ANC) on the degree of neutropenia for different doses.BI 2536 was administered as intravenous infusion over 60 min in the dose range from 25 to 250 mg. Three different administration schedules were explored: (a) day 1, (b) days 1, 2, and 3 or (c) days 1 and 8 within a 3 week treatment cycle. BI 2536 plasma concentrations and ANC obtained during the first treatment cycle from 104 patients were analysed using the population approach with NONMEM VI.Neutropenia was described by a semi-mechanistic model resembling proliferation at the stem cell compartment, maturation, degradation, and homeostatic regulation. BI 2536 acts decreasing proliferation rate. Simulations showed that (1) all MTD doses showed an acceptable risk of neutropenia, (2) when BI 2536 is given as 200 mg single administration, cycle duration can be reduced from 3 to 2 weeks, and (3) baseline ANC might be considered to individualise the dose of BI 2536.A semi-mechanistic population model was applied to describe the neutropenic effects of BI 2536. The model was used for simulations to support further clinical development. more...

Aims: BI 2536 is a selective and potent small-molecule inhibitor of polo-like kinase 1. We performed a multi-centre, multi-tumour phase II trial to investigate the efficacy, safety and pharmacokinetics of BI 2536 in five solid tumour types. Patients and methods: Patients with advanced head and neck, breast and ovarian cancer, soft tissue sarcoma and melanoma were selected according to protocol-defined general and tumour-specific criteria. They were >= 18 years old, had a good performance status, adequate bone marrow, renal and liver function, measurable progressive disease and had completed other relevant systemic treatments >4 weeks ago. BI 2536 200-250 mg was given intravenously on day 1 every 3 weeks until intolerance, progression or refusal. The study was based on a Simon two-stage design, with 12 patients entering in stage 1 and additional 25 patients to be entered in case of at least one response in the first stage. The rate of objective responses (RECIST criteria) was chosen as primary end-point. Results: Seventy six patients were included, 71 started treatment and received a median number of two cycles (four in ovarian cancer). Frequent grade 3-4 adverse events were neutropaenia (81.6%), thrombocytopaenia (19.7%), febrile neutropaenia (19.7%), anaemia (15.5%) and pain (9.9%). We did not observe confirmed objective responses. All cohorts were closed after the entry of 14-15 eligible non-responding patients. Pharmacokinetic analyses revealed multi-compartmental behaviour and a rapid distribution of BI 2536. Conclusions: BI 2536 showed limited antitumour activity according to the design of this trial in five different tumour types. Derivatives of BI 2536 with a more favourable pharmacological profile are currently explored further in prospective studies. (C) 2010 Elsevier Ltd. All rights reserved. more...

Purpose BI 2536 is a novel, potent, and highly specific inhibitor of polo-like kinase 1 (Plk1), which has an essential role in the regulation of mitotic progression. The aim of this trial was to identify the maximum tolerated dose (MTD) of BI 2536 and to determine the safety, pharmacokinetics, and antitumor activity in patients who had advanced solid tumors. Patients and Methods This phase I trial followed an open label, toxicity-guided, dose-titration design. Single doses of BI 2536 (25 to 250 mg) were administered as a 1-hour intravenous infusion; patients who experienced clinical benefit were eligible for additional treatment courses. Safety and pharmacokinetics were investigated. Tumor response was evaluated according to Response Evaluation Criteria in Solid Tumors Group guidelines. Results The MTD was defined at 200 mg in a total of 40 patients entered; reversible neutropenia constituted the dose-limiting toxicity (DLT) and the most frequent adverse event at the MTD (grade 3 to 4; 56%). Nausea (52%), fatigue (52%), and anorexia (44%) also were common and were mostly of mild to moderate intensity (Common Terminology Criteria of Adverse Events ≤ grade 2). One patient experienced a transient partial response. At doses equal to or greater than the MTD, 23% of patients experienced disease stabilization for 3 or more months. Dose-proportional increases in the maximum plasma concentration and total exposure were observed. BI 2536 showed a high total clearance and high distribution into tissue. Conclusion The MTD of BI 2536 when administered as a single-dose, 1-hour infusion was 200 mg; BI 2536 was well tolerated and showed a favorable pharmacokinetic profile. Antitumor activity of BI 2536 was observed. more...

In acute myeloid leukemia (AML), receptor tyrosine kinase ligands promote growth and survival and contribute to AML-associated marrow neoangiogenesis. We have tested simultaneous inhibition of vascular endothelial growth factor, fibroblast growth factor, and platelet-derived growth factor receptor signaling by novel indolinone derivatives using 14 myeloid, including 11 human leukemic, cell lines. Compounds inhibited colony formation of all cell lines in a dose-dependent fashion. Inhibitory concentrations for 50% of the colony formation/survival (IC50) for BIBF1000 were more...

Bcr-Abl constitutes a deregulated tyrosine kinase involved in the pathogenesis of chronic myeloid leukemia (CML) and a subset of acute lymphoblastic leukemia (ALL). Although activation of the transcription factor NF-kappaB/Rel has been demonstrated, mechanisms of NF-kappaB/Rel activation by Bcr-Abl remain obscure. In this paper we demonstrate activation of NF-kappaB/Rel by Bcr-Abl and for the first time by v-Abl. Furthermore, we investigated mechanisms of NF-kappaB/Rel induction by Bcr-Abl and v-Abl. Both Bcr-Abl and v-Abl induced NF-kappaB/Rel DNA binding in Ba/F3 cells. DNA binding was a result of nuclear translocation of p65/RelA, whereas p65/RelA expression was unaffected. Nuclear translocation of p65/RelA is at least partially due to increased IkappaBalpha degradation, which is independent of IkappaB kinase (IKK) activity. IKK activity is not deregulated by Bcr-Abl and v-Abl. NF-kappaB/Rel transactivation was dependent on abl kinase activity but independent of Grb2 and Grb10 binding tobcr sequences. In addition, NF-kappaB/Rel activation was dependent on Ras activity. Primary CML blasts showed constitutive p65/RelA NF-kappaB/Rel DNA binding activity. Thus NF-kappaB/Rel represents a potential target for molecular therapies in CML. more...

The aim of this study was to enhance the delivery and thus anti-tumoral efficiency of antisense bcl-2 oligonucleotides (ODN's).Bcl-2 overexpressing DoHH2 lymphoma and HeLa-cells were transfected with ODN's using a polycationic liposome preparation. Specific hybridization of antisense ODN's was demonstrated by gel-shift assays and in vitro transcription/translation studies. Cellular uptake of oligonucleotides was evaluated by fluorescence microscopy. Inhibition of bcl-2 translation was demonstrated by quantitative RT-PCR and Western Blot. TUNEL assay, ANNEXIN V-binding and Apo-2.7 expression were performed to evaluate induction of apoptosis.Using polycationic liposomes, a ODN transfection rate of 95% in HeLa and 45% in DoHH2 cells were demonstrated by fluorescence microscopy. 24 hours after transfection quantitative RT-PCR detected a 56% decrease of bcl-2 mRNA in antisense and a 7% decrease in sense transfected DoHH2 cells (p0.05). In HeLa-cells, bcl-2 expression was almost completely inhibited 72 hours after antisense ODN transfection. Antisense treated cells also showed significant induction of apoptosis.Polycationic liposome-mediated transfection of bcl-2 antisense ODN's causes enhanced cellular uptake and efficient bcl-2 downregulation in bcl-2 overexpressing cell lines. This delivery strategy may explain why significant induction of apoptosis was achieved at low oligonucleotide concentrations (approximately 200 pmol/5 x 10(5) tumor cells). more...

The BCR-ABL fusion, the molecular equivalent of the Philadelphia translocation, gains importance for treatment stratification in adult acute lymphoblastic leukemia (ALL). In this prospective study, samples from 478 patients with CD10(+) B-cell precursor ALL (c-ALL and pre-B ALL) underwent BCR-ABL reverse transcription-polymerase chain reaction (RT-PCR) analysis with double testing of positive samples. Patients were stratified according to the PCR result and treated in 2 German Multicenter Trials of Adult ALL. The outcome was followed and the prognostic impact of BCR-ABL was compared to clinical risk features. Of the 478 samples, 432 had an evaluable BCR-ABL result. Thirty-seven percent of the c-ALL and pre-B ALL patients were BCR-ABL(+) (p190, 77%; p210, 20%; simultaneous p190/p210, 3%). BCR-ABL positivity was associated with the high-risk features of older age (45 years versus 30 years median age; P =.0001) and higher white blood cell counts (23 500/microL versus 11 550/microL; P =.0001). Univariate and multivariate analyses revealed BCR-ABL as the leading factor for a poor prognosis (P =.0001) in comparison to clinical risk criteria. Irrespective of the breakpoint, presence of any BCR-ABL transcript predicted a lower chance of initial treatment response (68.4% versus 84.6%; P =.001) and a lower probability of disease-free survival at 3 years (0.13 versus 0.47; P =.0001). This bad outcome was not influenced by postinduction high-dose treatment stratifications. The results show a high prevalence of BCR-ABL fusion transcripts with predominance of p190. BCR-ABL RT-PCR is confirmed as a sensitive, rapid method to diagnose t(9;22), and p190 and p210 are unequivocally demonstrated as the most important predictors of poor long-term survival despite intensified chemotherapy. more...

Summary: Background This trial evaluated the maximum tolerated dose (MTD), safety, pharmacokinetics, and activity of volasertib, a selective Polo-like kinase 1 inhibitor that induces mitotic arrest and apoptosis, combined with cisplatin or carboplatin in patients with advanced/metastatic solid tumors (NCT00969761; 1230.6). Methods Sequential patient cohorts (3+3 dose-escalation design) received a single infusion of volasertib (100-350 mg) with cisplatin (60-100 mg/m2) or carboplatin (area under the concentration versus time curve [AUC]4-AUC6) on day 1 every 3 weeks for up to six cycles. Sixty-one patients received volasertib/cisplatin (n=30) or volasertib/carboplatin (n=31) for a median of 3.5 (range, 1-6) and 2.0 (range, 1-6) treatment cycles, respectively. Results The most common cycle 1 dose-limiting toxicities (DLTs) were thrombocytopenia, neutropenia and fatigue. MTDs (based on cycle 1 DLTs) were determined to be volasertib 300 mg plus cisplatin 100 mg/m2 and volasertib 300 mg plus carboplatin AUC6. Co-administration did not affect the pharmacokinetics of each drug. Partial responses were observed in two patients in each arm. Stable disease was achieved in 11 and six patients treated with volasertib/cisplatin and volasertib/carboplatin, respectively. Conclusions Volasertib plus cisplatin or carboplatin at full single-agent doses was generally manageable and demonstrated activity in heavily pretreated patients with advanced solid tumors., SCOPUS: ar.j, info:eu-repo/semantics/published more...

The conditioning regimen prior to stem cell transplantation in 36 patients with high-risk acute myeloid leukemia (AML) and myelodysplastic syndrome (MDS) was intensified by treating patients with a rhenium 188–labeled anti-CD66 monoclonal antibody. Dosimetry was performed prior to therapy, and a favorable dosimetry was observed in all cases. Radioimmunotherapy with the labeled antibody provided a mean of 15.3 Gy of additional radiation to the marrow; the kidney was the normal organ receiving the highest dose of supplemental radiation (mean 7.4 Gy). Radioimmunotherapy was followed by standard full-dose conditioning with total body irradiation (12 Gy) or busulfan and high-dose cyclophosphamide with or without thiotepa. Patients subsequently received a T-cell–depleted allogeneic graft from a HLA-identical family donor (n = 15) or an alternative donor (n = 17). In 4 patients without an allogeneic donor, an unmanipulated autologous graft was used. Infusion-related toxicity due to the labeled antibody was minimal, and no increase in treatment-related mortality due to the radioimmunoconjugate was observed. Day +30 and day +100 mortalities were 3% and 6%, respectively, and after a median follow-up of 18 months treatment-related mortality was 22%. Late renal toxicity was observed in 17% of patients. The relapse rate of 15 patients undergoing transplantation in first CR (complete remission) or second CR was 20%; 21 patients not in remission at the time of transplantation had a 30% relapse rate. more...

NF-kappaB/rel transcription factors are crucial regulators of development, differentiation and apoptosis of both lymphoid and myeloid lineages. There is increasing evidence for an involvement of NF-kappaB/rel proteins in lymphomagenesis and resistance of lymphoid tumors to the induction of apoptosis. Structural alterations of the NF-kappaB/rel genes NFkappaB2, c-rel and bcl-3 have been shown to result in increased NF-kappaB/rel activity. Because we observed strong constitutive NF-kappaB/rel binding activity in chronic lymphocytic leukemia of the B-cell type (B-CLL) which may contribute to resistance against cytotoxic drugs we studied the genomic organisation of NFkappaB2, c-rel and bcl-3 gene loci in a panel of lymphoproliferative disorders (n=81) with an emphasis on B-CLL (n=47). The method of genomic Southern blotting using cDNAs of the respective genes was used. In spite of the role of NF-kappaB/rel in myeloid maturation there is no data available as to the occurrence of NF-kappaB/rel rearrangements in chronic myeloproliferative syndromes (cMPS). For this reason we included a small panel of cMPS patients (n=16). Southern Blotting revealed a germline configuration of NFkappaB2, c-rel and bcl-3 loci in all NHL and cMPS patients examined. Our results demonstrate that structural alterations of NFkappaB2, c-rel and bcl-3 genes at the Southern Blotting level are rare events that do not contribute to lymphoid or myeloid transformation in the majority of NHL or cMPS patients. more...

Polo-like kinases (Plks) play an important role in cell cycle checkpoint controls and are over-expressed in acute myeloid leukaemia (AML). BI 2536, a novel Plk inhibitor, induces mitotic arrest and apoptosis. In this phase I/II trial of BI 2536 in 68 elderly patients with relapsed/refractory AML, three schedules were investigated (day 1, days 1-3, and days 1 + 8). Maximum tolerated dose was 350 and 200 mg in the day 1 and days 1 + 8 schedules, respectively. The day 1-3 schedule appeared equivalent to the day 1 schedule and was discontinued early. BI 2536 exhibited multi-compartmental pharmacokinetic behaviour. The majority of patients showed an increase of bone marrow cells in G2/M with a characteristic pattern of mitotic catastrophe. The overall response rate in the day 1 and day 1 + 8 schedules was 9% (5/54) with 2 complete and 3 partial responses. The majority of drug-related adverse events grade ≥3 were haematological. Taken together, Plk inhibition induced cell cycle arrest in AML blasts in vivo and BI 2536 monotherapy showed modest clinical activity in this poor prognosis patient group. more...

11574Background: BRD4 is a key epigenetic regulator facilitating transcriptional elongation of super-enhancer regulated oncogenes including MYC. BI 894999, a novel, potent and selective orally bioa... more...

TPS8067Background: Myeloma is an incurable disease that typically follows a relapsing course, with many patients (pts) requiring multiple lines of therapy. Outcomes in RRMM remain poor, particularl... more...

BACKGROUND: H2-histamine receptors mediate a wide range of physiological functions extending from stimulation of gastric acid secretion to induction of human promyelocyte differentiation. We have previously cloned the H2-histamine receptor gene and noted that only three amino acids on the receptor were sufficient to define its specificity and selectivity. Despite only modest overall amino acid homology (34% amino acid identity and 57.5% similarity) between the H2-histamine receptor and the receptor for another monoamine, the beta 2-adrenergic receptor, there is remarkable similarity at their critical ligand binding sites. We hypothesized that, if the specificity and selectivity of both receptors are invested in just three amino acids, it should be possible to convert one of the receptors into one that recognizes the ligand of the other by simple mutations at only one or two sites. MATERIAL AND METHODS: We explored the effect of two single mutations in the fifth transmembrane domain of the H2-histamine receptor, which encompasses the sites that determine H2 selectivity. The canine H2 receptor gene was mutated at Asp186 and Gly187 (Asp186 to Ala186 and Gly187 to Ser187) by oligonuceotide directed mutagenesis. The coding region of both the wild-type and mutated H2 receptors was subcloned into the eukaryotic expression vector, CMVneo, and stably transfected into Hepa cells and L cells. The biological activity of histamine and epinephrine on the expressed receptor was examined by measurement of cellular cAMP production and inositol trisphosphate formation. RESULTS: Hepa cells transfected with the Ala186-Ser187 mutant H2 receptor demonstrated a biphasic rise in cAMP in response to epinephrine with an early phase (ED50 approximately 10(-11) M) that could be inhibited by both propranolol and cimetidine. Epinephrine also induced IP3 generation in the same cells, a biological response that is characteristic of activation of the wild-type H2 but not of the beta-adrenergic receptor. L cells transfected with the Ala186-Ser187 mutant H2 receptor also responded to epinephrine in a cimetidine and propranolol inhibitable manner. CONCLUSIONS: We converted the H2-histamine receptor into a bifunctional one that has characteristics of both histamine and adrenergic receptors by two simple mutations. These results support the hypothesis that ligand specificity is determined by only a few key points on a receptor regardless of the structure of the remainder of the molecule. Our studies have important implications on the design of pharmacological agents targeted for action at physiological receptors. more...

This open-label phase i study with an accelerated titration design was performed to determine the maximum tolerated dose of BI 2536, a potent, highly selective small-molecule polo-like kinase 1 (Plk1) inhibitor. Patients with advanced solid tumours received a single 60-minute intravenous infusion of BI 2536 (50&ndash, 70 mg) on days 1&ndash, 3 of each 21-day treatment course. Recipients without disease progression or untenable toxicity could receive additional treatment courses. The maximum tolerated dose was determined based on dose-limiting toxicities. Other assessments included safety, pharmacokinetic profile, and antitumour activity according to the Response Evaluation Criteria in Solid Tumors. The study enrolled 21 patients. The maximum tolerated dose for BI 2536 was determined to be 60 mg for the study schedule. Dose-limiting toxicities included hematologic events, hypertension, elevated liver enzymes, and fatigue. The most frequently reported drug-related adverse events were mild-to-moderate fatigue, leukopenia, constipation, nausea, mucosal inflammation, anorexia, and alopecia. The pharmacokinetics of BI 2536 were linear within the dose range tested. Plasma concentration profiles exhibited multi-compartmental pharmacokinetic behaviour, with a terminal elimination half-life of 20&ndash, 30 hours. In the present study, BI 2536 showed an acceptable safety profile warranting further investigation of Plk1 inhibitors in this patient population. more...

Introduction BI 2536 is a potent, highly selective inhibitor of polo-like kinase (Plk) 1. This open-label, phase I study investigated the maximum tolerated dose (MTD), safety, efficacy, and pharmacokinetics (PK) of BI 2536 IV in combination with standard-dose pemetrexed in previously treated advanced or metastatic non–small-cell lung cancer. Patients and Methods A standard 3 + 3 design was used. The patients received 500 mg/m 2 pemetrexed and escalating doses of BI 2536 on day 1 every 3 weeks. The primary objective was the MTD of BI 2536 combined with pemetrexed. Secondary endpoints were response rate (Response Evaluation Criteria in Solid Tumors), overall safety, and PK. Results Forty-one patients received BI 2536 (100-325 mg). Two dose-limiting toxicities (DLT) occurred at BI 2536 325 mg (grade 3 pruritus and rash; grade 4 neutropenia). Therefore, the MTD for BI 2536 in combination with pemetrexed was 300 mg. After expanding the MTD dose level, 3 additional patients experienced DLTs, which resulted in expansion of the 250 mg cohort, in which 4 of the 13 additional patients experienced DLTs. Therefore, the recommended dose of BI 2536 was 200 mg. Most frequently reported drug-related adverse events were fatigue (71%), nausea (37%), and rash (34%). Two patients had durable confirmed partial responses; 21 (54%) patients had stable disease after the treatment cycle 2. PK analysis showed that BI 2536 and pemetrexed exposure were not altered when coadministered. Conclusion BI 2536 200 mg combined with standard-dose pemetrexed has an acceptable safety profile in relapsed non–small-cell lung cancer. The antitumor activity observed is encouraging and supports further investigation of Plk inhibitors. more...

Purpose: This phase I, open-label, dose-escalation study investigated the maximum tolerated dose (MTD) of BI 2536, a small-molecule polo-like kinase (Plk)–1 inhibitor, in two treatment schedules in patients with advanced solid tumors. Secondary objectives included evaluation of safety, efficacy, and pharmacokinetics. Experimental Design: Patients received a single i.v. dose of BI 2536 as a 1-hour infusion on days 1 and 8 or a single 24-hour infusion on day 1 of each 21-day treatment course. MTD determination was based on dose-limiting toxicities. Results: Forty-four and 26 patients received each treatment schedule, respectively. The MTD of BI 2536 in the day 1 and 8 schedule was 100 mg per administration (200 mg per course). The MTD for the second dosing schedule was not determined; a 225-mg dose was well tolerated. The most frequently reported treatment-related nonhematologic adverse events were gastrointestinal events and fatigue. Hematotoxicity as the most relevant side effect was similar in both schedules; neutropenia grades 3 and 4 were observed in 16 patients (36.4%) of the day 1 and 8 schedule and 13 patients (50%) of the 24-hour infusion. Fourteen patients (32%) treated in the day 1 and 8 dosing schedule had a best overall response of stable disease. Plasma concentrations of BI 2536 increased dose proportionally, with no relevant accumulation of exposure in the day 1 and 8 dosing schedule. The average terminal half-life was 50 hours. Conclusions: BI 2536 administered in either treatment schedule has adequate safety in patients with advanced solid tumors, warranting further clinical investigation of polo-like kinase–1 inhibitors. Clin Cancer Res; 16(18); 4666–74. ©2010 AACR. more...

To determine the maximum tolerated dose (MTD) and dose-limiting toxicities (DLTs) of BIBF 1120, a triple angiokinase inhibitor administered once-daily in patients with advanced multiple myeloma.This Phase I study included 17 patients. Planned dose escalations of BIBF 1120 were 100, 200, 250 and 300 mg. Safety and pharmacokinetic (PK) assessments were performed.Two DLTs (200 and 250 mg) occurred due to increased gamma-glutamyltransferase levels (CTC grade 3). The 250 mg dose was well tolerated; no dose escalation beyond 250 mg was made. The most common adverse events included diarrhoea, nausea and vomiting. No detectable deviation from dose linear PKs was observed. Regarding tumour control, two patients had stable disease foror = 4 months.BIBF 1120 was safe and well tolerated up to 250 mg/day. The MTD was not reached. more...

Purpose: Novel drugs including targeted approaches have changed treatment paradigms for multiple myeloma (MM) and may also have therapeutic potential in the poor-prognosis t(4;14) subset; t(4;14) results in overexpressed and activated fibroblast growth factor receptor 3 (FGFR3). Blocking this receptor tyrosine kinase (RTK) induces apoptosis in t(4;14)+ MM cells and decreases adhesion to bone marrow stromal cells (BMSC). Using combinations of novel drugs, we investigated potential enhancement of single-agent activities within the tumor cells, targeting of the marrow micromilieu, or circumvention of drug resistance in t(4;14)+ MM. Experimental Design: We tested effects on apoptosis and related signaling pathways in the t(4;14)+ MM subset, applying drug combinations including a FGFR3 tyrosine kinase inhibitor (RTKI), the proteasome inhibitor bortezomib, and dexamethasone. Results: RTKI, bortezomib, and dexamethasone were active as single agents in t(4;14)+ MM. RTK inhibition triggered complementary proapoptotic pathways (e.g., decrease of Mcl-1, down-regulation of p44/42 mitogen-activated protein kinase, and activation of proapoptotic stress-activated protein/c-Jun NH2-terminal kinases). Synergistic or additive effects were found by combinations of RTKI with dexamethasone or bortezomib. In selected cases of t(4;14)+ MM, triple combinations were superior to dual combinations tested. Prevention from MM cell apoptosis by BMSC or exogenous interleukin-6 was circumvented by drug combinations. In t(4;14)+, N-ras–mutated NCI-H929 cells, resistance to RTKI was overcome by addition of dexamethasone. Notably, the combination of RTKI and dexamethasone showed additive proapoptotic effects in bortezomib-insensitive t(4;14)+ MM. Conclusions: Combining novel drugs in poor-prognosis t(4;14)+ MM should take into account at least bortezomib sensitivity and probably Ras mutational status. more...

In multiple myeloma (MM), both vascular endothelial (VEGF) and basic fibroblast growth factor (bFGF) promote tumor growth and survival. We have used the novel indolinone BIBF 1000 to study effects of simultaneous inhibition of VEGF, FGF and transforming growth factor-β on MM cells and their interactions with bone marrow stroma cells (BMSCs). Both, in the absence and presence of myeloma-stroma cell contacts, BIBF 1000 abrogated BMSC-derived secretion of interleukin-6 (IL-6). In addition, BIBF 1000 directly induced apoptosis in t(4;14)–positive cell lines as well as in CD138+ marrow cells from patients with t(4;14) myeloma. To a similar extent, BIBF 1000 induced apoptosis in MM.1S and MM.1R cells carrying the translocation t(14;16). In case of MM.1S and other dexamethasone-sensitive t(14;16) cell lines, BIBF 1000 and dexamethasone had additive proapoptotic effects. Induction of apoptosis by BIBF 1000 was associated with inhibition of the mitogen-activated protein kinases (MAPK) pathway in t(4;14) and inhibition of the phosphatidyl-inositol-3 kinase/AKT pathway in t(14;16) cells. Apoptotic effects did not occur in t(4;14)–or t(14;16)–positive MM cells carrying n- or k-Ras mutations. The data provide the rationale for clinical evaluation of this class of targeted kinase inhibitors in MM with focus on defined cytogenetic subgroups. more...

The Bcr-Abl translocation t(9;22)(q34;q11 ) defines a subgroup of ALL patients with a dismal prognosis despite the introduction of intensified induction and consolidation regimen. Although Bcr-Abl induced NF-kappaB/Rel activation has previously been shown, the role of NF-kappaB/Rel in Ph+ leukemia is unclear. Using DNA binding assays, we demonstrate constitutive NF-kappaB/Rel activity in nuclear extracts from Ph+ ALL blasts, whereas Ph- primary blast cells and B-precursor cell lines lack NF-kappaB/Rel activity. NF-kappaB/Rel activity was shown in the ela2 and the b2a2 subtypes. Constitutive NF-kappaB/Rel activity in Ph+ blasts is not due to elevated endogenous IkappaB kinase (IKK) activity as shown by immune complex kinase assays. Since NF-kappaB/Rel is a transcriptional regulator of inhibitors of apoptosis we examined the expression of anti-apoptotic genes known to be induced by NF-kappaB/Rel by real time PCR analysis. We found no induction of TRAFI, TRAF2, cIAPI, cIAP2, XIAP, A20 or Bfl/Al in Ph+ ALL samples as compared to Ph-negative ALL controls. In summary, constitutive NF-kappaB/Rel activation independent of endogenous IKK activation may be a common finding in Ph+ ALL. However, targets of NF-kappaB/Rel mediated transcriptional regulation in this disease remain to be identified. more...

From December 1999 until July 2001, a phase I dose escalation study was performed with (186)Re-labeled bivatuzumab, a humanized monoclonal antibody against CD44v6, on patients with inoperable recurrent or metastatic head and neck cancer. The aim of the trial was to assess the safety and tolerability of intravenously administered (186)Re-bivatuzumab and to determine the maximum tolerated dose (MTD) of (186)Re-bivatuzumab. The data were also used for dosimetric analysis of the treated patients. Dosimetry is used to estimate the absorbed doses by nontarget organs, as well as by tumors. It can also help to explain toxicity that is observed and to predict organs at risk because of the therapy given.Whole-body scintigraphy was used to draw regions around sites or organs of interest. Residence times in these organs and sites were calculated and entered into the MIRDOSE3 program, to obtain absorbed doses in all target organs except for red marrow. The red marrow dose was calculated using a blood-derived method. Twenty-one studies on 18 patients, 5 female and 16 male, were used for dosimetry.The mean red marrow doses were 0.49 +/- 0.03 mGy/MBq for men and 0.64 +/- 0.03 mGy/MBq for women. The normal organ with the highest absorbed dose appeared to be the kidney (mean dose, 1.61 +/- 0.75 mGy/MBq in men and 2.15 +/- 0.95 mGy/MBq in women; maximum kidney dose in all patients, 11 Gy), but the doses absorbed are not expected to lead to renal toxicity. Other organs with doses exceeding 0.5 mGy/MBq were the lungs, the spleen, the heart, the liver, the bones, and the testes. The doses delivered to the tumor, recalculated to the MTD level of 1.85 GBq/m(2), ranged from 3.8 to 76.4 Gy, with a median of 12.4 Gy. A good correlation was found between platelet and white blood cell counts and the administered amount of activity per kilogram of body weight (r = -0.79).Dosimetric analysis of the data revealed that the range of doses to normal organs seems to be well within acceptable and safe limits. Tumor doses ranged from 4 to 76 Gy. Given the acceptable tumor doses, (186)Re-labeled bivatuzumab could be a good candidate for future adjuvant radioimmunotherapy in patients with minimal residual disease. more...

The human CD44 gene encodes type 1 transmembrane glycoproteins involved in cell-cell and cell-matrix interactions. The structural heterogeneity of the gene products is caused primarily by alternative splicing of at least 10 out of 20 exons. Certain CD44 variant isoforms, in particular those containing CD44 variant domain 6 (CD44v6), have been implicated in tumourigenesis, tumour cell invasion and metastasis. Here we will give an overview of immunohistochemically determined CD44v6 expression in human malignancies (primary epithelial and nonepithelial tumours as well as metastases) and normal tissues, and review several examples of the clinical use of CD44v6-specific antibodies. In nonmalignant tissues, CD44v6 expression is essentially restricted to a subset of epithelia. Intense and homogeneous expression of CD44v6 was reported for the majority of squamous cell carcinomas and a proportion of adenocarcinomas of differing origin, but was rarely seen in nonepithelial tumours. This expression pattern has made CD44v6 an attractive target for antibody-guided therapy of various types of epithelium-derived cancers. more...

B-cell chronic lymphocytic leukemia (B-CLL) is characterized by a resistance toward apoptosis-inducing agents. Nuclear factor-kappaB (NF-kappaB)/Rel has been shown to regulate the expression of antiapoptotic genes, such as members of the inhibitor of apoptosis protein (IAP) and tumor necrosis factor receptor-associated factor (TRAF) gene families. Expression and regulation of NF-kappaB/Rel-dependent inhibitors of apoptosis have not been collectively studied in B-CLL. We examined expression of known NF-kappaB/Rel-regulated antiapoptotic genes by RNAse protection assay, real-time polymerase chain reaction, and immunoblotting in patients with B-CLL. TRAF1 and to a lesser extent TRAF2 were overexpressed in B-CLL lymphocytes as compared with normal CD19(+) B cells. TRAF1 overexpression did not correlate with markers of disease progression or overall survival. Furthermore, we found high constitutive expression of the IAP genes c-IAP-1, c-IAP-2, and XIAP both in normal and B-CLL lymphocytes. Focusing on the regulation of TRAF1, NF-kappaB/Rel activity in B-CLL nuclear extracts was shown to bind to TRAF1 promoter elements. However, IkappaB kinase (IKK) activity was not increased in CLL lymphocytes as compared with normal CD19(+) B cells. The known IKK inhibitor sulfasalazine did not compromise TRAF1 expression. Thus, although our study revealed a common expression pattern of NF-kappaB/Rel-regulated inhibitors of apoptosis, our findings indicate an IKK-independent regulation of TRAF1 in B-CLL. more...

7557 Background: The benefit of sustained ErbB family blockade in NSCLC patients with acquired resistance (AR) to EGFR TKIs is unknown. We investigated afatinib, an irreversible blocker of EGFR (ErbB1), HER2 (ErbB2) and ErbB4 receptor tyrosine kinases, in patients with metastatic NSCLC, who had failed chemotherapy and E/G. Methods: This was a Phase III, randomized, open-label, multi-center trial. Patients with pathologically confirmed Stage IIIB/IV metastatic NSCLC after ≥1 line of chemotherapy who failed E/G received oral afatinib 50 mg until disease progression (Part A). After progression, patients with clinical benefit (≥12 wks) were eligible to continue afatinib 40 mg plus paclitaxel or receive investigator’s choice chemotherapy (Part B). Primary endpoint for Part A was PFS (RECIST 1.1; CT scan every 6 wks). Available tumor samples were collected for central EGFR mutation testing; local mutation data were also collected. An interim analysis of Part A, assessing afatinib monotherapy, is reported. Results: Part A enrolled April 2010 through to May 2011; 1154 patients received afatinib monotherapy. The majority had adenocarcinoma (85%), 57% were female, 43% were Asian, 54% were never smokers. Best response to prior E/G was CR (2%), PR (31%), SD (42%) and PD (20%). Median PFS for afatinib was 3.3 mths; 88 patients (8%) achieved an objective tumor response, 648 (56%) had SD. For EGFR mutation positive patients (n=49, centrally confirmed), PFS was 4.2 vs. 2.6 mths for EGFR mutation negative patients (n=35). When applying clinical enrichment criteria for AR, PFS was 4.2 mths for those with enrichment (n=597) vs. 2.8 mths for those without (n= 557; logrank test p more...

3018^ Background: Volasertib (V) is a first in class, selective and potent cell cycle kinase inhibitor that induces mitotic arrest and apoptosis by targeting Plk. This phase I study evaluates dose-limiting toxicities (DLT), maximum tolerated dose (MTD), safety and pharmacokinetics (PK) of V combined with cisplatin (Cis) or carboplatin (Ca). Methods: Sequential cohorts of patients (pts) received a single 2h infusion of V with Cis (arm A) or Ca (arm B) every 3 wks. Cis and Ca were given for up to 6 cycles (Cy); V was continued until progression or intolerance. MTD was the highest dose at which ≤1/6 pts experienced a DLT in Cy 1. MTD cohorts were expanded to 12 DLT-evaluable pts to further characterize safety. Results: As of January 11 2012, 61 pts (arm A: 30; arm B: 31) were treated. Pt characteristics were (arm A/B): median age 55/58 yrs, male 16/18 pts, ECOG PS 0: 13/14 pts, PS 1: 17/17 pts. Tumors included (pts): non-small cell lung cancer (15); sarcoma (8); colorectal cancer (6); melanoma (4); urothelial cancer (4); other (24). Pts received V + Cis for a median [range] of 3.5 Cy [1-6], V + Ca for 2 Cy [1-6] and V for 3.5 Cy [1-20] in arm A and 2 Cy [1-14] in B. PK analyses are ongoing. MTD was reached at V 300 mg + Cis 100 mg/m2 and V 300 mg + Ca AUC 6. Five partial responses (PR) were seen. 15/30 pts in arm A and 12/31 in B achieved stable disease (SD) or PR. PR or SD for >6 Cy was observed in 6/30 pts and 5/31 pts in arms A and B, respectively. Conclusions: In this phase I study, V in combination with Cis or Ca at full single-agent doses was well tolerated. Furthermore, several objective responses and cases of sustained SD were observed in heavily pretreated pts with advanced solid tumors.[Table: see text] more...

Adrenocorticotropic hormone (ACTH) and [alpha]-, [beta]-, and [gamma]-melanocyte-stimulating hormone (MSH) are products of propiomelanocortin post-translational processing. These compounds are collectively labeled as melanocortins (MC). Aside from their established effects on the regulation of the adrenal cortex (ACTH) and melanocytes ([alpha]-MSH), the melanocortins have been implicated in a broad array of physiological events. Melanocortins mediate their effects through cell membrane receptors belonging to the superfamily of seven transmembrane G-protein-linked receptors. Using the technique of polymerase chain reaction with primers based on conserved areas of the seven transmembrane G-protein-linked receptor family, the authors recently isolated an [open quotes]orphan[close quotes] subfamily of this receptor group. Within the past year, two of these receptors were identified as specific for [alpha]-MSH (MC1) and ACTH (MC2). They have recently described a third melanocortin receptor (MC3) that appears to recognize the core heptapeptide sequence of melanocortins with equal potency and efficacy and identified its presence in the brain, placenta, and gut. Using the FISH technique, they localized the ACTH and the melanocortin-3 receptors to chromosome loci 18p11.2 and 20q12.3-q13.2, respectively. 19 refs., 1 fig. more...

Using the technique of the polymerase chain reaction primed with oligonucleotides based on the homologous transmembrane regions of seven transmembrane G protein-linked receptors, we isolated three full-length human genes that encode a novel subgroup of this receptor family. Recently, two of these receptors were identified as specific for alpha-melanocyte-stimulating hormone (alpha-MSH) and adrenocorticotropic hormone. We report the molecular cloning and pharmacologic characterization of a third member of this subgroup. The gene for this receptor encodes a protein of 361 amino acids in length. Its pharmacology characterizes it as an MSH receptor specific to the heptapeptide core common to adrenocorticotropic hormone and alpha-, beta-, and gamma-MSH. By Northern blot hybridization and polymerase chain reaction, it is expressed in brain, placental, and gut tissues but not in melanoma cells or in the adrenal gland. These findings may yield insight into the physiology of peptides derived from pro-opiomelanocortin post-translational processing. more...

3061 Background: BI 6727 is a highly potent and selective inhibitor of the serine-threonine Polo-like kinase 1 (Plk1), a key regulator of cell cycle progression. BI 6727 is a second generation dihy... more...

We undertook these studies to characterize the molecular basis of the interaction of histamine with the H2 receptor. Key areas of homology in the structures of the histamine H2 and beta 2 adrenergic receptor suggested specific transmembrane amino acids that might be important for binding of histamine. A third transmembrane aspartic acid of the histamine receptor (Asp98), thought to serve as a counter anion that interacts with the cationic amine moiety of histamine, was mutated to Asn98, and the mutated receptor was expressed in Hepa cells. Removal of the negatively charged amino acid abolished both binding of the H2 receptor antagonist [methyl-3H]tiotidine and histamine stimulated increases in cellular cAMP content. Mutation of a fifth transmembrane aspartic acid (Asp186) to Ala186 or Asn186 by itself or in conjunction with mutation of another fifth transmembrane amino acid (Thr190 to Ala190) resulted in a loss of [methyl-3H] tiotidine binding, although the generation of cAMP in response to histamine was maintained. The histamine receptor with only a Thr190 to Ala190 or Cys190 mutation retained the ability to bind [methyl-3H]tiotidine, but both the affinity and efficacy of binding were reduced. These data lead us to propose a model for histamine binding in which Asp98 is essential for histamine binding and action, Asp186 defines H2 selectivity, and Thr190 is important in establishing the kinetics of histamine binding, but is not essential for H2 selectivity. more...

8108 Background: BI 2536 is a potent, selective inhibitor of polo-like kinase 1 (Plk1), a regulator of mitotic progression. BI 2536 demonstrated favorable tolerability and antitumor activity in phase I trials. We investigated the antitumor efficacy, safety and PK of BI 2536 in patients (pts) with sensitive relapse SCLC. Methods: This open label single arm phase II study followed a Gehan two-stage design. Primary objective was to determine the antitumor efficacy of BI 2536 in SCLC pts with disease recurrence ≥60 days after completion of first-line chemotherapy. 18 pts had to complete 2 courses to be evaluable for stage 1 analysis. In case of ≥2 partial or complete antitumor responses (RECIST criteria), stage 2 accrual would continue until 40 pts were entered. Patients received 200 mg BI 2536 as a 1h i.v. infusion on Day 1 every 3 weeks. Dose escalation to 250 mg (cycle 3 onwards) was encouraged in pts with 60 days after completion of first-line therapy. Of 23 pts, no objective antitumor responses were observed, 7 had stable disease as best response, 14 had progression, 2 were not evaluable. A median of 2 courses were given, up to a maximum of 12 in 1 pt. The PFS rate at 3 months was 25%. Due to the lack of antitumor responses, trial accrual was terminated after stage 1. Overall, BI 2536 was well tolerated. Frequent AEs were neutropenia (48%), fatigue (39%), nausea (30%), anemia, vomiting, constipation (26% each), and thrombocytopenia (22%). Drug related grade 3/4 AEs were neutropenia (13%/26%), grade 3/4 thrombocytopenia (1 pt each), grade 3/4 anemia (1 pt each), grade 4 sepsis (1 pt), Grade 4 ARDS (1 pt) and Grade 3 fatigue (1 pt). PK analyses indicate that BI 2536 has high clearance (>1,000 mL/min) and quickly distributes in multiple compartments in a large volume of distribution (>1,000 L). Estimated elimination half-life was >25 h. Conclusions: BI 2536 was well tolerated in relapsed SCLC pts, but demonstrated no convincing antitumor efficacy after stage I of the study. Therefore, BI 2536 will not be assessed further as a single agent in SCLC. [Table: see text] more...

We utilized the technique of polymerase chain reaction with oligonucleotide primers based upon the nucleotide sequence of the canine H2 histamine receptor gene which we recently isolated to clone its human homologue. Transfection of a construct of this gene in Colo-320 DM cells led to the expression of a receptor that bound to [methyl-3H] tiotidine and was linked to 3′,5′ cyclic adenosine monophosphate (cAMP) generation in response to histamine. Both cAMP generation and [methyl-3H] tiotidine binding were inhibited with the H2 histamine receptor selective antagonist cimetidine but not diphenhydramine or thioperamide which are, respectively, H1 and H3 histamine receptor antagonists. These data confirm that we have successfully cloned a novel gene encoding the human H2 histamine receptor. more...

Background: BI 2536 is a highly potent, selective inhibitor of Polo-like kinase 1 (Plk1), a key regulator of mitotic progression. BI 2536 has demonstrated favorable tolerability and antitumor activity in Phase I trials in patients with solid tumors. Antitumor activity of BI 2536 was also shown in preclinical non-Hodgkin’s lymphoma (NHL) models. We determined the maximum tolerated dose (MTD), overall safety, pharmacokinetics (PK) and efficacy of BI 2536 given as an intravenous infusion once every 3 weeks in patients with relapsed or refractory aggressive NHL of T- or B-cell origin. Methods: Sequential cohorts of 3–6 patients with relapsed or refractory aggressive NHL received 1-hour infusions of BI 2536 following a toxicity-guided Phase 1 doseescalation design. Patients relapsed after peripheral stem cell transplantation and transplantation-naive patients were entered into different strata and the respective MTD determined independently. A single administration was given every 21 days. Patients with clinical benefit were eligible for further treatment courses after recovery from toxicity after a 3-week observation period. A total of 41 patients were entered into the trial: 24 patients in the transplant-naive (non-tr) stratum; and 17 patients in the transplant-failure (tr) stratum. Patients were treated at dose levels from 50 to 200 mg. Results: The safety profile was similar in both strata with the MTD determined independently at 175 mg for both non-tr and tr patients. Neutropenia (tot: 33%; CTCAE Grade (gr)3/4: 21%), anemia (tot: 29%; gr3: 4%), thrombocytopenia (tot: 29%; gr3/4: 17%), fatigue (tot: 25%; gr3: 4%) and nausea (tot=gr1/2: 25%) were the most frequent adverse events in non-tr; and thrombocytopenia (tot: 59%; gr3/4: 41%), anemia (tot=gr1/2: 41%), fatigue (tot=gr1/2: 41%) and neutropenia (tot: 41%; gr3/4: 21%) were most frequent in tr patients. Dose-limiting toxicities (DLTs) consisted of reversible thrombocytopenia (six patients) and neutropenia (three patients). No relevant non-specific toxicity was observed. Pharmacokinetic analysis showed dose proportionality of Cmax and AUC0–∞ with a high clearance (~1,400 mL/min) and a high volume of distribution (>1,000 L). Patients were treated for up to 6 courses without evidence of cumulative toxicity. Three complete responses (CRs) and one partial response were observed. Stable disease as best response was noted in three (18%) of tr patients and nine (38%) of non-tr patients. All responders had relapsed after prior peripheral stem cell transplants and were treated at doses of 150–200 mg. Three of the four responders had a peripheral T-cell lymphoma (PTCL) NHL; one CR was observed in a patient with diffuse large B-cell lymphoma. The overall response rate (ORR) in the tr stratum was 23.5%; in the aggregate of both tr and non-tr, the ORR amounted to 9.7%. With three out of five patients responding, an ORR of 60% was observed in the T-cell subset. However, the responses were of short duration. Conclusion: BI 2536 has a favorable safety and PK profile in patients with NHL. Safety profile and PK properties are comparable to data obtained in solid tumor patient populations. Encouraging, albeit transient, anti-lymphoma efficacy was observed in patients suffering from PTCL after autologous stem-cell transplantation. more...

Background: BI 2536 is a potent and selective inhibitor of Plk-1, which plays a crucial role in the regulation of mitosis. Inhibition of Plk-1 leads to mitotic arrest and apoptosis. Therefore, Plk-1 inhibitors are currently in clinical testing as anti-proliferative agents in cancer patients. BI 2536, the first specific Plk-1 inhibitor in clinical testing, demonstrated strong anti-proliferative effects on AML cell lines in vitro and in in vivo models. To investigate target inhibition in the malignant cell, bone marrow samples from patients who participated in a Phase I/II study of BI 2536 single-agent therapy in elderly patients with relapsed or refractory AML were analyzed. Methods: Pharmacodynamic analyses including immunocytochemistry (ICC) and flow cytometry (FACS) of bone marrow (BM) were performed to examine cell morphology, phosphorylation of histone H3 (phospho-H3), and induction of apoptosis. Samples were acquired before and 24 h after the first administration of BI 2536. FACS analysis included propidium iodide (PI)-FACS to determine the percentage of cells residing in various cell cycle stages (G0/1-, S- and G2/M-phases) and Annexin V-Cy5-staining for apoptosis. Immunocytochemistry included staining for phosphorylated histone H3 and TUNEL assay for apoptosis. Results: At the time of analysis, data from 28 patients treated at doses in the range of 50 to 400 mg BI 2536 were available. BM taken after BI 2536 administration showed an increase of phospho-H3 positive cells as compared to baseline prior to treatment. There was a trend toward a positive correlation between phospho-H3 increase and increase in dosage of BI 2536 (p=0.16) when treatment at low doses (50 to 60 mg) of BI 2536 were compared to treatment at higher doses (100 to 400 mg). Furthermore, trends were observed that an increase of phospho-H3 is positively correlated with both a higher percentage of cells in G2/M and an increase in apoptotic cells. The typical morphology of cells in mitotic arrest could be demonstrated by ICC. Interestingly, when patients with progressive disease after one cycle (PD) were compared to patients with disease stabilization or response (nonPD), a statistically significant positive correlation between PD and increase in phospho-H3 compared to baseline was found (p=0.03). Also, there was a clear trend for an increase in the percentage of cells in G2/M phase and apoptosis in patients with PD compared to patients with nonPD. Preliminary evaluation of other disease characteristics including karyotype (normal vs complex), secondary AML, complete response in previous treatments, baseline value for blasts in the bone marrow or in the peripheral blood, did not reveal any signs of correlation with the clinical response to BI 2536 treatment. Conclusion: BI 2536 treatment increases the number of cells in G2/M phase (as detected by FACS analysis and phospho-H3 staining) and the number of apoptotic cells within 24 h after administration. In line with the clinical observation of rapid blast reduction both in the BM and the peripheral blood, these findings indicate that BI 2536 induces mitotic arrest and apoptosis of the malignant target cells in AML patients. The biological meaning of the correlation between the BM findings and the clinical response to BI 2536 is unclear, but if substantiated by more data, these results may suggest a predictive value of BM examinations for the response to BI 2536. more...