Srinivasan Madhusudan

Cisplatin plus Gemcitabine versus Gemcitabine for Biliary Tract Cancer

BACKGROUND There is no established standard chemotherapy for patients with locally advanced or metastatic biliary tract cancer. We initially conducted a randomized, phase 2 study involving 86 patients to compare cisplatin plus gemcitabine with gemcitabine alone. After we found an improvement in progression-free survival, the trial was extended to the phase 3 trial reported here. METHODS We randomly assigned 410 patients with locally advanced or metastatic cholangiocarcinoma, gallbladder cancer, or ampullary cancer to receive either cisplatin (25 mg per square meter of body-surface area) followed by gemcitabine (1000 mg per square meter on days 1 and 8, every 3 weeks for eight cycles) or gemcitabine alone (1000 mg per square meter on days 1, 8, and 15, every 4 weeks for six cycles) for up to 24 weeks. The primary end point was overall survival. RESULTS After a median follow-up of 8.2 months and 327 deaths, the median overall survival was 11.7 months among the 204 patients in the cisplatin-gemcitabine group and 8.1 months among the 206 patients in the gemcitabine group (hazard ratio, 0.64; 95% confidence interval, 0.52 to 0.80; P<0.001). The median progression-free survival was 8.0 months in the cisplatin-gemcitabine group and 5.0 months in the gemcitabine-only group (P<0.001). In addition, the rate of tumor control among patients in the cisplatin-gemcitabine group was significantly increased (81.4% vs. 71.8%, P=0.049). Adverse events were similar in the two groups, with the exception of more neutropenia in the cisplatin-gemcitabine group; the number of neutropenia-associated infections was similar in the two groups. CONCLUSIONS As compared with gemcitabine alone, cisplatin plus gemcitabine was associated with a significant survival advantage without the addition of substantial toxicity. Cisplatin plus gemcitabine is an appropriate option for the treatment of patients with advanced biliary cancer. (ClinicalTrials.gov number, NCT00262769.)

Docetaxel versus active symptom control for refractory oesophagogastric adenocarcinoma (COUGAR-02): an open-label, phase 3 randomised controlled trial

BACKGROUND Second-line chemotherapy for patients with oesophagogastric adenocarcinoma refractory to platinum and fluoropyrimidines has not shown benefits in health-related quality of life (HRQoL). We assessed whether the addition of docetaxel to active symptom control alone can improve survival and HRQoL for patients. METHODS For this open-labelled, multicentre trial, we recruited patients aged 18 years or older from 30 UK centres. Patients were eligible if they had an advanced, histologically confirmed adenocarcinoma of the oesophagus, oesophagogastric junction, or stomach that had progressed on or within 6 months of treatment with a platinum-fluoropyrimidine combination. Patients could have an Eastern Cooperative Oncology Group performance status of 0-2. We randomly assigned patients using a central, computerised minimisation procedure to receive docetaxel plus active symptom control, or active symptom control alone (1:1; stratified by disease status, disease site, duration of response to previous chemotherapy, and performance status). Docetaxel was given at a dose of 75 mg/m(2) by intravenous infusion every 3 weeks for up to six cycles. The primary endpoint was overall survival, analysed by intention to treat. This is the report of the planned final analysis. This study is an International Standardised Randomised Controlled Trial, number ISRCTN13366390. FINDINGS Between April 21, 2008, and April 26, 2012, we recruited 168 patients, allocating 84 to each treatment group. After a median follow-up of 12 months [IQR 10-21]) and 161 (96%) deaths (80 in the docetaxel group, 81 in the active symptom control group), median overall survival in the docetaxel group was 5.2 months (95% CI 4.1-5.9) versus 3.6 months (3.3-4.4) in the active symptom control group (hazard ratio 0.67, 95% CI 0.49-0.92; p=0.01). Docetaxel was associated with higher incidence of grade 3-4 neutropenia (12 [15%] patients vs no patients), infection (15 [19%] patients vs two [3%] patients), and febrile neutropenia (six [7%] patients vs no patients). Patients receiving docetaxel reported less pain (p=0.0008) and less nausea and vomiting (p=0.02) and constipation (p=0.02). Global HRQoL was similar between the groups (p=0.53). Disease specific HRQoL measures also showed benefits for docetaxel in reducing dysphagia (p=0.02) and abdominal pain (p=0.01). INTERPRETATION Our findings suggest that docetaxel can be recommended as an appropriate second-line treatment for patients with oesophagogastric adenocarcinoma that is refractory to treatment with platinum and fluoropyrimidine. FUNDING Cancer Research UK.

A phase II study of etanercept (Enbrel), a tumor necrosis factor alpha inhibitor in patients with metastatic breast cancer.

Purpose: Tumor necrosis factor (TNF) α is a key player in the tumor microenvironment and is involved in the pathogenesis of breast cancer. Etanercept is a recombinant human soluble p75 TNF receptor that binds to TNF-α and renders it biologically unavailable. In the current study, we sought to determine the toxicity, biological activity, and therapeutic efficacy of Etanercept in metastatic breast cancer. Experimental Design: We initiated a Phase II, nonrandomized, open-labeled study in patients with progressive metastatic breast cancer refractory to conventional therapy (Phase I toxicity data were available in patients with rheumatoid arthritis). Etanercept was administered subcutaneously at a dose of 25 mg twice weekly until disease progression. Results: Sixteen patients were recruited [median age 53 years (range, 34 to 74)]. A total of 141.6 weeks of therapy was administered (median of 8.1 weeks). Seven patients received ≥12 weeks of therapy. The most common side effects were injection site reactions (6), fatigue (5), loss of appetite (2), nausea (1), headache (1), and dizziness (1). Brief period of disease stabilization was seen in 1 patient lasting for 16.4 weeks. Immunoreactive TNF-α was elevated within 24 hours of therapy and persisted until the end of treatment (days 7, 28, 56, and 84). Phytohemagglutinin stimulates the production of interleukin-6 and CCL2 in peripheral blood cells, and the ability of Etanercept to modulate this response was assessed in a cytokine release assay. A consistent decrease in interleukin-6 and CCL2 level was seen compared with pretreatment values in serial blood samples (days 1, 7, 28, 56, and 84). Conclusions: Our study shows the safety and biological activity of Etanercept in breast cancer and provides data to assess pharmacodynamic endpoints of different schedules of Etanercept and combinations with chemotherapy or other biological therapies.

Human AP endonuclease 1 (APE1): from mechanistic insights to druggable target in cancer.

DNA base excision repair (BER) is critically involved in the processing of DNA base damage induced by alkylating agents. Pharmacological inhibition of BER (using PARP inhibitors), either alone or in combination with chemotherapy has recently shown promise in clinical trials. Human apurinic/apyrimidinic endonuclease 1(APE1) is an essential BER protein that is involved in the processing of potentially cytotoxic abasic sites that are obligatory intermediates in BER. Here we provide a summary of the basic mechanistic role of APE1 in DNA repair and redox regulation and highlight preclinical and clinical data that confirm APE1 as a valid anticancer drug target. Development of small molecule inhibitors of APE1 is an area of intense research and current evidence using APE1 inhibitors has demonstrated potentiation of cytotoxicity of alkylating agents in preclinical models implying translational applications in cancer patients.

Study of Etanercept, a Tumor Necrosis Factor-Alpha Inhibitor, in Recurrent Ovarian Cancer

PURPOSE Convincing data support the link between inflammation and ovarian cancer. Tumor necrosis factor-alpha (TNF-alpha), a major mediator of inflammation, is chronically produced in the ovarian tumor microenvironment and may enhance tumor growth and invasion by inducing the secretion of cytokines, proangiogenic factors, and metalloproteinases. Etanercept is a recombinant human soluble p75 TNF receptor that binds to TNF-alpha and renders it biologically unavailable. In the current study, we sought to determine the toxicity, biologic activity, and therapeutic efficacy of etanercept in recurrent ovarian cancer. PATIENTS AND METHODS We initiated a phase I-B, nonrandomized, open-label study in patients with recurrent ovarian cancer. Etanercept was administered subcutaneously at a dose of 25 mg twice weekly (cohort one) and 25 mg thrice weekly (cohort two) until disease progression. RESULTS Thirty patients were recruited (cohort one, 17 patients; cohort two, 13 patients). Eighteen of the 30 patients (cohort one, 11 patients; cohort two, seven patients) completed > or = 12 weeks of treatment. Six patients achieved prolonged disease stabilization (cohort one, two patients [40 and 25 weeks]; cohort two, four patients [34, 24, 22, and 24 weeks]). A significant rise in immunoreactive TNF was seen in all patients (pretreatment compared with end of treatment). A phytohemagglutinin-stimulated whole-blood cytokine assay showed a significant fall in interleukin-6 (cohort one [11 of 17]) and CCL2 (cohort one [13 of 17]) levels. Common adverse effects were injection-site reactions and fatigue. CONCLUSION We provide evidence for the biologic activity and safety of etanercept in recurrent ovarian cancer. Our data suggest possible clinical activity that must be confirmed in future studies.

Gemcitabine and capecitabine with or without telomerase peptide vaccine GV1001 in patients with locally advanced or metastatic pancreatic cancer (TeloVac): an open-label, randomised, phase 3 trial

BACKGROUND We aimed to assess the efficacy and safety of sequential or simultaneous telomerase vaccination (GV1001) in combination with chemotherapy in patients with locally advanced or metastatic pancreatic cancer. METHODS TeloVac was a three-group, open-label, randomised phase 3 trial. We recruited patients from 51 UK hospitals. Eligible patients were treatment naive, aged older than 18 years, with locally advanced or metastatic pancreatic ductal adenocarcinoma, and Eastern Cooperative Oncology Group performance status of 0-2. Patients were randomly assigned (1:1:1) to receive either chemotherapy alone, chemotherapy with sequential GV1001 (sequential chemoimmunotherapy), or chemotherapy with concurrent GV1001 (concurrent chemoimmunotherapy). Treatments were allocated with equal probability by means of computer-generated random permuted blocks of sizes 3 and 6 in equal proportion. Chemotherapy included six cycles of gemcitabine (1000 mg/m(2), 30 min intravenous infusion, at days 1, 8, and 15) and capecitabine (830 mg/m(2) orally twice daily for 21 days, repeated every 28 days). Sequential chemoimmunotherapy included two cycles of combination chemotherapy, then an intradermal lower abdominal injection of granulocyte-macrophage colony-stimulating factor (GM-CSF; 75 μg) and GV1001 (0·56 mg; days 1, 3, and 5, once on weeks 2-4, and six monthly thereafter). Concurrent chemoimmunotherapy included giving GV1001 from the start of chemotherapy with GM-CSF as an adjuvant. The primary endpoint was overall survival; analysis was by intention to treat. This study is registered as an International Standard Randomised Controlled Trial, number ISRCTN4382138. FINDINGS The first patient was randomly assigned to treatment on March 29, 2007, and the trial was terminated on March 27, 2011. Of 1572 patients screened, 1062 were randomly assigned to treatment (358 patients were allocated to the chemotherapy group, 350 to the sequential chemoimmunotherapy group, and 354 to the concurrent chemoimmunotherapy group). We recorded 772 deaths; the 290 patients still alive were followed up for a median of 6·0 months (IQR 2·4-12·2). Median overall survival was not significantly different in the chemotherapy group than in the sequential chemoimmunotherapy group (7·9 months [95% CI 7·1-8·8] vs 6·9 months [6·4-7·6]; hazard ratio [HR] 1·19, 98·25% CI 0·97-1·48, p=0·05), or in the concurrent chemoimmunotherapy group (8·4 months [95% CI 7·3-9·7], HR 1·05, 98·25% CI 0·85-1·29, p=0·64; overall log-rank of χ(2)2df=4·3; p=0·11). The commonest grade 3-4 toxic effects were neutropenia (68 [19%] patients in the chemotherapy group, 58 [17%] patients in the sequential chemoimmunotherapy group, and 79 [22%] patients in the concurrent chemoimmunotherapy group; fatigue (27 [8%] in the chemotherapy group, 35 [10%] in the sequential chemoimmunotherapy group, and 44 [12%] in the concurrent chemoimmunotherapy group); and pain (34 [9%] patients in the chemotherapy group, 39 [11%] in the sequential chemoimmunotherapy group, and 41 [12%] in the concurrent chemoimmunotherapy group). INTERPRETATION Adding GV1001 vaccination to chemotherapy did not improve overall survival. New strategies to enhance the immune response effect of telomerase vaccination during chemotherapy are required for clinical efficacy. FUNDING Cancer Research UK and KAEL-GemVax.

Phase I study of MetXia-P450 gene therapy and oral cyclophosphamide for patients with advanced breast cancer or melanoma.

Purpose: MetXia-P450 is a novel recombinant retroviral vector that encodes the human cytochrome P450 type 2B6 gene (CYP2B6), Escherichia coli lacZ, and neomycin resistance marker genes. Cytochrome P450 enzymes are primarily expressed in the liver and convert the prodrug cyclophosphamide to an active phosphoramide mustard and acrolein. Gene-based delivery of CYP2B6 to the tumor site leads to local prodrug activation and higher concentrations of the active metabolites at the target site. Experimental Design: MetXia-P450 was directly injected into metastatic cutaneous tumor nodules on days 1 and 2 and nodules biopsied on day 7. Oral cyclophosphamide (100 mg/m2) was administered between days 8 and 22. Subsequent cycles of oral cyclophosphamide were repeated for 2 of 4 weeks. Gene transfer levels in biopsy samples were measured by histologic and quantitative PCR analyses. Safety assessments were made using PCR for vector dissemination to the blood after injection and using PCR and serologic analyses to detect replicating virus. Secondary end points included clinical response, toxicity, and evaluation of antitumor immune responses by measurement of carcinoembryonic antigen and 5T4 antibodies. Results: Twelve patients with breast cancer (n = 9) and melanoma (n = 3) received three dose levels of MetXia-P450 (∼8 × 105, ∼8 × 106, and ∼8 × 107 lacZ transferring units/mL). The product was safe and well tolerated. The lacZ transgene was detected in biopsy material by immunohistochemistry in 10 of 12 patients and integrated viral sequences by PCR in 3 of 6 patients. One (8%) patient with breast cancer had a partial response and received 7 months of oral cyclophosphamide. Four (33%) patients had stable disease for ≥3 months and the rest had progressive disease. Preliminary immunologic analyses were suggestive of an antitumor response in two patients (partial response in one patient and stable disease in one patient). Conclusion: MetXia was safe and well tolerated. Gene transfer was detected at all dose levels, and the initial suggestion of an antitumor response indicates that MetXia-P450 should undergo further clinical assessment.

Acquired Resistance to Temozolomide in Glioma Cell Lines: Molecular Mechanisms and Potential Translational Applications

Treatment for glioblastoma multiforme includes the alkylating agent temozolomide combined with ionizing radiation. Persistent O6-guanine methylation by temozolomide in O6-methylguanine methyl transferase negative tumors causes cytotoxic lesions recognized by DNA mismatch repair, triggering apoptosis. Resistance (intrinsic or acquired) presents obstacles to successful temozolomide treatment, limiting drug efficacy and life expectancy. Two glioma cell lines, SNB19 and U373, sensitive to temozolomide (GI50 values 36 and 68 µM, respectively) were exposed to increasing temozolomide concentrations (1–100 µM). Variant cell lines (SNB19VR, U373VR) were generated that displayed acquired temozolomide resistance (GI50 values 280 and 289 µM, respectively). Cross-resistance to mitozolomide was observed in U373VR cells only. In clonogenic and MTT assays, methylguanine methyltransferase (MGMT) depletion using O6-benzylguanine sensitized U373VR cells to temozolomide, indicating the resistance mechanism involves MGMT re-expression. Indeed, Western blot analyses revealed MGMT protein in cell lysates. In SNB19VR cells, down-regulation of MSH6 message and protein expression may confer temozolomide tolerance. Inhibition of poly(ADP-ribose) polymerase-1 (a key base excision repair (BER) enzyme) partially restored sensitivity, and DNA repair gene arrays demonstrated up-regulation (>5-fold) of BER gene NTL1 in SNB19VR cells. In conclusion, we have developed two glioma cell lines whose distinct mechanisms of acquired resistance to temozolomide, involving expression of MGMT, or inactivation of DNA mismatch repair and recruitment of BER enzymes, are consistent with clinical observations. These cell lines provide valuable models for the development of strategies to combat temozolomide resistance.

Biomarkers of response to therapy in oesophago-gastric cancer

Cancer of the oesophagus, gastro-oesophageal junction (GOJ) and stomach remains a major health problem worldwide. The evidence base for the optimal management of patients with operable oesophago-gastric cancer is evolving. Accepted approaches include preoperative chemotherapy followed by surgery (oesophageal cancer), chemo-radiotherapy alone (oesophageal cancer) and perioperative chemotherapy (gastric and gastro-oesophageal adenocarcinomas). The underlying principles behind neoadjuvant therapy are to improve resectability of the tumour by tumour shrinkage/downstaging and to treat occult metastatic disease as early as possible. The response rate to cytotoxic therapy is about 40% in oesophago-gastric cancer. Available evidence suggests that a favourable histopathological response to cytotoxic therapy may be a useful positive predictive marker in oesophago-gastric cancer. However, the ability to predict tumour response in routine clinical practice is difficult and is an area of intense investigation. There is evolving evidence for the role of predictive biomarkers in cancer in general and oesophago-gastric cancer in particular. We provide an overview on the current status of radiological and biological predictive biomarkers. We have focussed on clinical translational investigations and, where appropriate, provided pre-clinical insights. Whether predictive markers will be routinely incorporated in clinical practice remains to be seen as biomarker research is expensive and the data generated from these investigations are complex. It is clear that a concerted international effort between academia and industry is critical if personalised medicine as a practical reality for our cancer patients is to be realised.

Synthetic lethal targeting of DNA double-strand break repair deficient cells by human apurinic/apyrimidinic endonuclease inhibitors

An apurinic/apyrimidinic (AP) site is an obligatory cytotoxic intermediate in DNA Base Excision Repair (BER) that is processed by human AP endonuclease 1 (APE1). APE1 is essential for BER and an emerging drug target in cancer. We have isolated novel small molecule inhibitors of APE1. In this study, we have investigated the ability of APE1 inhibitors to induce synthetic lethality (SL) in a panel of DNA double‐strand break (DSB) repair deficient and proficient cells; i) Chinese hamster (CH) cells: BRCA2 deficient (V‐C8), ATM deficient (V‐E5), wild type (V79) and BRCA2 revertant [V‐C8(Rev1)]. ii) Human cancer cells: BRCA1 deficient (MDA‐MB‐436), BRCA1 proficient (MCF‐7), BRCA2 deficient (CAPAN‐1 and HeLa SilenciX cells), BRCA2 proficient (PANC1 and control SilenciX cells). We also tested SL in CH ovary cells expressing a dominant‐negative form of APE1 (E8 cells) using ATM inhibitors and DNA‐PKcs inhibitors (DSB inhibitors). APE1 inhibitors are synthetically lethal in BRCA and ATM deficient cells. APE1 inhibition resulted in accumulation of DNA DSBs and G2/M cell cycle arrest. SL was also demonstrated in CH cells expressing a dominant‐negative form of APE1 treated with ATM or DNA‐PKcs inhibitors. We conclude that APE1 is a promising SL target in cancer.