Maccon Keane

Prospective validation of a 21-gene expression assay in breast cancer

BACKGROUND Prior studies with the use of a prospective-retrospective design including archival tumor samples have shown that gene-expression assays provide clinically useful prognostic information. However, a prospectively conducted study in a uniformly treated population provides the highest level of evidence supporting the clinical validity and usefulness of a biomarker. METHODS We performed a prospective trial involving women with hormone-receptor-positive, human epidermal growth factor receptor type 2 (HER2)-negative, axillary node-negative breast cancer with tumors of 1.1 to 5.0 cm in the greatest dimension (or 0.6 to 1.0 cm in the greatest dimension and intermediate or high tumor grade) who met established guidelines for the consideration of adjuvant chemotherapy on the basis of clinicopathologic features. A reverse-transcriptase-polymerase-chain-reaction assay of 21 genes was performed on the paraffin-embedded tumor tissue, and the results were used to calculate a score indicating the risk of breast-cancer recurrence; patients were assigned to receive endocrine therapy without chemotherapy if they had a recurrence score of 0 to 10, indicating a very low risk of recurrence (on a scale of 0 to 100, with higher scores indicating a greater risk of recurrence). RESULTS Of the 10,253 eligible women enrolled, 1626 women (15.9%) who had a recurrence score of 0 to 10 were assigned to receive endocrine therapy alone without chemotherapy. At 5 years, in this patient population, the rate of invasive disease-free survival was 93.8% (95% confidence interval [CI], 92.4 to 94.9), the rate of freedom from recurrence of breast cancer at a distant site was 99.3% (95% CI, 98.7 to 99.6), the rate of freedom from recurrence of breast cancer at a distant or local-regional site was 98.7% (95% CI, 97.9 to 99.2), and the rate of overall survival was 98.0% (95% CI, 97.1 to 98.6). CONCLUSIONS Among patients with hormone-receptor-positive, HER2-negative, axillary node-negative breast cancer who met established guidelines for the recommendation of adjuvant chemotherapy on the basis of clinicopathologic features, those with tumors that had a favorable gene-expression profile had very low rates of recurrence at 5 years with endocrine therapy alone. (Funded by the National Cancer Institute and others; ClinicalTrials.gov number, NCT00310180.).

Breast-cancer adjuvant therapy with zoledronic acid

BACKGROUND Data suggest that the adjuvant use of bisphosphonates reduces rates of recurrence and death in patients with early-stage breast cancer. We conducted a study to determine whether treatment with zoledronic acid, in addition to standard adjuvant therapy, would improve disease outcomes in such patients. METHODS In this open-label phase 3 study, we randomly assigned 3360 patients to receive standard adjuvant systemic therapy either with or without zoledronic acid. The zoledronic acid was administered every 3 to 4 weeks for 6 doses and then every 3 to 6 months to complete 5 years of treatment. The primary end point of the study was disease-free survival. A second interim analysis revealed that a prespecified boundary for lack of benefit had been crossed. RESULTS At a median follow-up of 59 months, there was no significant between-group difference in the primary end point, with a rate of disease-free survival of 77% in each group (adjusted hazard ratio in the zoledronic acid group, 0.98; 95% confidence interval [CI], 0.85 to 1.13; P=0.79). Disease recurrence or death occurred in 377 patients in the zoledronic acid group and 375 of those in the control group. The numbers of deaths--243 in the zoledronic acid group and 276 in the control group--were also similar, resulting in rates of overall survival of 85.4% in the zoledronic acid group and 83.1% in the control group (adjusted hazard ratio, 0.85; 95% CI, 0.72 to 1.01; P=0.07). In the zoledronic acid group, there were 17 confirmed cases of osteonecrosis of the jaw (cumulative incidence, 1.1%; 95% CI, 0.6 to 1.7; P<0.001) and 9 suspected cases; there were no cases in the control group. Rates of other adverse effects were similar in the two study groups. CONCLUSIONS These findings do not support the routine use of zoledronic acid in the adjuvant management of breast cancer. (Funded by Novartis Pharmaceuticals and the National Cancer Research Network; AZURE Current Controlled Trials number, ISRCTN79831382.).

TRAIL receptor signalling and modulation : Are we on the right TRAIL?

Tumour necrosis factor-related apoptosis-inducing ligand or Apo2 ligand (TRAIL/Apo2L) is a member of the tumour necrosis factor (TNF) superfamily of cytokines that induces apoptosis upon binding to its death domain-containing transmembrane receptors, death receptors 4 and 5 (DR4, DR5). Importantly, TRAIL preferentially induces apoptosis in cancer cells while exhibiting little or no toxicity in normal cells. To date, research has focused on the mechanism of apoptosis induced by TRAIL and the processes involved in the development of TRAIL resistance. TRAIL-resistant tumours can be re-sensitized to TRAIL by a combination of TRAIL with chemotherapeutics or irradiation. Studies suggest that in many cancer cells only one of the two death-inducing TRAIL receptors is functional. These findings as well as the aim to avoid decoy receptor-mediated neutralization of TRAIL led to the development of receptor-specific TRAIL variants and agonistic antibodies. These molecules are predicted to be more potent than native TRAIL in vivo and may be suitable for targeted treatment of particular tumours. This review focuses on the current status of TRAIL receptor-targeting for cancer therapy, the apoptotic signalling pathway induced by TRAIL receptors, the prognostic implications of TRAIL receptor expression and modulation of TRAIL sensitivity of tumour cells by combination therapies. The mechanisms of TRAIL resistance and the potential measures that can be taken to overcome them are also addressed. Finally, the status of clinical trials of recombinant TRAIL and DR4-/DR5-specific agonistic antibodies as well as the pre-clinical studies of receptor-selective TRAIL variants is discussed including the obstacles facing the use of these molecules as anti-cancer therapeutics.

The effects of adding zoledronic acid to neoadjuvant chemotherapy on tumour response: exploratory evidence for direct anti-tumour activity in breast cancer

Background:Pre-clinical studies have demonstrated synergistic anti-tumour effects of chemotherapy (CT) and zoledronic acid (ZOL). Within the AZURE trial, designed to determine whether the addition of ZOL to neoadjuvant therapy improves disease outcomes, a subgroup received neoadjuvant CT. We report a retrospective evaluation comparing pathological response in the primary tumour between treatment groups.Methods:In total, 205 patients received neoadjuvant CT±ZOL (CT+ZOL, n=102; CT, n=103). The primary end point was pathologically assessed residual invasive tumour size (RITS) at surgery. Secondary end points were pathological complete response (pCR) rate and axillary nodal involvement. Following review of surgical pathology reports (n=195), outcome differences between groups were assessed adjusting for potential response modifiers.Results:Baseline characteristics and CT treatments were similar. In multivariate analysis, allowing for biological and clinical factors known to influence tumour response, the adjusted mean RITS in CT and CT+ZOL groups were 27.4 and 15.5 mm, respectively, giving a difference in means of 12 mm (95% confidence interval: 3.5–20.4 mm; P=0.006). The pCR rate was 6.9% in the CT group and 11.7% in the CT+ZOL group (P=0.146). There was no difference in axillary nodal involvement (P=0.6315).Conclusion:These data suggest a possible direct anti-tumour effect of ZOL in combination with CT, warranting formal evaluation in prospective studies.

Adjuvant zoledronic acid in patients with early breast cancer: final efficacy analysis of the AZURE (BIG 01/04) randomised open-label phase 3 trial

BACKGROUND The role of adjuvant bisphosphonates in early breast cancer is uncertain. We therefore did a large randomised trial to investigate the effect of the adjuvant use of zoledronic acid on disease-free survival (DFS) in high-risk patients with early breast cancer. METHODS In the AZURE trial, an open-label, international, multicentre, randomised, controlled, parallel-group phase 3 trial, women (age ≥18 years) with stage II or III breast cancer were randomly assigned (1:1) by a central automated 24-h computer-generated telephone minimisation system (balanced for number of involved axillary lymph nodes, tumour stage, oestrogen receptor status, type and timing of systemic therapy, menopausal status, statin use, and treatment centre) to receive standard adjuvant systemic treatment alone (control group) or with 4 mg intravenous zoledronic acid every 3-4 weeks for six doses, then every 3 months for eight doses, followed by every 6 months for five doses, for a total of 5 years of treatment. The primary endpoint was disease-free survival (DFS). Secondary endpoints were invasive DFS (IDFS), overall survival, time to bone metastases, time to distant recurrence, and subgroup analyses of variables included in the randomisation. All patients have completed study treatment. Results from the intention-to-treat final analysis of this fully recruited study are presented after a median follow-up of 84 months (IQR 66-93). This final efficacy analysis was planned to take place after 940 DFS events. This trial is registered with ClinicalTrials.gov, NCT00072020. FINDINGS 3360 women were recruited from 174 centres in seven countries between Sept 4, 2003, and Feb 16, 2006. The number of DFS events did not differ between groups: 493 in the control group and 473 in the zoledronic acid group (adjusted hazard ratio [HR] 0·94, 95% CI 0·82-1·06; p=0·30). IDFS (HR 0·93, 95% CI 0·82-1·05; p=0·22), overall survival (0·93, 0·81-1·08; p=0·37), and distant recurrences (0·93, 0·81-1·07; p=0·29) were much the same in both groups. Zoledronic acid reduced the development of bone metastases, both as a first event (HR 0·78, 95% CI 0·63-0·96; p=0·020) and at any time during follow-up (0·81, 0·68-0·97; p=0·022). The effects of zoledronic acid on DFS were not affected by oestrogen-receptor status. However, zoledronic acid improved IDFS in those who were over 5 years since menopause at trial entry (n=1041; HR 0·77, 95% CI 0·63-0·96) but not in all other (premenopause, perimenopause, and unknown status) menopausal groups (n=2318; HR 1·03, 95% CI 0·89-1·20). 33 cases of suspected osteonecrosis of the jaw have been reported, with 26 confirmed on central review, all in the zoledronic acid group (1·7%, 95% CI 1·0-2·4). INTERPRETATION These results suggest no overall benefit from the addition of zoledronic acid to standard adjuvant treatments for early breast cancer. However, zoledronic acid does reduce the development of bone metastases and, for women with established menopause, improved disease outcomes. FUNDING Novartis Global and NIHR Cancer Research Network.

2-weekly versus 3-weekly docetaxel to treat castration-resistant advanced prostate cancer: a randomised, phase 3 trial

BACKGROUND Docetaxel administered every 3 weeks is a standard treatment for castration-resistant advanced prostate cancer. We hypothesised that 2-weekly administration of docetaxel would be better tolerated than 3-weekly docetaxel in patients with castration-resistant advanced prostate cancer, and did a prospective, multicentre, randomised, phase 3 study to compare efficacy and safety. METHODS Eligible patients had advanced prostate cancer (metastasis, a prostate-specific-antigen test result of more than 10·0 ng/mL, and WHO performance status score of 0-2), had received no chemotherapy (except with estramustine), had undergone surgical or chemical castration, and had been referred to a treatment centre in Finland, Ireland, or Sweden. Enrolment and treatment were done between March 1, 2004, and May 31, 2009. Randomisation was done centrally and stratified by centre and WHO performance status score of 0-1 vs 2. Patients were assigned 75 mg/m(2) docetaxel intravenously on day 1 of a 3-week cycle, or 50 mg/m(2) docetaxel intravenously on days 1 and 15 of a 4-week cycle. 10 mg oral prednisolone was administered daily to all patients. The primary endpoint was time to treatment failure (TTTF). We assessed data in the per-protocol population. This study is registered with ClinicalTrials.gov, number NCT00255606. FINDINGS 177 patients were randomly assigned to the 2-weekly docetaxel group and 184 to the 3-weekly group. 170 patients in the 2-weekly group and 176 in the 3-weekly group were included in the analysis. The 2-weekly administration was associated with significantly longer TTTF than was 3-weekly administration (5·6 months, 95% CI 5·0-6·2 vs 4·9 months, 4·5-5·4; hazard ratio 1·3, 95% CI 1·1-1·6, p=0·014). Grade 3-4 adverse events occurred more frequently in the 3-weekly than in the 2-weekly administration group, including neutropenia (93 [53%] vs 61 [36%]), leucopenia (51 [29%] vs 22 [13%]), and febrile neutropenia (25 [14%] vs six [4%]). Neutropenic infections were reported more frequently in patients who received docetaxel every 3 weeks (43 [24%] vs 11 [6%], p=0·002). INTERPRETATION Administration of docetaxel every 2 weeks seems to be well tolerated in patients with castration-resistant advanced prostate cancer and could be a useful option when 3-weekly single-dose administration is unlikely to be tolerated. FUNDING Sanofi.

Abstract S4-5: Adjuvant Treatment with Zoledronic Acid in Stage II/III Breast Cancer. The AZURE Trial (BIG 01/04)

Background: The ABCSG XII trial demonstrated a 32% risk reduction in disease-free survival (DFS) events with Zoledronic acid (ZOL) use in a cohort of premenopausal women treated with endocrine therapy at 62 months median follow-up .[1,2] This strategy is increasingly being adopted in the wider breast cancer population. The AZURE trial is an academic study designed to determine whether treatment with ZOL added to standard adjuvant therapy improves DFS and bone metastasis-free survival (BMFS) in a broader range of patients with stage II/III breast cancer. Materials and methods: 3360 patients from 174 centres were randomized to receive (neo) adjuvant chemotherapy (CT) and/or endocrine therapy (ET) +/− ZOL 4mg iv every 3-4 weeks for 6 doses, then 3 monthly x 8 and 6 monthly x 5 to complete 5 years treatment. The primary DFS endpoint was to be determined after 940 DFS events, providing 80% power to detect a 17% reduction in hazard rates (HR) for DFS events. The rate of events on study has been slower than expected, resulting in an estimated final analysis in 2012. In light of the clinical interest in the results of AZURE, the DMEC has agreed to a second interim analysis after 705 events with boundaries set by an independent statistician, unaware of results at the first interim analysis, for both efficacy (HR ∼0.82, alpha spend 1%) and lack of clinical benefit (HR ∼0.935 with a 5% risk of declaring a false negative result). Contingent on DMEC advice, efficacy results that breach either boundary will be presented. A DMEC recommendation not to release the results at the second interim analysis may indicate that the point HR estimate lies between these boundaries. Results: Patient characteristics including stage, number of positive axillary nodes, CT type, ER status, menopausal status and statin use were well balanced. 3208 patients (96%) received (neo) adjuvant CT (93% anthracyclines, 23% taxanes). 152 patients received ET alone. As of June 9th 2010, with a median follow up of 49 (IQR 42-56) months, there have been 695 DFS events. The safety population comprised 3340 patients (ZOL 1665; control 1675). The addition of ZOL to standard treatment did not significantly impact on chemotherapy delivery. Serious adverse events (SAE) were similar in both treatment arms. To date 13 confirmed (0.83%; 95% CI 0.38%, 1.28%) and 12 possible (1.12%; 95% CI 0.39%, 1.85%) cases of osteonecrosis of the jaw (ONJ) in the ZOL arm have occurred. Discussion: AZURE is one of the largest phase III studies of adjuvant bisphosphonates, and the results from this study will help define the role of adjuvant ZOL in the management of early breast cancer. [1] Gnant M et al. NEJM 2009; 360(7):679-691 [2] Gnant M et al. ASCO 2010 Proceedings; abs #533. Citation Information: Cancer Res 2010;70(24 Suppl):Abstract nr S4-5.

Adjuvant Chemotherapy Guided by a 21-Gene Expression Assay in Breast Cancer

BACKGROUND The recurrence score based on the 21‐gene breast cancer assay predicts chemotherapy benefit if it is high and a low risk of recurrence in the absence of chemotherapy if it is low; however, there is uncertainty about the benefit of chemotherapy for most patients, who have a midrange score. METHODS We performed a prospective trial involving 10,273 women with hormone‐receptor–positive, human epidermal growth factor receptor 2 (HER2)–negative, axillary node–negative breast cancer. Of the 9719 eligible patients with follow‐up information, 6711 (69%) had a midrange recurrence score of 11 to 25 and were randomly assigned to receive either chemoendocrine therapy or endocrine therapy alone. The trial was designed to show noninferiority of endocrine therapy alone for invasive disease–free survival (defined as freedom from invasive disease recurrence, second primary cancer, or death). RESULTS Endocrine therapy was noninferior to chemoendocrine therapy in the analysis of invasive disease–free survival (hazard ratio for invasive disease recurrence, second primary cancer, or death [endocrine vs. chemoendocrine therapy], 1.08; 95% confidence interval, 0.94 to 1.24; P=0.26). At 9 years, the two treatment groups had similar rates of invasive disease–free survival (83.3% in the endocrine‐therapy group and 84.3% in the chemoendocrine‐therapy group), freedom from disease recurrence at a distant site (94.5% and 95.0%) or at a distant or local–regional site (92.2% and 92.9%), and overall survival (93.9% and 93.8%). The chemotherapy benefit for invasive disease–free survival varied with the combination of recurrence score and age (P=0.004), with some benefit of chemotherapy found in women 50 years of age or younger with a recurrence score of 16 to 25. CONCLUSIONS Adjuvant endocrine therapy and chemoendocrine therapy had similar efficacy in women with hormone‐receptor–positive, HER2‐negative, axillary node–negative breast cancer who had a midrange 21‐gene recurrence score, although some benefit of chemotherapy was found in some women 50 years of age or younger. (Funded by the National Cancer Institute and others; TAILORx ClinicalTrials.gov number, NCT00310180.)

Early growth response-1 is a regulator of DR5-induced apoptosis in colon cancer cells

Background:Tumour necrosis factor-related apoptosis-inducing ligand (TRAIL) induces tumour cell apoptosis by binding to death receptor 4 (DR4) and DR5. DR4 and DR5 activation however can also induce inflammatory and pro-survival signalling. It is not known how these different cellular responses are regulated and what the individual role of DR4 vs DR5 is in these processes.Methods:DNA microarray study was carried out to identify genes differentially expressed after DR4 and DR5 activation. RT–PCR and western blotting was used to examine the expression of early growth response gene-1 (Egr-1) and the proteins of the TRAIL signalling pathway. The function of Egr-1 was studied by siRNA-mediated knockdown and overexpression of a dominant-negative version of Egr-1.Results:We show that the immediate early gene, Egr-1, regulates TRAIL sensitivity. Egr-1 is constitutively expressed in colon cancer cells and further induced upon activation of DR4 or DR5. Our results also show that DR4 mediates a type II, mitochondrion-dependent apoptotic pathway, whereas DR5 induces a mitochondrion-independent, type I apoptosis in HCT15 colon carcinoma cells. Egr-1 drives c-FLIP expression and the short splice variant of c-FLIP (c-FLIPS) specifically inhibits DR5 activation.Conclusion:Selective knockdown of c-FLIPS sensitises cells to DR5-induced but not DR4-induced apoptosis and Egr-1 exerts an effect as an inhibitor of the DR5-induced apoptotic pathway, possibly by regulating the expression of c-FLIPS.

Differential activation of JNK1 isoforms by TRAIL receptors modulate apoptosis of colon cancer cell lines

Tumour necrosis factor-related apoptosis-inducing ligand (TRAIL) induces apoptosis on binding to its receptors, death receptor 4 and 5 (DR4, DR5). TRAIL can also activate c-Jun N-terminal kinase (JNK) through the adaptor molecules, TNF receptor-associated factor 2 (TRAF2) and receptor-interacting protein (RIP). The role of JNK in TRAIL-induced tumour cell apoptosis is unclear. In this study, we demonstrate that JNK is activated by TRAIL in colon cancer cells. Inhibition of JNK with L-JNKI reduced rhTRAIL-induced cell death but enhanced cell death induced by selective activation of DR4 or DR5. This difference was unrelated to receptor internalisation or differential activation of c-Jun, but activation of different JNK isoforms. Our data demonstrate that JNK1, but not JNK2 is activated by rhTRAIL in the examined colon cancer cell lines. Although rhTRAIL activated both the long and short isoforms of JNK1, selective activation of DR4 or DR5 led to predominant activation of the short JNK1 isoforms (JNK1α1 and/or JNK1β1). Knockdown of JNK1α1 by shRNA enhanced apoptosis induced by TRAIL, agonistic DR4 or DR5 antibodies. On the other hand, knockdown of the long JNK1 isoforms (JNK1α2 and JNK1β2) had the opposite effect; it reduced TRAIL-induced cell death. These data indicate that the short JNK1 isoforms transmit an antiapoptotic signal, whereas the long isoforms (JNK1α2 or JNK1β2) act in a proapoptotic manner.