Citations

Abstract Background: In 2007, the American Society of Clinical Oncology and College of American Pathologists (ASCO/CAP) recommended new guidelines to define HER2 positivity by immunohistochemistry (3+ IHC: uniform intense membrane staining of >30% of invasive tumor cells) or fluorescent in situ hybridization (FISH+: HER2/CEP17 ratio >2.2). The original criteria used for enrollment in the pivotal N9831 trial conducted from 2000-2005 were IHC>10% (3+) or FISH ≥2.0.Purpose: To investigate the impact of the ASCO/CAP guidelines on patient (pt) eligibility and disease free survival (DFS) compared to the originally used FDA-approved definitions.Patients/Methods: This analysis included 2268 patients from the HER2+ N9831 adjuvant trastuzumab phase III trial. IHC was centrally performed at the Mayo Clinic using the DAKO HercepTest™ and was re-analyzed to determine the percent of tumor cells with 0, 1+, 2+, and 3+ staining intensities. DFS was compared between pts randomized to standard chemotherapy +/- concurrent trastuzumab (Arms A and C of the trial) within IHC/FISH subgroups using Cox proportional hazards regression stratified by hormone receptor and nodal status.Results: Based on 2268 pts, 83 pts (3.7%) were 3+ by FDA-approved guidelines but not 3+ by the ASCO/CAP guidelines. Among these pts, 77% (64/83) were FISH amplified centrally by ASCO/CAP guidelines. Only 1.5% (34/2268) of pts eligible under FDA-approved definitions from N9831 in this sample set would not meet the ASCO/CAP guidelines. In 1441 patients randomized to standard chemotherapy +/- concurrent trastuzumab, the exploratory DFS hazard ratio (HR) associated with trastuzumab in IHC/FISH subgroups were as follows: ASCO/CAP HER2+ HR=0.59 (95% CI: 0.47-0.75; p<0.001); FDA-approved HER2+ HR=0.60 (95% CI: 0.47-0.75; p<0.001); ASCO/CAP HER2 normal(not positive) HR=0.74 (95% CI: 0.35-1.57; p=0.43); and FDA-approved HER2 normal HR=0.53 (95% CI: 0.22-1.26; p=0.15).Conclusions: A small percentage (1.5%) of N9831 patients did not meet ASCO/CAP 2007 guidelines when applied retrospectively. Trastuzumab effect appeared similar for HER2+ patients regardless of ASCO/CAP or FDA-approved guidelines. Additional DFS analyses by IHC/FISH subgroups will be presented. We gratefully acknowledge support from the Breast Cancer Research Foundation, Genentech, and NIH CA25224 for this study. Citation Information: Cancer Res 2009;69(24 Suppl):Abstract nr 701.

Abstract Background: Gene expression profiling is an important tool in prediction, prognosis, and cancer modeling. The DASL® assay (cDNA –mediated Annealing, extension, Selection, and Ligation; Illumina; San Diego, CA) is a gene expression profiling system designed for degraded RNAs such as those derived from formalin-fixed paraffin-embedded (FFPE) tumor samples. FFPE tumor samples are invaluable resources for cancer research, being the most widely available materials for which patient outcomes are known. Purpose: The objective of this study was to evaluate the performance of Illumina's Whole Genome DASL platform (24,526; 24K) at distinguishing HER2-positive (HER2+) from HER2-negative (HER2-) FFPE breast tumors. Methods: We measured the relative gene expression of biological and technical replicates using nine HER2+ and 11 HER2- primary breast tumors from patients that were all node-positive and estrogen receptor-positive. The HER2 status of the tumors was determined using the Dako Hercept Test following the 2007 ASCO/CAP guidelines for HER2 positivity. The non-background corrected probe-expression data were exported from Illumina BeadStudio software and normalized using the software R and the function fastlo. Log2 transformed expression values were used to obtain the relative gene expression per probe. Spearman correlations were calculated between both types of replicates. Unequal-variance t-tests were conducted to test for differences in expression levels between HER2+ and HER2- tumors. An overall ERBB2 gene expression level was calculated by taking the average of the three ERBB2 probes for each tumor across all replicates. To simulate the possible outcome if only one replicate of each tumor was measured for gene expression, differential expression was tested for all possible combinations of single replicate probe expression values. Results: Spearman correlations for technical and biological replicates within tumor samples ranged from 0.9804 – 0.9946, and 0.9822 – 0.9945, respectively. The overall ERBB2 gene expression was significantly lower in HER2- tumors than in HER2+ tumors (p = 0.0000018). The top five most differentially expressed genes (p < 0.0005) included ERBB2, GRB7, C17ORF37, PERLD1, and ASF1B. For each of the three ERBB2 probes, there were 65,536 possible ways to combine the individual probe expression values. The p-values for testing differential expression in each of these 65,536 combinations of HER2+ tumors versus HER2- tumors ranged from 0.00029 – 0.0091 for ILMN_1717902, 0.0000078 – 0.00026 for ILMN_1728761, and 0.000000002 – 0.0000032 for ILMN-2352131. Conclusions: Illumina's Whole Genome Platform provided highly reproducible gene expression values among biological and technical replicates of breast tumors. This platform also demonstrated significant differential ERBB2 gene expression between HER2+ and HER- breast tumors, which correctly classified the breast tumors as HER2+ and HER2-. Citation Information: Cancer Res 2009;69(24 Suppl):Abstract nr 6127.

Abstract BackgroundRIBBON-1, a Phase III, multicenter, randomized, placebo-controlled trial in patients with previously untreated MBC was designed to evaluate the efficacy and safety of adding bevacizumab (B) to chemotherapy regimens including capecitabine (Cape; n=615); a taxane (T; n=307), or an anthracycline (Anth; n=315) compared with chemotherapy alone. Pre-specified analyses of progression-free survival PFS) in the Cape and pooled T/Anth cohorts, the primary endpoints of the study were increased upon addition of B. In the Cape cohort, median PFS increased from 5.7 to 8.6 mo (HR=0.69, p-value=0.0002) and in the T/Anth cohort, it increased from 8.0 to 9.2 mo (HR=0.64; p-value<0.0001).MethodsRECIST was used to determine objective response rate (ORR). Patients without a post-baseline tumor assessment were considered non-responders. The primary analysis for the ORR and the duration of response was performed using only patients with measurable disease at baseline. ORR was formally compared between the two treatment arms using the Mantel-Haenszel Chi-squared test, using the randomization stratification factors. Fisher's exact test was also performed. Clinical benefit rate (CBR) was defined as the proportion of patients with a complete or partial response or with stable disease at Week 24. Exploratory analyses of CBR and time to response are provided here. Summary statistics of time to response at Week 9 will be presented as will the final analysis of the secondary endpoint of overall survival.ResultsAnalyses of ORR and CBR are provided in the following table. CapeTaxaneAnth PL (n=206)B (n=409)PL (n=104)B (n=203)PL (n=103)B (n=212)Patients with measurable disease, n (%)161 (78.2)325 (61.1)85 (81.7)161 (79.3)92 (89.3)184 (86.8)ORR, %23.635.435.350.340.252.2CR, n (%)1 (0.6)7 (2.2)3 (3.5)4 (2.5)2 (2.2)3 (1.6)PR, n (%)37 (23.0)108 (33.2)27 (31.8)77 (47.8)35 (38.0)93 (50.5)Difference between arms, %11.815.012.0p-value, unstratified0.00940.030.073Duration of ORMedian, mo7.29.28.68.46.08.1HR (95% CI)0.61 (0.39–0.96)0.75 (0.45–1.27)0.53 (0.34–0.83)Log-rank p-value0.03260.2840.0047Clinical benefit rate*, %47.164.365.471.969.980.7p-value, unstratified<0.00010.240.04CR+PR at Week 9, %20.229.438.354.544.451.1B=bevacizumab, cape=capecitabine, CI=confidence interval, HR=hazard ratio, OR=objective response, PL=placebo, T/Anth=taxane/anthracycline.*For randomized patients.ConclusionsIn patients with HER2-negative MBC, the addition of bevacizumab to capecitabine, taxane or anthracycline chemotherapy induces more responses and those responses occur more rapidly. For capecitabine and anthracycline the addition of bevacizumab leads to objective responses that last longer. Citation Information: Cancer Res 2009;69(24 Suppl):Abstract nr 6084.

Abstract Background: Despite the impressive results of the trastuzumab adjuvant therapy trials, 15% of patients (pts) with HER2 overexpressing or amplified breast cancer developed tumor relapse at 4 years. Lapatinib is a reversible tyrosine kinase inhibitor against ErbB1 and ErbB2. The current study was developed to assess the cardiac safety and feasibility of adding lapatinib to paclitaxel and trastuzumab following doxorubicin and cyclophosphamide as part of adjuvant therapy.Methods: A single-arm phase II study of doxorubicin (A, 60 mg/m2 day 1) and cyclophosphamide (C, 600 mg/m2 day 1) [q2w or q3w for 4 cycles]; followed by PTL: paclitaxel (P, 80 mg/m2 weekly), trastuzumab (T, 4 mg/kg loading dose then 2 mg/kg weekly), and lapatinib (L, 1000 mg PO daily which was later amended to 750 mg) [total of 12 weeks of treatment]; followed by T (6 mg/kg day 1, every 3 weeks) and L (1000 mg PO daily) [total of 40 weeks of treatment] was conducted. The primary endpoint was severe symptomatic congestive heart failure (CHF: New York Heart Association Class III or IV with a drop in left ventricular ejection fraction [LVEF] of at least 10 percentage points to below 50% confirmed by cardiologist) with secondary endpoints of cardiac death and grade 3+ cardiac adverse events (AEs).Results: From April 2007 to October 2008, 109 evaluable pts were enrolled and initiated study treatment. Median age was 51 (range 28-72). Median follow-up is 10.2 months. No pts experienced CHF or cardiac death while on active treatment. One pt had CHF and 1 pt died due to myocardial infarction (MI) after they went off study treatment. The pt with CHF refused further treatment after completing 2 cycles of PTL and had CHF 1.8 months post PTL discontinuation with LVEF of 45% and dyspnea on exertion. The pt with MI went off study treatment due to non-cardiac AEs after completing 2 cycles of PTL and died 2.9 months post PTL discontinuation. Mean LVEF at baseline was 63.6% (N=109, SD=5.7); end of AC was 63.3% (N=104, SD=6.4); end of PTL was 59.9% (N=89, SD=8.3); and cycle 12 was 59.7% (N=66, SD=7.1). Grade 3+ non-hematologic AEs with incidence >5% include diarrhea (N=42, 39% & included both 1000mg & later amended 750mg dosing), fatigue (N=15, 14%), nausea (N=7, 6%), vomiting (N=7, 6%), acne (N=6, 6%), and hypokalemia (N=6, 6%). Pre-treatment cardiac marker data (troponin-T, troponin-I, brain natriuretic peptide, creatine kinase MB isoenzyme) will also be presented.Conclusions: Preliminary data suggest that the inclusion of L does not add to the cardiac profile of T. Data from ALTTO and long-term follow data are needed to confirm the cardiac safety of this regimen. Cardiac marker data are being analyzed in the context of changes in LVEF. We gratefully acknowledge support from the Breast Cancer Research Foundation and GlaxoSmthKline for this study. Citation Information: Cancer Res 2009;69(24 Suppl):Abstract nr 3086.

Abstract Abstract #6074 Objective: To analyze blood-serum biomarkers (HER2, EGFR, uPA, and TIMP-1) for prediction of response to treatment in conjunction with Study 96-32-55, a multicenter phase II trial assessing the efficacy and tolerability of two irinotecan schedules in anthracycline- or taxane-refractory MBC patients.
 Methods: MBC patients who experienced disease progression after one to three chemotherapy regimens, including at least one anthracycline- or taxane-based regimen, were randomly assigned to irinotecan in 6-week cycles comprising 100 mg/m2 weekly for 4 weeks, then a 2-week rest or 240 mg/m2 every 3 weeks. During this study the serum samples from each patient were collected at a possibility of three time points; prior to treatment, at the first occurrence of response, and at completion of or withdrawal from treatment. Response prediction was analyzed using predetermined elevated/non-elevated serum biomarker cutoffs. Percentage changes from baseline to first response and disease progression were analyzed.
 Results: In the weekly arm, the objective response (CR+PR) rate was 23% (95% CI, 13% to 37%), and in the every-3-weeks arm, the objective response rate was 14% (95% CI, 6% to 26%). Serum biomarker levels were determined for HER2, EGFR, uPA, and TIMP-1 prior to treatment for 91 patients. Of these 91 patients, 17 had serum measurements at first response, and 38 had serum measurements at their completion of study due to disease progression. Only 7 patients had serum-levels collected at all three event-points. The baseline levels of sHER2, EGFR, uPA, and TIMP-1 were not different among responders and non-responders (Fisher's Exact p=0.41, 0.26, 0.68, 0.75). sHER2 level increased by 20.6% from baseline to disease progression (p=0.01). TIMP-1 level was 15.2% lower than baseline at first response (p=0.03), 16.5% higher than baseline at disease progression (p=0.01), and for 7 patients with all three event-point measures, TIMP-1 was 21.8% lower than baseline at first response and was 22.3% higher than baseline at disease progression (p=0.02, 0.04).
 Conclusions: In this study, serum levels of HER2, EGFR, uPA, and TIMP-1 do not appear to be predictive of response. However, interpretation of the data is compromised due to limited availability of serum at all 3 points. Serum HER2 appears higher at disease progression than at baseline. Serum TIMP-1 appears to decrease from baseline to first response, and then appears to increase at disease progression. Citation Information: Cancer Res 2009;69(2 Suppl):Abstract nr 6074.

Abstract Background: African Americans are underrepresented in cancer research trials. We propose that a collaborative effort with African American churches that brings cancer education events and a translational research project to the community with a mobile research unit will facilitate cancer awareness education and increase participation in translational research protocols among African Americans. Methods: From February 2010 through April 2010, Mayo Clinic partnered with predominantly African American Jacksonville churches, with the Volunteers in Medicine Clinic for the working uninsured, and with the local chapter of the Leukemia and Lymphoma Society, to provide 12 education programs focused on five behaviors that reduce cancer risk: proper nutrition, adequate fitness, tobacco cessation, breastfeeding, and limited alcohol consumption. Also offered during these programs was on site access to a translational cancer research pilot project evaluating the prevalence of t(14,18) and monoclonal proteins in African Americans and other racial/ethnic minorities. Translational research participation was facilitated by a mobile research unit (MRU), a converted RV that has sample collection, processing, and storage capabilities. Participants were surveyed on health knowledge and their willingness to participate in research. Results: A total of 236 of 318 participants (74%) returned written questionnaires provided during the education events. Forty-three percent resided in a health zone of Jacksonville that has worse health outcomes compared with any other sections of the city. Eighty-three percent responded that they had some type of health insurance and 88% had a primary care provider. Sixty-seven percent had never received any information on multiple myeloma, and 57% had never received information about clinical research studies. Sixty percent responded that they would participate in research studies and 61 % indicated that they would participate if the research focused on health issues that impacted African Americans. A total of 175 of 318 participants (55%) were enrolled in the translational research feasibility study. Seventy-two percent of the research study participants were female and 77% were African American, 19% Caucasians, and 4% Hispanic or of other racial/ethnic group. Ninety-three percent agreed to have their samples used for research of other health conditions and 85% agreed to have their samples shared with other research institutions. Conclusions: A high rate of participation in a blood collection translational research project was observed among this predominantly African American population. Most participants expressed a positive attitude towards research participation. Partnerships with African American churches in projects that provide community education and bring research to church venues with a mobile research unit are effective in improving health literacy related to a targeted topic, and in increasing translational research participation among African Americans. Funded by Mayo Clinic Comprehensive Cancer Center (NCI grant number P50-CA01508), Mayo Clinic CTSA (NCRR/NIH Grant Number 1 UL1 RR024150) and Active SPORE CA 90297052. Citation Information: Cancer Epidemiol Biomarkers Prev 2010;19(10 Suppl):A38.

The ECG lead monitoring algorithm was developed to provide a tool to assist bedside nurses in monitoring ST segments and dysrhythmias.

Abstract Heat shock protein 90 (HSP90), which regulates the functions of multiple oncogenic signaling pathways, has emerged as a novel anticancer therapeutic target, and multiple small-molecule HSP90 inhibitors are now in clinical trials. Although the effects of HSP90 inhibitors on oncogenic signaling pathways have been extensively studied, the effects of these agents on tumor suppressor signaling pathways are currently unknown. Here, we have examined how HSP90 inhibitors affect LATS1 and the related protein LATS2, two kinases that relay antiproliferative signals in the Hippo tumor suppressor pathway. Both LATS1 and LATS2 were depleted from cells treated with the HSP90 inhibitors 17-allylamino-17-demethoxygeldanamycin (17-AAG), radicicol, and PU-H71. Moreover, these kinases interacted with HSP90, and LATS1 isolated from 17-AAG–treated cells had reduced catalytic activity, thus showing that the kinase is a bona fide HSP90 client. Importantly, LATS1 signaling was disrupted by 17-AAG in tumor cell lines in vitro and clinical ovarian cancers in vivo as shown by reduced levels of LATS1 and decreased phosphorylation of the LATS substrate YAP, an oncoprotein transcriptional coactivator that regulates genes involved in cell and tissue growth, including the CTGF gene. Consistent with the reduced YAP phosphorylation, there were increased levels of CTGF, a secreted protein that is implicated in tumor proliferation, metastasis, and angiogenesis. Taken together, these results identify LATS1 and LATS2 as novel HSP90 clients and show that HSP90 inhibitors can disrupt the LATS tumor suppressor pathway in human cancer cells. Cancer Res; 70(21); 8642–50. ©2010 AACR.

Purpose We examined associations between tumor characteristics (human epidermal growth factor receptor 2 [HER2] protein expression, HER2 gene and chromosome 17 copy number, hormone receptor status) and disease-free survival (DFS) of patients in the N9831 adjuvant trastuzumab trial. Patients and Methods All patients (N = 1,888) underwent chemotherapy with doxorubicin and cyclophosphamide, followed by weekly paclitaxel with or without concurrent trastuzumab. HER2 status was determined by immunohistochemistry (IHC) and fluorescent in situ hybridization (FISH) at a central laboratory, Mayo Clinic, Rochester, MN. Patients with conflicting local positive HER2 expression results but normal central laboratory testing were included in the analyses (n = 103). Results Patients with HER2-positive tumors (IHC 3+, FISH HER2/centromere 17 ratio ≥ 2.0, or both) benefited from trastuzumab, with hazard ratios (HRs) of 0.46, 0.49, and 0.45, respectively (all P < .0001). Patients with HER2-amplified tumors with polysomic (p17) or normal (n17) chromosome 17 copy number also benefited from trastuzumab, with HRs of 0.52 and 0.37, respectively (P < .006). Patients who received chemotherapy alone and had HER2-amplified and p17 tumors had a longer DFS than those who had n17 (78% v 68%; P = .04), irrespective of hormone receptor status or tumor grade. Patients with HER2-normal tumors by central testing (n = 103) seemed to benefit from trastuzumab, but the difference was not statistically significant (HR, 0.51; P = .14). Patients with hormone receptor–positive or –negative tumors benefited from the addition of trastuzumab, with HRs of 0.42 (P = .005) and 0.60 (P = .0001), respectively. Conclusion These results confirm that IHC or FISH HER2 testing is appropriate for patient selection for adjuvant trastuzumab therapy. Trastuzumab benefit seemed independent of HER2/centromere 17 ratio and chromosome 17 copy number.