bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract P3-14-07: A Phase 1 Study To Assess the Safety, Tolerability and PK of ARRY-380 — An Oral HER2 Inhibitor
Authors: Moulder SL , Baetz T , D ‘Aloisio S , Fernetich G , Sajan B , Freeman B , Barrett E , Guthrie KM , Kass CL , Chia SKL. MD
Published: April 2011

Background: ARRY-380 is a potent, orally active small molecule that selectively inhibits HER2 signaling in vitro and in vivo. Methods: Patients (pts) with HER2+ MBC or other tumor types with published evidence of HER2 expression were treated with ARRY-380 as a single oral dose on C1 D1 followed by continuous dosing on C1 D3, in a BID schedule. Dose escalation was performed using standard DLT rules after all pts within a cohort completed the first cycle. Serial PK assessments were performed on D 1, 2, 3 and on D 15 of C1. Preliminary Results: As of 1 June, 2010, 30 pts have been treated at doses ranging from 25 mg to 800 mg BID. Pts had a median age of 57 years (range: 31-77 years). Cancer types included HER2+ MBC (n=23), CRC (n=6), and salivary gland (n=1). All HER2+ MBC pts had received prior trastuzumab and most (78%) had received prior lapatinib. The MTD (600 mg BID) has been determined in the dose escalation study. ARRY-380 was generally well-tolerated. One DLT of reversible grade 3 AST and one DLT of reversible grade 3 AST/ALT were noted at the 800 mg BID dose level. Other related AEs have been predominantly mild and include Grade 1 nausea, rash, and fatigue and Grade 2 fatigue. No treatment-related cardiac events have been reported. PK analyses indicate a trend for increasing Cmax and AUC with increasing dose, a median Tmax of 2 hours, and a mean t1/2 of 5.3 hours across doses. Of the 15 HER2+ MBC pts evaluable for response at doses ≥200 mg BID, 1 (7%) pt had a confirmed PR and 8 (53%) had SD accompanied by regression of tumor lesions. Of the 8 pts with SD, 5 pts demonstrated SD for ≥4 months. The rate of PR + SD ≥4 months was 43%. The HER2+ MBC pt with a PR had complete regression of all target and non-target lesions and a change in CEA from 71.1 to 3.5 ng/mL [NR = 0.0-3.0 ng/mL]. This pt is currently in Cycle 5 and has received 6 prior lines of therapy for advanced disease with a best response of PD on lapatinib. Conclusions: ARRY-380 has demonstrated a generally well-tolerated safety profile with a low incidence and severity of rash and diarrhea. ARRY-380 has shown promising signs of antitumor activity in a heavily pretreated HER2+ MBC population (43% of pts with PR or SD ≥4 months). These safety and preliminary efficacy data compare favorably to historical data of other HER2-targeted agents.

Citation Information: Cancer Res 2010;70(24 Suppl):Abstract nr P3-14-07.
Abstract P3-14-14: Activation of SRC and MAPK Mediates Resistance to Lapatinib in ER-/ERBB2+ Breast Cancer
Authors: Jegg A-M , Ward T , Iorns E , Gallas M , Pegram M.
Published: April 2011

Background

Resistance to lapatinib represents a significant problem in the treatment of ERBB2+ breast cancer, especially after failure of prior trastuzumab therapy. A number of resistance mechanisms have been identified in recent years that limit the anti-tumor effects of lapatinib. These mechanisms include the activation of ERα signaling, and sustained activation of AKT either through coexpression of receptor tyrosine kinases (IGF-IR, AXL, MET, ERBB3), loss of PTEN or activating mutations in PIK3CA. However, these mechanisms have been shown to confer resistance primarily in ER+/ERBB2+ breast cancer cells; the mechanisms of resistance to lapatinib in ER-/ERBB2+ breast cancer (∼ 50% of ERBB2+ breast cancers) remain poorly understood.

We hypothesized that resistance to lapatinib in ER-/ERBB2+ breast cancer cells can be mediated by activation of alternative signaling pathways which bypass the PI3K/AKT signaling axis. Materials and Methods

To investigate the mechanisms of acquired resistance to lapatinib we generated ER-/ERBB2+ and ER+/ERBB2+ breast cancer cells resistant to lapatinib at the clinically relevant concentration of 2.6 uM (SK-lapR and BT-lapR). Resistance to lapatinib was achieved by exposure of SKBR3 and BT474 cells to increasing concentrations of the drug, and resistant cells were selected through serial passage. The activation status of multiple signaling pathways in lapatinib sensitive and resistant cells was determined by western blot analysis, and the role of individual signaling pathways in promoting resistance to lapatinib was investigated in cell proliferation assays using selective small molecule inhibitors. Results and Discussion

Consistent with previous studies, we show that inhibition of ERα signaling using 100 nM fulvestrant in combination with lapatinib restored lapatinib sensitivity and effectively leads to growth arrest and cell death in ER+/ERBB2+ BT-lapR cells (P<0.001). However, none of the previously proposed mechanisms of resistance to lapatinib were apparent in our SK-lapR cells (ER-/ERBB2+). Strikingly, these cells which are stably resistant to lapatinib show no activation of AKT even in the absence of lapatinib. Rather, they exhibit increased phosphorylation of SRC kinase (3.9 fold, analyzed with ImageJ) and no inhibition of ERK1/2 after treatment with lapatinib compared to sensitive SKBR3 cells. Inhibition of SRC activity with 1uM AZD0530 in combination with lapatinib significantly reduced growth of SK-lapR cells compared to treatment with lapatinib alone (P<0.05). These data reveal potential novel mechanisms for lapatinib resistance in ER-/ERBB2+ breast cancer, and may indicate therapeutic strategies to prevent or overcome lapatinib resistance.

Citation Information: Cancer Res 2010;70(24 Suppl):Abstract nr P3-14-14.
Abstract P3-14-15: Lapatinib Plus Capecitabine in Trastuzumab Pre-Treated HER2- Positive Metastatic Breast Cancer: The Canadian Lapatinib Expanded Access Program Experience
Authors: Chang J , Chia S , Prady C , Haider K , Grenier D , Lopez P , Ghedira S.
Published: April 2011

BACKGROUND:

The global lapatinib expanded access programme (LEAP) provided access to lapatinib (L) combined with capecitabine (C) for women with HER2- positive metastatic breast cancer (MBC) who were previously treated with an anthracycline (A), taxane (T) and trastuzumab (TRAS) in 45 countries. The eligibility criteria and the dose of given drugs resembled that used in the phase III clinical trial EGF 100151, with the exception that patients (pts)previously exposed to C and pts with an ECOG performance status of 2 could be included as well. L is a dual tyrosine kinase inhibitor of EGFR and HER2. L plus C is an effective treatment option in T-refractory HER2-positive MBC. 148 Canadian pts were enrolled. METHODS:

LEAP enrolled patients with ErbB2-positive, locally advanced and MBC showing progressive disease following prior therapies with A, T, TRAS-containing regimens. Pts received L (1,250 mg/day) and C (2,000 mg/m2/day, days 1-14, every 21 days). We analyzed 148 Canadian LEAP pts recruited from seven centres for demographics, duration of therapy, L compliance, left ventricular function, progression-free survival (PFS), and overall survival (OS). RESULTS:

Data cut off for these analyses was Jan 10, 2010. [Data were not available for all 148 pts. The median age was 52 y (range 29-80 y) for 147 pts (145 women, 2 men). Ninety-five pts (64.6%) had no prior C and 52 (35.4%) had prior C (n=147). Study medication was discontinued for 142 pts (96.6%): 110 (74.8%) for progressive disease; 17 (11.6%) for adverse events; 8 (5.4%) for patient preference; and 3 (2%) were transitioned to the commercial supply of L. The median duration of therapy was 18.95 wk, ranging 0.1 to 96 wk (n=137). L compliance data showed that 73.7% of pts received ≥80% of the L dose (n=122). Median baseline left ventricular function (LVF) was 60%, ranging 50% to 81% (n=142). The median LVF at study end was 61%, ranging 37% to 83% (n = = 96). There was an amendment in June 2008 to end the collection of disease progression and/or death dates. Median PFS and OS were 22.1 wk (95% CI: 18.9-26.7), and 48 wk (95% CI: 42.3-…), respectively (n=71). CONCLUSIONS:

L combined with C is an effective option for women with refractory HER2-positive MBC. The results seen in the Canadian subset enrolled in LEAP were similar to the results of EGF 100151 and the Global LEAP in the efficacy and safety data.

Citation Information: Cancer Res 2010;70(24 Suppl):Abstract nr P3-14-15.
Abstract P3-14-16: Can Resistance to Trastuzumab Be Reversed by Endocrine Therapy? Results from a Proof-of-Principle Trial in Postmenopausal Patients with Hormonal Receptor (HR) Positive, HER-2 Positive Advanced Breast Cancer (SAKK 23/03)
Authors: Koeberle D , Ruhstaller T , Jost L , Pagani O , Zaman K , von Moos R , Crowe S , Thuerlimann B
Published: April 2011

Introduction: Trastuzumab (T) is a cornerstone in the treatment of patients with HER2-overexpressing advanced breast cancer and development of resistance to T is a major therapeutic problem. HER-2 is part of a highly interactive signaling network that may impair efficacy of endocrine therapy. A sequential treatment design was chosen in this trial to ensure complete resistance to single agent therapy before receiving both a non-steroidal aromatase inhibitor (AI) and T. Any kind of clinical activity with combined treatment of AI and T after progression of single agent treatments could indicate restoration of sensitivity as a consequence of cross-talking and networking between both pathways.

Methods: Key eligibility criteria included postmenopausal patients (pts.) with advanced, measurable, HER-2 positive (assessed by FISH, ratio (≥2)), HR positive disease and progression on prior treatment with a non-steroidal AI, e.g. letrozole or anastrozole, either in an adjuvant or advanced setting. Pts. received standard dose T monotherapy either weekly or three-weekly in step 1 and upon disease progression, continued T in combination with letrozole in step 2. The primary endpoint was clinical benefit response (CBR: CR, PR or SD for at least 24 weeks (+/− 1 week) according to RECIST) in step 2.

Results: Thirteen pts. were enrolled in five centers in Switzerland. In step 1, six pts. (46%) achieved CBR. Median time to progression (TTP) was 161 days (Range: 50 - 627). Based on data collected until the end of May 2010, CBR was observed in seven out of the eleven evaluable pts. (64%) in step 2, including one pt. with partial response. Four of the seven pts. within step 2 that achieved CBR also had CBR in step 1. Seven out of eleven pts. have documented tumor progression during step 2 treatment. Median TTP for all eleven pts. was 184 days (range 61 - 471). Mean time on study treatment (TTP in step 1 plus TTP in step 2) for pts. reaching step 2 was 380 days (range 174 - 864). Adverse events were generally mild. Conclusion: Results of this proof-of-principle trial suggest that complete resistance to both AI and T can be overcome in a proportion of pts. by combined treatment of AI and T, as all pts. served as their own control. Our results appear promising for a new treatment strategy which offers a chemotherapy-free and well-tolerated option for at least a subset of the pts. with HR positive, HER-2 positive breast cancer. Further trials will need to corroborate this finding.

Citation Information: Cancer Res 2010;70(24 Suppl):Abstract nr P3-14-16.
Abstract P3-14-18: Result of Phase I Part of a Phase I/II Study of Lapatinib (L) Plus Oral Vinorelbine (oVNR) in Metastatic Breast Cancer (MBC) Patients Overexpressing ErbB2
Authors: Lu Y-S , Lin C-H , Shen Y-C , Lan CY , Cheng A-L , Huang C-S.
Published: April 2011

Background: Lapatinib(L) is effective in overexpressing ErbB2 MBC patients. Oral vinorelbine (oVNR) has similar efficacy to that of the injection formulation and has demonstrated generally favorable tolerability. Oral medications provide convenient treatment and satisfactory life quality. We investigated the combination of L + VNR, seeking the recommended dose for further phase II studies.

Methods: This open-label, single-center, single-arm phase I study aimed to determine the maximum tolerated dose (MTD) and recommended dose of L+oVNR to women with ErbB2+ MBC, in progression after trastuzumab-taxane based treatment. Patients refractory to any line of anticancer treatment including vinorelbin was allowed to join in this study, except lapatinib. Cohorts of 3 patients received oral vinorelbine at escalating doses of 30-80 mg/day on day 1 and day 8 every 3 weeks (q3w) combined with lapatinib at escalating doses of 1000-1250 mg/day given po continuously, with each successive cohort until the MTD (2/3, 2/6 patients experiencing dose-limiting toxicity [DLT]) was reached. Additional patients received repeated administration at the recommended dose. G-CSF as primary prophylaxis of febrile neutropenia was not permitted.

Results: 15 patients have been enrolled. During dose escalation, 2 of 3 experienced DLT at dose level of oral vinorelbine 60mg/kg day 1 and day 8 q3w plus lapatinib 1000mg/day. One with grade 3 diarrhea and the other with prolonged grade 3 neutropenia. This dose level was declared as MTD. Therefore we stop recruiting patient into next dose level of oral vinorelbine 60mg/kg plus lapatinib 1250mg/day. Recommended dose was lapatinib 1000mg/day with oral vinorelbine 50mg/kg day 1, day 8 q3w. There is no DLT experienced in 12 patients with the dose level below MTD. The most frequent treatment-related adverse events included diarrhea, skin rash, and fatigue, mostly grade 1-2. As of June 2010, 6 patients are still on treatment. Efficacy signals in other 9 patients included 2 maintained stable disease (SD) > 6 months (both of them had unconfirmed PR during treatment), 1 unconfirmed partial response (PR), 1 SD < 6 months, and 4 progressive disease (3 of them had refractory to vinorelbine before). Phase II study with the recommended dose was just starting. Final result of the efficacy signal in phase I part and the toxicity profile of more patients receiving recommended dose will be presented at the meeting.

Conclusions: The combination of lapatinib and oral vinorelbine is tolerable and has antitumor activity in heavily pretreated ErbB2+ MBC. Further phase II study is underway.

Citation Information: Cancer Res 2010;70(24 Suppl):Abstract nr P3-14-18.
Abstract P3-15-01: Primary G-CSF Prophylaxis during the First Two Cycles Only or Throughout All Chemotherapy Cycles in Breast Cancer Patients at Risk of Febrile Neutropenia: Final Results from a Phase III Trial of the Dutch Breast Cancer Trialists Group (BOOG)
Authors: Aarts MJB , Peters FPJ , Dercksen WMW , Mandigers CMPW , Stouthard J , Nortier HWR , Kamm YJL , van Warmerdam LJC , van der Rijt CCD , van Gastel S , Joore M , Kessels A , Tjan-Heijnen VCG.
Published: April 2011

Background: In patients at risk of febrile neutropenia (FN), the highest incidence of FN is seen during the first chemotherapy cycles. This observation questions the effectiveness of continued G-CSF prophylaxis during later chemotherapy cycles.

Methods: We conducted a multicenter phase III study in breast cancer. Patients were randomized to primary G-CSF prophylaxis during the first two chemotherapy cycles only (experimental arm) or to primary G-CSF prophylaxis throughout all chemotherapy cycles (standard arm). Patients were eligible if treated with 3-weekly chemotherapy, being at risk of FN according to criteria of international guidelines. This was an equivalence study, aimed to include 230 patients, with FN as primary endpoint. Results: An independent Data Safety Monitoring Board recommended premature closure of the study because of an unacceptable increase in FN rate in the experimental arm. In total, 85 patients enrolled the experimental arm and 84 patients the standard arm till 15 th December 2009. Baseline characteristics were well belanced: age ≥65years in 93% versus 95%, ECOG PS of zero in 82% versus 88%, treated with docetaxel containing chemotherapy in (neo)-adjuvant setting in 97% versus 100%, respectively. At least one episode of FN was seen in 27 (32%) patients of the experimental arm compared with 4 (5%) patients in the standard arm (P<0.0001). Notably, in the experimental arm FN occurred predominantly in the third cycle, whereas in the standard arm FN was seen throughout all cycles. Fever, infection and/or mucositis led to serious adverse events in 31 (36%) patients in the experimental arm versus in 11 (13%) patients in the standard arm (P<0.001). In both arms, 5% of patients stopped or changed treatment because of toxicity. Most patients experienced only one FN episode. In these, chemotherapy was mostly given as planned, but in 61 % of patients with secondary G-CSF prophylaxis and in 21% of patients with secondary antibiotic prophylaxis (in the experimental arm). Conclusion: We conclude that in patients at risk of FN, primary G-CSF prophylaxis cannot be limited to the first chemotherapy cycles, despite the well known increased FN incidence in the first cycles without prophylaxis. With only 2 cycles protected, FN risk is increased in the third cycle. Support: The Netherlands organization for health research and development (ZonMw) and sanofi-aventis Netherlands B.V.

Citation Information: Cancer Res 2010;70(24 Suppl):Abstract nr P3-15-01.
Abstract P3-15-03: Prospective Evaluation of the Drug-Metabolizing Enzyme Polymorphisms and Toxicity Profile of Docetaxel in Korean Patients with Early-Stage Breast Cancer Receiving Adjuvant Chemotherapy
Authors: Kim K-p , Ahn J-H , Jung K-H , Kim S-B , Lim H-S.
Published: April 2011

Background

Interindividual variability of pharmacokinetics may account for unpredictable toxicity of docetaxel. Pharmacogenetic analysis on the metabolic pathway of docetaxel has been performed in hopes of identifying predicting factors related to docetaxel-related toxicities. Methods

From March 2007 to June 2008, female patients with early-stage breast cancer receiving docetaxel-containing adjuvant chemotherapy were included in this study. The 4 cycles of planned dose of docetaxel (100 mg/m2) was performed as adjuvant chemotherapy, following 4 cycles of adriamycin and cyclophosphamide. We evaluated toxicity profile of docetaxel according to the drug-metabolizing enzyme or transporter polymorphisms reported to affect the pharmacokinetics of docetaxel. The single nucleotide polymorphisms (SNPs) of CYP3A5 gene (6986 A>G), ABCB1 gene (1236 C>T, 3435 C>T, 2677 G>T(A)), ABCC2 gene (-24C>T, 1249 G>A, 3972 C>T, rs1276549) and SCLO1B3 gene (334 T>G, 699 G>A, rs11045585) were sequenced from individuals. Toxicities including neutropenia was evaluated for the 1st cycle of docetaxel and defined according to the NCI CTCAE version 3.0. Results

Pharmacogenetic analysis was performed in 218 Korean women who had received the planned chemotherapy. With regard to ABCB1 3435 C>T, ABCB1 3435 T/T had significantly higher risks of neutropenia (P=0.015). Meanwhile, allele frequencies for CYP3A5 6986 G and ABCB1 3435 T revealed a trend for neutropenia (P=0.107 and 0.068). We could not find any other association between genotypes and other toxicities. Discussion

Although ABCB1 3435 T/T was significantly associated with docetaxel-related neutropenia in our study population, polymorphism of pharmacogenetic genes related to docetaxel metabolism did not appear to be evidently associated with docetaxel-related adverse events. Further external validation of these genes may be required to allow them to enter clinical practice.

Citation Information: Cancer Res 2010;70(24 Suppl):Abstract nr P3-15-03.
Abstract P3-15-04: The Clinical Course of Metastatic Breast Cancer Patients with a Congestive Heart Failure after an Epirubicin-Based Treatment
Authors: Ryberg M , Nielsen DL , Jensen MT , Nielsen G , Andersen PK.
Published: April 2011

Background. Patients treated with epirubicin are at risk for the development of congestive heart failure (CHF). However, only limited data are available, which is describing the outcome for these patients. Materials and methods. The patients were recruited consecutively from 1,097 anthracycline-naïve patients with metastatic breast cancer, who were admitted to Herlev Hospital from November 1983 - November 2003 for an epirubicin-based chemotherapy. No patients had a history of heart disease. The outcome of 125 metastatic breast cancer patients, with a CHF Class ≥ II according to New York Heart Association (NYHA) guidelines after an epirubicin-based treatment, was performed retrospectively. Univariate ordinal logistic regression was used to study the association between NYHA class at time of CHF (II, III or IV) and possible risk factors. Two multiple Cox regression analyses (events: mortality of cardiotoxicity and mortality of other causes) were performed to evaluate the risk of cardiac death.

Results. CHF was developed within a median time from cessation of epirubicin on 3 months (range 0 to 66.7 months). The distribution of patients in the NYHA class II, III, IV and NYHA class ≥2 was; 35 patients (28%), 38 patients (30.4%), 39 patients (31.2%) and 13 patients (10.4%), respectively. No predictive factors, including increasing cumulative dose of epirubicin, co-morbidities and age had an influence on the severity of CHF. The median survival of all patients was 7.3 months (range 0.01 to +121.1 months). Deaths caused by CHF occurred in 27 patients (21.6%) in a median time on 1.2 months (range 0 - 77 months), including 6 patients (4.8%) dying of cardiac failure < 10 days after the diagnosis. Only the NYHA class patient had was the only risk factor for dying of CHF (p = 0.0003). Compared to patients in NYHA class II and III, the risk of cardiac death in patients with NYHA class IV was increased with 93% and 74%, respectively.

Survival in patients with stable CHF was comparable to other patients with metastatic breast cancer.

Discussion. Although the information is obtained retrospectively during a period of twenty years; the study is important, as only limited information is available on the course of CHF patients. The study showed that the risk of cardiac death were closely related to the degree of cardiac toxicity as the risk of cardiac death was 93% or 74% higher for patients in NYHA class IV than for patients in NYHA class II or III. Moreover, the outcome for the patients when CHF is stabilized can be expected to be as patients without CHF. Furthermore, no predictive factors were identified, including increasing cumulative dose of epirubicin for the severity of cardiotoxicity developed.

Citation Information: Cancer Res 2010;70(24 Suppl):Abstract nr P3-15-04.
Abstract P3-15-07: Late Toxicity and Patient Self-Assessment of Breast Appearance/Satisfaction on RTOG 0319: A Phase II Trial of Three Dimensional Conformal Radiation Therapy (3DCRT) Partial Breast Irradiation (PBI) Following Lumpectomy for Stage I and II Breast Cancer
Authors: Chafe SJ , Moughan J , Vicini FA , Wong JWC , Pass HA , Rabinovich RA , Arthur DW , Petersen IA , White JR , McCormick B.
Published: April 2011

Background:

Ongoing phase III trials are comparing accelerated PBI around the lumpectomy site to standard whole breast irradiation in early stage breast cancer. Recent studies have questioned the late toxicity from PBI delivered with 3DCRT. We evaluated late toxicity on patients (pts) treated on RTOG 0319.

Methods:52 pts with stage I-II breast cancer, ≥3 cm, negative margins, and < 3 nodes positive were treated with 3DCRT PBI and received 38.5 Gy in 10 BID fractions over 5 days. Median follow-up was 5.3 years (min-max: 1.7-6.4 years). Five pts were deceased at 5 years. NCI CTCAE v3.0 was used to grade adverse events (AEs). At 3 years, 50 pts were available for AE reporting or late toxicity assessment.

Results: The treatment-related AEs were primarily grade 1 36.5%, 50% grade 2, and only 3 (5.8%) pts had grade 3 AEs. The grade 3 AEs in these pts were radiation dermatitis, skin fibrosis, telangectasia, and myositis. No baseline self-assessment of breast appearance is available. Patient self-assessment of breast appearance and satisfaction was available in 45 (87%) pts at 1 year and 36 (72%) at 3 years. At 1 year, 82% self reported excellent/good breast appearance and at 3 years, 64%. Pts who reported < excellent/good breast appearance had significantly more grade ≥2 skin AEs (telangectasia, skin atrophy, hyperpigmentation, erythema, skin fibrosis, radiation dermatitis, and breast or nipple/areolar deformity). At 3 years, 31 of 36 pts were satisfied with their treatment, but all 36 would choose 3DCRT PBI again over standard breast RT. Treatment factors were evaluated to establish correlation with patient reported breast appearance. Preliminary observations suggest surgical bed volume, PTV volume, number of beams used, and possible use of bolus are associated with patient reported outcomes (PRO). Surgical factors impacting PRO were not collected.

Conclusions: The majority of pts were satisfied with their treatment results at 3 years and would all choose to have 3DCRT PBI again. With a median of 5.3 years, the rate of grade 3 AEs is low. Technical factors from this phase II trial were modified for the phase III NSABP B-39/RTOG 0413 trial, in which bolus is not allowed and both target volumes and required coverage were modified, increasing expectation of improved PRO. Supported by RTOG U10 CA21661, CCOP U10 CA37422 and ATC U24 CA 81647 NCI grants.

Citation Information: Cancer Res 2010;70(24 Suppl):Abstract nr P3-15-07.
Abstract P4-01-05: Lapatinib Resistance Confers Cross-Resistance to Microtubule Inhibitors in ErbB2-Overexpressing Breast Cancer Cells
Authors: Araki K , Gallas M , Liu X , Jegg A-M , Ward T , Pegram M.
Published: April 2011

ErbB2 gene amplification occurs in approximately 20% of all breast cancers and is associated with an aggressive subtype with poor clinical prognosis. Acquired resistance to erbB2-targeted therapies is a common occurance even in patients who show an initial clinical response to trastuzumab or lapatinib-based regimens. Optimal treatment for relapsed disease following erbB2 targeted therapies has not been defined. Therefore, we sought to evaluate whether cytotoxic agents have cross-resistance to lapatinib in cell lines that have acquired resistance to lapatinib. Here we established a model of acquired lapatinib resistance derived from erbB2 overexpressing breast cancer cell lines, BT474 and SKBR3. Lapatinib resistant cell lines, BTLapR and SKLapR, were generated by growth in the continuous presence of 2.6 uM lapatinib (peak plasma concentration in human subjects) resulting in IC50 of 4.72 uM (BTLapR) and 0.083 uM (BT474) (P<0.0001). IC50 of SKLapR and SKBR3 were 17.72 uM and 0.1637 uM (P<0.0001), respectively. Next, resulting cells were treated with different classes of chemotherapeutics (2 taxanes, 2 vinca-alkaloids, 1 epothilone, 1 anthracycline, 1 antimetabolite, 2 platinums, and 1 alkylating agent) in order to address cross-resistance to lapatinib. BT474 and BTLapR as well as SKBR3 and SKLapR were exposed to each chemotherapeutic agent and cell viability was evaluated at 72h using a cell proliferation assay (AlamarBlue®). We found that BTLapR was significantly cross-resistant to 5 microtubule inhibitors (paclitaxel, docetaxel, vinorelbine, vinblastine, and ixabepilone), but SKLapR was not (shown in table below).

IC 50 of Significant Drugs on Lapatinib Resistant BT474 cells

Previous studies have shown that resistance to lapatinib can switch cell survival signaling from the erbB2 to ER pathway, and that ER signaling can stimulate beta III tubulin expression. We therefore hypothesize the differential responses to microtubule inhibitors in BTLapR and SKLapR might be explained by the fact that BT474 has both estrogen (ER) and progesterone receptor (PR), but SKBR3 express neither. Moreover, beta-III tubulin is known to be a key resistance mechanism of microtubule inhibition. Western blot analysis of baseline protein expression in both acquired lapatinib-resistant and parental cells indicates that ER signaling is involved in stimulating beta III tubulin and is linked to the development of resistance to microtubule inhibitors. These findings point to a potential mechanism to address cross resistance between lapatinib and microtubule-interacting agents, which could have potential clinical implications when considering salvage therapies following lapatinib treatment.

Citation Information: Cancer Res 2010;70(24 Suppl):Abstract nr P4-01-05.
Abstract P4-02-04: Impact of CYP2D6 Genotype and Side-Effects on Adherence Rates to Tamoxifen in Premenopausal Breast Cancer Patients
Authors: Hubalek M , Oberguggenberger A , Meraner V , Beer B , Giesinger J , Oberacher H , Sperner-Unterweger B , Marth C.
Published: April 2011

Background: Only few studies have investigated the issue of breast cancer patients’ adherence to tamoxifen therapy and factors influencing adherence behavior. Especially in the context of different CYP2D6 genotypes adherence to tamoxifen has not been extensively studied yet. Variations in the CYP2D6 genotype, as well as patients taking inhibitors of CYP2D6 (e.g. antidepressants) contribute to different side effects and adherence rates to adjuvant tamoxifen.

Materials and Methods: 106 premenopausal breast cancer patients who met inclusion criteria were consecutively included in the study at the outpatient unit of the Department of Gynecology, Innsbruck Medical University. Within their routine after care appointment patients completed a comprehensive PRO assessment including the FACT-B/ES, the HADS and a self-report questionnaire on adherence behavior (SMAQ). The multi-method approach comprised the Simplified Medication Adherence Questionnaire, a semi-structured interview, physicians’ ratings and blood levels for tamoxifen metabolites. Additionally, the CYP2D6 genotype was determined in all patients participating in this part of the study.

Results: 19% patients were poor metabolizer (PM), 51% intermediate metabolizer (IM), 29% extensive metabolizer (EM) and 7% ultra-rapid metabolizer (UM). Significant group differences with regard to tamoxifen and endoxifen serum concentrations were found between the metabolization groups (p=0.044). UMs had the lowest tamoxifen and highest endoxifen concentrations. Only 3.2% of the patients analyzed had no measurable tamoxifen concentrations in their serum. All non-compilant patients were from the extensive metabolizer group. However, during follow-up 25% (2/6) of patients with UM genotype, 13% (4/30) with EM genotype, 1% (1/50) with IM genotype and 0% (0/19) with the PM genotype discontinued their tamoxifen therapy due to therapy related side effects. In addition anti-depressants were more frequently prescribed in the extensive metabolizer group leading to lower levels of the active metabolite endoxifen. Conclusion: A trend towards higher hot flashes was observed in the EM and UM group. We did not find significant differences in QOL between

CYP2D6 metabolizing groups. Trend level significance was found for global QOL (on a descriptive level) physical well-being and emotional well-being. Non-adherence to tamoxifen therapy is more frequent in patients with CYP2D6 EM and UM genotype. These preliminary data may explain part of the current controversy over CYP2D6 genotype predicting response to tamoxifen and suggest that patients most likely to benefit from tamoxifen are paradoxically most likely to stop their tamoxifen therapy prematurely.

Citation Information: Cancer Res 2010;70(24 Suppl):Abstract nr P4-02-04.
Abstract P4-02-08: STAT Signaling in Endocrine Sensitive and Resistant Breast Cancer — A Potential Target for Therapy?
Authors: Huang R , Faratian D , Harrison DJ , Langdon SP.
Published: April 2011

The STAT (for signal transducer and activators of transcription) family of molecules regulate cell growth, survival, and differentiation as important integrators of cytokine and growth factor receptor signaling. STAT3 and STAT5 have been implicated in endocrine resistance in breast cancer. We aimed to identify new modifiers of endocrine sensitivity in breast cancer using functional proteomic profiling of the estrogen/tamoxifen-sensitive MCF-7 cell line and its partially sensitive (LCC1) and fully resistant (LCC9) variants. Using an unsupervised analysis of the expression of 125 paired phosphorylated and non-phosphorylated epitopes in key oncogene and tumour suppressor pathways on antibody arrays, STAT1, STAT3, and their phosphorylated epitopes were differentially expressed between endocrine resistant and parental controls. Differential expression was confirmed by qRT-PCR and western blotting. The STAT1 inhibitor EGCG inhibited cell number in endocrine insensitive and resistant LCC1/LCC9 lines, while the STAT3 inhibitors static and WP1066 were equally effective in endocrine-resistant and parental lines. Reduction in cell number was shown to be due to induction of apoptosis rather than inhibition of proliferation. The effects of the STAT inhibitors were additive, rather than synergistic, when tested in combination with tamoxifen in vitro. Finally, expression of STAT 1 and STAT3 were measured by quantitative immunofluorescence in 54 invasive breast cancers and expression levels of STAT1 and STAT3 transcript analysed in 550 breast cancers from publicly available gene expression datasets (GSE2990, GSE12093, GSE6532). High expression of STAT1, and low expression of STAT3, were associated with a worse distant metastasis free survival (DMFS; RR 1.42 and 0.46; log-rank p=0.042 and P<0.0001, respectively). When treatment with tamoxifen was considered, STAT1 expression was nearly predictive of DMFS (log-rank p=0.067), while STAT3 expression was predictive of DMFS (log-rank P<0.0001). These results suggest that STAT signaling may be important in endocrine resistance, and that STAT inhibitors, particularly STAT1 inhibitors, may be effective therapies in breast cancer even in the resistant setting.

Citation Information: Cancer Res 2010;70(24 Suppl):Abstract nr P4-02-08.
Abstract P4-02-15: Overlapping Therapy with High-Dose Tamoxifen and CYP2D6 Inhibitors Modifies Medication Adherence in Breast Cancer Patients
Authors: Markward NJ , Tinnirello J , Khalid M , Frueh FW , Milayna S , Aubert RE , Epstein RS.
Published: April 2011

Background: Tamoxifen is a selective estrogen receptor modulator (SERM) commonly used to treat and prevent estrogen-receptor-positive (ERP) breast cancer. However, despite its established efficacy, recent studies indicate that 1 in 2 patients will discontinue use of the drug prior to completing the recommended five-year treatment cycle. The alleviation of unpleasant side effects through personalized targeting of tamoxifen dose and, as appropriate, concomitant administration of a CYP2D6 inhibitor could increase adherence and increase the likelihood that patients will receive the amount of the drug needed to protect them against disease onset and/or recurrence. Materials and Methods: A retrospective cohort study was implemented to assess whether overlapping therapy with highdose tamoxifen and CYP2D6 inhibitors influence medication adherence during the first 540 days of therapy. Subjects (N = 17,242) were identified in an integrated medical and pharmacy claims database and selected for inclusion in the study if they 1) filled a new tamoxifen prescription between 7/1/2008 and 6/30/2009 and 2) were continuously eligible to receive pharmacy benefits during the 6 months preceding and 12 months after initiation of therapy. Eligible participants were then cross-classified according to tamoxifen dose (NORMAL, HIGH) and CYP2D6 inhibitor status (NO, YES), and a Cox regression model was fitted to the data to obtain covariate-adjusted hazard ratios (HR) and 95% confidence intervals for each treatment combination relative to the HIGH/NO group.

Results: The lowest and highest hazard ratios were observed in the NORMAL/YES [HR = 0.924 (0.88, 0.97)] and HIGH/NO [HR = 2.33 (2.07, 2.62)] categories. Moreover, the risk of discontinuation was significantly reduced when high-dose tamoxifen was administered in concert with a CYP2D6 inhibitor [HR = 1.31 (0.953, 1.802)]. Discussion: These study suggests that joint treatment with high-dose tamoxifen and a CYP2D6 inhibitor may improve tamoxifen adherence in breast cancer patients. The findings also highlight the need for 1) education and research regarding tamoxifen dosing and 2) careful consideration for when–and under what circumstances–concomitant medications should be prescribed to enhance patient tolerance of tamoxifen-related negative symptoms.

Citation Information: Cancer Res 2010;70(24 Suppl):Abstract nr P4-02-15.
Abstract P4-05-05: Stromal Response to 14-Day Preoperative Therapy in Postmenopausal Oestrogen Receptor Positive Breast Cancer
Authors: Lo C , Busch S , Lee AG , Searle G , Lamb R , Cramer A , Winter MC , Coleman RE , Dixon M , Bundred NJ , Landberg G.
Published: April 2011

Background: Stromal-epithelial interaction is a key factor in tumour progression. Cancer-associated fibroblasts (CAFs) and macrophage infiltration have been associated with early relapse in breast cancer. Bisphosphonates are effective inhibitors of osteoclast activation in metastatic breast cancer but also have a general inhibitory effect on breast cancer progression. In order to monitor a potential tumour stromal response in breast cancer during treatment with an aromatase inhibitor and a bisphosphonate we analysed pre-and post-treatment samples from a neoadjuvant window study and focused on the presence of macrophages and CAFs.

Materials and methods: Tissue microarrays (TMAs) from surgical samples and pre-operative core biopsies were immunohistochemically stained for aSMA (CAF marker), CD68 (macrophages) and epithelial proliferation (Ki67). In order to validate if the presence of macrophages and aSMA could be monitored by the TMA approach, we initially analysed a screening cohort of 144 breast cancer samples. We then studied pre-and post-treatment samples from 110 postmenopausal ER-positive invasive breast cancer patients randomised to receive 14 days of preoperative treatment (placebo, Letrozole, or Letrozole plus Zoledronate). Results: In the screening cohort, we observed significant links between aSMA positive fibroblasts and disease recurrence as well as between CD68 positive macrophages and tumour size, grade, lymph node positivity and recurrence. This validated the use of TMAs for stromal analyses and furthersupported a link with key tumour biological events. In both treatment arms, there was a significant drop in absolute Ki67 value compared to placebo (-9.3% Letrozole and -13.1% combination reduction versus 1% increase, P<0.001). Post-treatment CD68 (median 35, range 3 to 117) was significantly linked to a Ki67 drop (p=0.045). Interestingly, this effect was mainly observed in the combination treatment group (p=0.002). aSMA expression was unaffected during treatment in 52%, increased in 35% and decreased in 13% of cases. Patients with aSMA reduction post treatment had a larger Ki67 fall compared to patients with increase or no change in aSMA (p=0.007).

Conclusion: Short term treatment response in the epithelial component of cancers was paralleled by specific responses in the tumour stromal component. These novel findings suggest that bisphosphonates and aromatase inhibitors have major effects on tumour stroma in vivo which might augment their inhibitory effect on tumour progression.

Citation Information: Cancer Res 2010;70(24 Suppl):Abstract nr P4-05-05.
Abstract P4-06-12: The Urokinase-Type Plasminogen Activator Is a Direct JAG1- Mediated Notch Target in Breast Cancer
Authors: Reedijk MJ , Shimizu M , Cohen B , Goldvasser P , Berman H , Virtanen C.
Published: April 2011

Aberrant activation of the Notch signaling system and over-expression of the Notch ligand JAG1 are associated with poor outcome in breast cancer. The plasminogen activator system, which includes urokinase-type plasminogen activator (uPA), has been validated as a marker of recurrence, high metastasis risk and death in breast malignancy. By using microarray profiling of breast cancer cell lines that had undergone siRNA-mediated abrogation of Notch signaling we uncovered a link between activated Notch and uPA expression. An association between elevated expression of the Notch ligand JAG1, uPA and the basal breast cancer subtype was confirmed in breast cancer cell lines. The association between JAG1 and uPA expression persisted in a survey of primary carcinomas of the breast. We found that Notch knockdown reduced transcription of uPA and phenocopied uPA knockdown in breast cancer cells. Through mutational analysis we identified a CBF-1 binding site in the uPA promoter that is required for direct transcriptional regulation by Notch. These data suggest that JAG1-induced Notch activation results in breast cancer progression through up-regulation of the plasminogen activator system and directly links these two poor prognosis pathways.

Citation Information: Cancer Res 2010;70(24 Suppl):Abstract nr P4-06-12.
Abstract P4-06-14: Induction of Blimp-1 in Breast Cancer Cells Is Mediated by Activator Protein-1 and Promotes Transforming Growth Factor (TGF)-ß-Induced Epithelial-to-Mesenchymal Transition
Authors: Romagnoli M , Belguise K , Wang X , Sonenshein GE.
Published: April 2011

The vast majority of breast cancers has been shown to display aberrant constitutive expression of NF-kB subunits, which enhances their mobility and promotes their survival and malignancy. Recently, we demonstrated that the RelB NF-kB subunit promotes migration of breast cancer cells, in part via repression of estrogen receptor (ER)α transcription. Furthemore, we showed that inhibition of the ERα promoter was mediated by the zinc finger transcriptional repressor B lymphocyte-induced maturation protein (Blimp-1), transcribed from the PRDM1 gene, which is induced by RelB-mediated activation of a Bcl-2/Ras pathway. Indeed, we showed for the first time that Blimp-1 is present in breast cancer and is more highly expressed in ERα negative breast cancer cells where it plays an essential role in their migratory and invasive abilities. Blimp-1 is a well-known zinc finger repressor, which is critical for terminal differentiation of B and T cells and for migration of primordial germ cells. Here we have examined the mechanism of activation of Blimp-1 expression in breast cancer cells. Given the role of Ras to c-Raf signaling in migration of cancer cells, we tested the role of this downstream mediator. A constitutively active c-Raf led to induction of Blimp-1 in breast cancer lines while a dominant negative variant reduced its levels. RelB/p52 NF-kB complexes induced c-Jun, c-Fos, Fra-1 and Fra-2 Activator Protein-1 (AP-1) subunits, which activated the PRDM1 gene promoter. The transforming growth factor (TGF)-β induces a more mesenchymal phenotype of breast cancer cells. Treatment of NMuMG breast epithelial cells with TGF-β led to Blimp-1 activation through AP-1 and epithelial-to-mesenchymal transition (EMT). Importantly, Blimp-1 knockdown counteracted TGF-mediated EMT and wound healing in both NMuMG and MDA-MB-231 breast cells. Thus, activation of AP-1 complexes via a Ras/Raf/Erk signaling pathway mediates induction of PRDM1 gene transcription and Blimp-1 expression in ERα negative breast cancer cells, which is essential for TGF-β-induced mesenchymal phenotype. Therefore, downstream mediators of Blimp-1 activity might emerge as new therapeutic targets for treatment of patients with ERα negative breast cancer.

Citation Information: Cancer Res 2010;70(24 Suppl):Abstract nr P4-06-14.
Abstract P4-06-21: Matrix Metalloproteinase 2 in Tumor Cell-Induced Platelet Aggregation: Regulation by Cryptotanshinone
Authors: Linggeng Y , Yin L , Chenhu H , Junshan R , Lei Z , Li T.
Published: April 2011

Background: A correlation exists between the ability of tumor cells to aggregate platelets and their tendency to metastasize. Tumor cell-induced platelet aggregation (TCIPA) facilitates the embolization of the vasculature with tumor cells and the formation of metastatic foci. It is well documentedthat matrix metalloproteinases (MMPs) play an integral part in tumor spread and the metastatic cascade.

Material and Methods: Therefore, we have examined the role of MMPs during TCIPA and its regulation by Cryptotanshinone in vitro. Human breast cancer cell line MCF-7 and MDA-MB-231 induced TCIPA in a concentration dependent manner that was monitored by aggregometry. This aggregation resulted in the release of MMP-2 from platelets and cancer cells, as measured by RT-PCR.

Results:MCF-7 cells released significantly more MMP-2 than MDA-MB-231 cells and were more efficacious in inducing TCIPA. Inhibition of MMP-2 with phenanthroline (1-1000 mM), a synthetic inhibitor of MMPs, and by neutralizing anti-MMP-2 antibody (10 mg/ml) reduced TCIPA induced by MCF-7 cells. TCIPA was abolished by simultaneous inhibition of platelet function with acetylsalicylic acid (100 mM; thromboxane pathway inhibitor), apyrase (250 mg/ml; ADP pathway inhibitor), and phenanthroline. Cryptotanshinone inhibited TCIPA and MMP-2 release from platelets and tumor cells.

Discussion:We conclude that (a) human Human breast cancer cells aggregate platelets via mechanism (s) that are mediated, in part, by MMP-2; (b) Cryptotanshinone inhibits TCIPA, in part, by attenuating the release of MMP-2.

Citation Information: Cancer Res 2010;70(24 Suppl):Abstract nr P4-06-21.
Abstract P4-06-23: KLF8 Is Regulated by Its Novel Interacting Protein PARP-1
Authors: Lu H , Wang X , Urvalek AM , Li T , Yu L , Zhu J , Lin Q , Zhao J.
Published: April 2011

Krüppel-like factor 8 (KLF8) plays an important role in the progression of breast cancer invasion and metastasis. Using mass spectrometry and co-immunoprecipitation, we identified PARP-1, a chromatin-associated enzyme that catalyzes protein poly(ADP-ribosyl)ation (PARylation), as a novel KLF8 interacting protein. Co-IP and western blotting indicated that KLF8 is a PARylation substrate. Mutation of the cystein residues in the first and second zinc-finger motifs of KLF8 abolishes its interaction with PARP-1. Luciferase assays showed KLF8 activation on the cyclin D1 promoter was significantly reduced when PARP-1 was physically deleted or functionally inhibited or when KLF8-PARP-1 interaction was disrupted. Surprisingly, immunofluorescent staining demonstrated that KLF8 was localized in the nuclei of wide type MEFs, but was mislocalized in the perinuclear cytoplasm of the PARP-1-/- cells, which could be rescued by re-expression of PARP-1. The detection of γH2A.X, comet assay and clonogenic assay indicated KLF8 play a DNA repair role in a PARP-1- dependent matter. Cychloheximade-chase assay revealed the stability of KLF8 dramatically reduced by loss of PARP-1 or when KLF8-PARP-1 interaction was disrupted, meanwhile ubiquitination assay showed heavier ubiquitin chains of KLF8. This work has identified PARP-1 as a novel binding partner and post-translational modifier of KLF8 and a critical regulator of KLF8 localization, functions and stability in the cells and suggested a critical role of the interaction between KLF8 and PARP1 during metastatic progression of breast cancer. Support by: NIH, ACS and Komen grants to J.Z.

Citation Information: Cancer Res 2010;70(24 Suppl):Abstract nr P4-06-23.