Residual Cancers Research Articles

O3–056 - Utility of MRI and us for Evaluation of Minor Residual Diseases After Receiving Neoadjuvant Therapies to Breast Cancer

Background and Object: To decide operative procedure of primary breast cancer (PBC) in the patients who received neoadjuvant chemotherapy (NAC) or primary endocrine therapy(PET), images to monitor the response of breast cancerto their treatment are hoped. However, it is unclear if magnetic resonance imaging (MRI) and ultrasonography (US) are useful for evaluation of minor residual invasive cancers after treatment. Therefore we retrospectively compared the histological change with the change of image findings before and after the therapy to know image findings of suggesting therapeutic effect.

Multispectral Mueller polarimetric imaging detecting residual cancer and cancer regression after neoadjuvant treatment for colorectal carcinomas

This work is devoted to a first exploration of Mueller polarimetric imaging for the detection of residual cancer after neoadjuvant treatment for the rectum. Three samples of colorectal carcinomas treated by radiochemotherapy together with one untreated sample are analyzed ex vivo before fixation in formalin by using a multispectral Mueller polarimetric imaging system operated from 500 to 700 nm. The Mueller images, analyzed using the Lu-Chipmann decomposition, show negligible diattenuation and retardation. The nonirradiated rectum exhibits a variation of depolarization with cancer evolution stage. At all wavelengths on irradiated samples, the contrast between the footprint of the initial tumor and surrounding healthy tissue is found to be much smaller for complete tumor regression than when a residual tumor is present, even at volume fractions of the order of 5%. This high sensitivity is attributed to the modification of stromal collagen induced by the cancer. The depolarization contrast between treated cancer and healthy tissue is found to increase monotonously with the volume fraction of residual cancer in the red part of the spectrum. Polarimetric imaging is a promising technique for detecting short-time small residual cancers, which is valuable information for pathological diagnosis and patient management by clinicians.

Functional proteomics characterization of residual breast cancer after neoadjuvant systemic chemotherapy

BackgroundThe purpose of this study was to determine the functional proteomic characteristics of residual breast cancer and hormone receptor (HR)-positive breast cancer after neoadjuvant systemic chemotherapy, and their relationship with patient outcomes. MethodsReverse phase protein arrays of 76 proteins were carried out. A boosting approach in conjunction with a Cox proportional hazard model defined relapse predictors. A risk score (RS) was calculated with the sum of the coefficients from the final model. Survival outcomes and associations of the RS with relapse were estimated. An independent test set was used to validate the results. ResultsTest (n = 99) and validation sets (n = 79) were comparable. CoxBoost revealed a three-biomarker (CHK1pS345, Caveolin1, and RAB25) and a two-biomarker (CD31 and Cyclin E1) model that correlated with recurrence-free survival (RFS) in all residual breast cancers and in HR-positive disease, respectively. Unsupervised clustering split patients into high- and low risk of relapse groups with different 3-year RFS (P ≤ 0.001 both). RS was a substantial predictor of RFS (P = 0.0008 and 0.0083) after adjustment for other substantial characteristics. Similar results were found in validation sets. ConclusionsWe found models that independently predicted RFS in all residual breast cancer and in residual HR-positive disease that may represent potential targets of therapy in this resistant disease.

The role of radiotherapy-resistant stem cells in breast cancer recurrence.

Although the effectiveness of anti cancer therapies has improved dramatically, breast cancer (BC) remains the second leading cause for cancer-related death among women in the western world [1]. The observed improvement in the survival rate and remission for primary tumors are essentially related to the advances in early diagnosis with more sensitive imaging technology [2,3]. However, the death rate remains unacceptably high for BC patients due to hard-to-treat metastatic and recurrent tumors, necessitating new, effective approaches and anticancer agents. Accumulating evidence suggests that cancer stem cells (CSCs), which are present in many cancers including BC, mediate tumor metastasis and contribute to relapse due to their resistance to current conventional therapies [4,5]. These unique cells possess stem cell-like characteristics, such as the capacity of self-renewal, which makes the tumor capable of regenerating its entire bulk. CSCs are resistant to proapoptotic factors, rendering them a formidable adversary to anticancer agents. In part, this is related to their quiescence capacity, which holds them in a standby mode in their niche microenvironment, sheltering them from radiation and other anticancer agents, since these agents are effective only on highly proliferative cells [6,7]. In addition to their powerful DNA repair machinery, breast CSCs (BCSCs) express ALDH, which is suspected to play a part in their death-resistance phenotype by being involved with cell detoxification machinery [8,9]. Thus, BCSCs represent a major challenge in the battle against BC, and an opportunity to develop more effective target to treat metastatic lesions.

Abstract A91: Developing novel treatment strategies for triple-negative breast cancer metastasis

Abstract Despite the significant advances in breast cancer biology, there has been limited progress in the treatment of advanced breast cancer, with little change in the overall survival for women with metastatic breast cancer over the last several decades. There were approximately 180,000 new cases of breast cancer diagnosed in the United States in 2007. About 15% of these, accounting for 25,000 to 30,000 cases each year, are “triple-negative breast cancer” (TNBC) in that the tumors do not express the estrogen receptor (ER), progesterone receptor (PR), or HER-2/neu. Unlike therapies targeted against steroid hormone receptors or HER2, there is no Food and Drug Administration (FDA) approved targeted drugs for the treatment of TNBC, and hence these cancers are associated with the worst prognosis with patients ultimately succumbing to metastatic disease. Systemic therapies are effective initially in controlling and reversing tumor growth; however, residual cancers will invariably re-grow despite this initial response causing recurrence or metastasis at distant sites. The identification of new therapeutic targets against TNBC is crucial to improving the outcome of this subtype. Our goal is to eliminate TNBC metastasis by developing novel therapies capable of attacking and destroying the “lethal seeds” driving these cancers and metastases, the “tumor initiating cells” (TICs) or “cancer stem cells” (CSCs). Hypothesis: Targeting cancer stem cells will eliminate metastasis, ultimately improving survival in TNBC. Data from paired clinical human breast cancer biopsies showed that standard therapy every three weeks kills dividing daughter cells but not tumor-initiating cells (TICs), so that tumor tissue obtained after therapy is enriched for “tumor-initiating” or “cancer stem” cells. Using patient-derived tumorgraft models, we further discovered that despite complete eradication of primary TNBC tumors by multiple cycles of standard chemotherapy, metastases in distant sites ultimately developed, causing death. We analyzed the gene expression of CSC isolated from patient biopsies to identify therapeutic targets regulating CSCs. Based on the signaling pathways identified through this microarray analysis, we treated mice with small molecule inhibitors of STAT3 or the Notch signaling pathway and showed remarkable reduction of CSCs in primary tumors. We expect that the addition of CSC-targeted therapy to standard chemotherapy will eliminate metastasis. Single agent targeted therapy may only reduce metastatic burden, suggesting a cocktail of targeted-therapies will be required to eradicate metastasis, in which case, we would combine multiple therapies that target CSCs. Citation Format: Melissa D. Landis, Lacey A. Burey, Jenny C. Chang. Developing novel treatment strategies for triple-negative breast cancer metastasis. [abstract]. In: Proceedings of the AACR Special Conference on Tumor Invasion and Metastasis; Jan 20-23, 2013; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2013;73(3 Suppl):Abstract nr A91.

Transanal Endoscopic Microsurgery for Residual Rectal Cancer (ypT0-2) Following Neoadjuvant Chemoradiation Therapy

Significant tumor downstaging among patients with rectal cancer following neoadjuvant chemoradiation has raised the issue of offering patients with small residual cancers restricted to the bowel wall an alternative treatment strategy to total mesorectal excision. Transanal endoscopic microsurgery may allow proper primary tumor resection with promising oncological outcomes, less postoperative morbidity, and minimal long-term sexual, urinary, and fecal continence disorders in comparison with radical resection. The aim of this study was to determine the oncological outcomes of patients with residual rectal cancers restricted to the rectal wall (ypT0-2) following neoadjuvant chemoradiation and transanal endoscopic microsurgery. This study considered a prospective cohort of patients with residual rectal cancers following neoadjuvant chemoradiation treated by transanal endoscopic microsurgery and no additional systemic therapy. This study was a single-institution experience. Patients with adenocarcinoma of the rectum located no more than 7 cm from the anal verge and endorectal ultrasound- or magnetic resonance-staged cT2-4N0-2M0 treated by neoadjuvant chemoradiation (50.4-54 Gy and 5-fluorouracil-based chemotherapy) were eligible for the study. Patients with small residual tumors (≤3 cm) radiologically staged ycT0-2N0 were treated by transanal endoscopic microsurgery. Transanal endoscopic microsurgery was performed. The primary outcome measured was local recurrence. Of the 27 patients treated by transanal endoscopic microsurgery, 3 had ypT0, 6 had ypT1, and 18 had ypT2 cancers. All patients underwent R0 transanal endoscopic microsurgery excision. Local recurrence was observed in 4 (15%) patients after a median follow-up of 15 months. Only lymphovascular invasion was an independent predictive factor for local failure (p = 0.04). Tumor size, ypT status, T-status downstaging, lateral/radial margins, and tumor regression grade were not predictors of local failure. This study was limited by the small sample size and limited follow-up. A local failure rate of 15% after transanal endoscopic microsurgery for patients with residual rectal cancers restricted to the bowel wall (ypT0-2) may limit the indication of this procedure to highly selected patients as an alternative to standard radical total mesorectal excision.

Influence of Neoadjuvant Chemotherapy on HER2/neu Status in Invasive Breast Cancer

IntroductionReliably estimating HER2/neu expression in breast cancer is important for predicting patient prognosis and optimizing adjuvant therapeutic strategies. In this retrospective cohort study, effects of NAC on HER2/neu status in invasive breast cancer were evaluated, and the related factors were analyzed. Patients and MethodsOne hundred thirty-one patients with primary breast cancer were treated with anthracycline- and/or taxane-based NAC. HER2/neu status was evaluated by IHC on core needle biopsies of primary tumors before NAC and surgical resection specimens of post-NAC residual breast cancers or tumor-positive axillary lymph nodes. Thirty-two pairs of specimens with discordant HER2/neu IHC scores were analyzed by fluorescence in situ hybridization (FISH). ResultsA significant difference in HER2/neu status by IHC between core needle biopsies and surgical resection specimens in patients receiving NAC was observed. After NAC, 23.4% (29 of 124) of tumors showed downregulated HER2/neu expression by IHC. Alterations of HER2/neu IHC scores did not significantly correlate with tumor subtype, pathologic response to NAC, adjuvant regimen, or time interval from the last chemotherapy to surgery. HER2/neu protein overexpression level was associated with favorable pathologic response to anthracycline and taxane-based chemotherapy. However, tumors with altered HER2/neu IHC scores after NAC revealed stable HER2/neu gene amplification/nonamplification by FISH analysis. ConclusionNeoadjuvant chemotherapy for breast carcinoma resulted in the HER2/neu status alteration by IHC, but they have stable gene amplification status by FISH. HER2/neu protein overexpression indicated greater sensitivity to neoadjuvant anthracycline- and taxane-based chemotherapy. Thus, retesting HER2/neu IHC status in residual tumors after NAC should be considered in order to optimize adjuvant systemic therapy.

Profiling of residual breast cancers after neoadjuvant chemotherapy identifies DUSP4 deficiency as a mechanism of drug resistance

Neoadjuvant chemotherapy (NAC) induces a pathological complete response (pCR) in ~30% of patients with breast cancer. However, many patients have residual cancer after chemotherapy, which correlates with a higher risk of metastatic recurrence and poorer outcome than those who achieve a pCR. We hypothesized that molecular profiling of tumors after NAC would identify genes associated with drug resistance. Digital transcript counting was used to profile surgically resected breast cancers after NAC. Low concentrations of dual specificity protein phosphatase 4 (DUSP4), an ERK phosphatase, correlated with high post-NAC tumor cell proliferation and with basal-like breast cancer (BLBC) status. BLBC had higher DUSP4 promoter methylation and gene expression patterns of Ras-ERK pathway activation relative to other breast cancer subtypes. DUSP4 overexpression increased chemotherapy-induced apoptosis, whereas DUSP4 depletion dampened the response to chemotherapy. Reduced DUSP4 expression in primary tumors after NAC was associated with treatment-refractory high Ki-67 scores and shorter recurrence-free survival. Finally, inhibition of mitogen-activated protein kinase kinase (MEK) synergized with docetaxel treatment in BLBC xenografts. Thus, DUSP4 downregulation activates the Ras-ERK pathway in BLBC, resulting in an attenuated response to anti-cancer chemotherapy.

Abstract 162: Biomarker identification of residual triple negative breast cancer (TNBC) after standard neoadjuvant chemotherapy (NCT)

Abstract Backgrounds Residual TNBC after standard NCT has a poor prognosis. The aim of this study was to identify biomarkers that correlate with prognosis and can be used as novel therapeutic targets in this disease Methods 54 residual TNBC were identified. Protein was extracted and 76 antibiotics to proteins related to breast cancer signaling were measured by reverse phase protein arrays (RPPA). Univariable and multivariable Cox proportional hazard model ratios were fitted for each protein and simultaneously. Cross-validation was also performed. The coefficient in the multivariable Cox model were used to calculate a risk score (RS) for each sample. Results The unsupervised clustering analysis of the proteomic profiles for the 54 residual breast cancers and 76 proteins divided the tumors into two groups without significant survival difference. Multivariable analysis using the top 25 proteins from univariable analysis at a false discovery rate (FDR) of 0.3 showed that IGFPB2, LKB1, and S6 were significant predictors in relapse free survival (RFS). RS model calculated based on multivariable analysis was 0.3746*IGFPB2-0.8852*LKB1+0.6012*S6, AUC=0.807. Employing crossvalidation the model was found to be reproducible. Multivariable analysis suggested that RS was an independent predictor of RFS (HR: 2.705, 95% CI 1.549 to 4.723, P-value=0.0005). Conclusions We identified a RS of three biomarkers that predict outcome in patients with residual TNBC that could be studied as therapeutic targets for this disease. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 162. doi:1538-7445.AM2012-162

Tu1736 Temporal Changes in Colorectal Cancer Screening Practices in the Department of Veterans Affairs Between 2004 and 2010

diameter ( SD) of the EGCs was 26.0 9.5mm and 28 (70%) lesions had the diameter of more than 20mm. The immediately secondary treatments with argon plasma coagulation (APC) or additional ESD (AESD) were performed in 14 (35%) and operations were done in 4 (10%) just after 1st ESD. The residual cancers were found in 2 surgical specimens of 4 patients who underwent operation. No lymph node metastasis was noted in these patients. There was no the local and distant recurrence for mean follow-up period (40.0 months 25.2, median SE) in 18 patients with immediately secondary treatment. Twenty-two (55%) cases without secondary treatment were followed up with endoscopic surveillance. Among 22 patients, 3 (13.6%) had the local recurrence during follow-up endoscopic surveillance (30.0 months 19.0, median SE). The EGCs with local recurrence were treated by Re-ESD (2 cases) and operation (1 case). Conclusions: Immediately secondary treatment was effective for the EGC patients who were regarded as laterally incomplete resection in pathologic report after ESD. The endoscopic surveillance should be performed meticulously in the patients without immediately secondary treatment. Endoscopic procedure can be firstly considered as less invasive treatment in selected patients with incompletely resected and locally recurred EGCs after ESD.

Gene Expression, Molecular Class Changes, and Pathway Analysis after Neoadjuvant Systemic Therapy for Breast Cancer

To examine gene expression differences between pre- and post-neoadjuvant systemic therapy (NST) specimens of breast cancers and identify biologic changers that may lead to new therapeutic insights. Gene expression data from prechemotherapy fine needle aspiration specimens were compared with resected residual cancers in 21 patients after 4 to 6 months of NST. We removed stroma-associated genes to minimize confounding effects. PAM50 was used to assign molecular class. Paired t test and gene set analysis were used to identify differentially expressed genes and pathways. The ER and HER2 status based on mRNA expression remained stable in all but two cases, and there were no changes in proliferation metrics (Ki67 and proliferating cell nuclear antigen expression). Molecular class changed in 8 cases (33.3%), usually to normal-like class, which was associated with low residual cancer cell cellularity. The expression of 200 to 600 probe sets changed between baseline and post-NST samples. In basal-like cancers, pathways driven by increased expression of phosphoinositide 3-kinase, small G proteins, and calmodulin-dependent protein kinase II and energy metabolism were enriched, whereas immune cell-derived and the sonic hedgehog pathways were depleted in residual cancer. In non-basal-like breast cancers, notch signaling and energy metabolism (e.g., fatty acid synthesis) were enriched and sonic hedgehog signaling and immune-related pathways were depleted in residual cancer. There was no increase in epithelial-mesenchymal transition or cancer stem cell signatures. Our data indicate that energy metabolism related processes are upregulated and immune-related signals are depleted in residual cancers. Targeting these biologic processes may represent promising adjuvant treatment strategies for patients with residual cancer.

P1-03-02: The Norton-Simon Hypothesis and Cancer Stem Cells: How Cancer Stem Cells May Explain the Effectiveness of Dose-Dense Chemotherapy.

Abstract Background: Systemic therapies are effective initially in controlling and reversing tumor growth; however, residual cancers will invariably re-grow despite this initial response. We have published data from paired human breast cancer samples that standard therapy every three weeks kills dividing daughter cells but not tumor-initiating cells (TICs), so that samples obtained after therapy are enriched for CD44+/CD24−/low putative “tumor-initiating” or “cancer stem” cells, indicating that standard treatment regimens are missing the critical targets, TICs. Interestingly, we have recent data in human breast tumors that indicate that TICs may in fact be chemosensitive initially, with a decrease in TICs observed within two days of chemotherapy, but shortly thereafter, TICS are actually induced by chemotherapy. Materials and Methods: To evaluate TICs response to chemotherapy, mice with human breast tumor xenograft lines BCM-2665a and BCM-2147 were treated with vehicle, 10-, or 33-mg/kg docetaxel, and then tumors were collected for TIC assays and molecular analysis at both 48 and 72h after treatment. Using Affymetrix gene expression microarrays and reverse phase protein array (RPPA) analysis with 119 different validated antibodies, we identified pathways involved in regulation of TICs. Results: According to flow cytometric analysis for TIC markers and mammosphere (MS) formation efficiency, BCM-2665 TICs were reduced by docetaxel treatment compared to vehicle-treated at 48h post 10 mg/kg docetaxel (4-fold decrease) and 48 and 72h post 33 mg/kg docetaxel compared to control (14- and 2-fold decrease, respectively). Although the BCM-2147 TICs did not significantly decrease at any time points or doses tested, they were clearly induced at 72h post-treatment compared to control. Additionally, BCM-2665 TICs were increased within 72h post 10 mg/kg docetaxel, indicating that these time points are ideal for defining the mechanisms responsible for induction of TICs. Ingenuity Pathway Analysis of Affymetrix microarray data for both BCM-2665 and BCM-2147 revealed induction of inflammatory pathways, suggesting leukocyte infiltration associated with induction of TICs. Furthermore, RPPA analysis confirmed gene expression changes from the microarray data, and implicated apoptosis and inflammatory pathways. Sixteen of 28 proteins significantly changed with activation of CSC are involved in development of leukocytes. Discussion: These findings are consistent with the Norton-Simon Hypothesis in that chemotherapy regimens given more frequently may in fact eliminate TICs, thereby explaining the proven increased effectiveness of dose-dense chemotherapy. Based on when TICs became chemoresistant, we are comparing dose-dense treatment (4 mg/kg docetaxel every 3 days) to a traditional single dose of 32 mg/kg, in an effort to eliminate the tumor cells that cause tumor recurrence. Furthermore, our analysis of gene expression at both the RNA and protein level implicated the immune cells as TICs inducers. Since our immunocompromised mice lack T- and B- cells but have active macrophages, macrophages are indicated as inducers of TICs. We are focusing our current efforts at identifying how immune cells activate TICs and thus enhance tumorigenesis. Citation Information: Cancer Res 2011;71(24 Suppl):Abstract nr P1-03-02.

P3-17-02: Targeting the Autophagy Pathway for Drug Resistance of Breast Tumor-Initiating Cells.

Abstract Background: Our clinical data and experimental evidence reveal that the tumor-initiating cells (TICs) in the original tumor are intrinsically resistant to conventional chemotherapy and radiation therapy and greatly enriched in residual breast cancers after such treatments. We have published a gene expression signature of such breast TICs, and our pathway analysis on the gene signature suggests that the activation of autophagy pathway may be an intrinsic characteristic of the TICs. This motivates us to further investigate the role of the autophagy pathway in TICs self-maintenance and their resistance to hormonal and chemo therapies, as well as their response to TIC-targeted therapies. Methods: A collective 84 well-documented autophagy pathway genes were used to compare the activation of autophagy pathway in different microarray datasets, 1) flow-sorted CD44+/CD24−/low cancer cells vs. all other cells (representing 20 patients), and cancer-derived MSs vs. corresponding primary bulk tumors (representing 17 patients), 2) before vs. after letrozole and doxetaxel treatments (representing 30 patient pairs), 3) before vs. after letrozole treatment (representing 176 patient pairs), and 4) before vs. after lapatinib treatment (representing 115 patient pairs). We applied the Significance Analysis of Microarrays (SAM) algorithm to analyze the expression data of 211 normalized probes for the 84 genes. Low-density RTPCR array for the 84 genes was used to confirm the differential expressed genes on tumor tissues of 18 letrozol treated patient pairs. Two human-cancer-in-mouse triple-negative xenograft tumor lines were treated with Notch pathway inhibitor alone or in combining with docetaxel. The tumor growth, mammosphere formation efficiency, and the expression of autophagy marker proteins were evaluated. Results: Overall, the enrichment analysis of the 84-gene set in all the above microarray datasets showed that the autophagy genes are significantly enriched in the differential expressed genes. In particular, 28 out of the 84 autophagy genes are significantly up-regulated in the TICs populations while 5 other autophagy genes are down-regulated with the false discovery rate (FDR) less than 0.05. For the letrozole and doxetaxel treatment microarray analysis, 34 out of the 84 autophagy genes are significantly up-regulated in the after-treatment group (p<0.01), and only 3 out of the 84 autophagy genes are down-regulated (FDR<0.05). Interestingly, significantly more autophagy genes (15 out of the 84) are down-regulated in the after-lapatinib treatment group (p<0.05), and less genes (13 out of 84) are up-regulated after lapatinib treatment, thus confirming previous observations that lapatinib may affect TICs. We confirmed these findings in two human cancer xenograft studies of Notch pathway inhibitor (GSI) to target TICs, which showed significantly reduced expressions of LC3 and p62, two autophagic markers. In both xenograft models, GSI reduced mammosphere formation efficiency compared to vehicle control (p<0.05). Conclusion: Activation of the autophagy pathway in TICs is a promising target to combat the drug resistance of breast cancer to conventional systemic therapy. Citation Information: Cancer Res 2011;71(24 Suppl):Abstract nr P3-17-02.

'Omic approaches to preventing or managing metastatic breast cancer

Early detection of metastasis-prone breast cancers and characterization of residual metastatic cancers are important in efforts to improve management of breast cancer. Applications of genome-scale molecular analysis technologies are making these complementary approaches possible by revealing molecular features uniquely associated with metastatic disease. Assays that reveal these molecular features will facilitate development of anatomic, histological and blood-based strategies that may enable detection prior to metastatic spread. Knowledge of these features also will guide development of therapeutic strategies that can be applied when metastatic disease burden is low, thereby increasing the probability of a curative response.

P59. Role of PET CT in management of recurrent or residual head and neck cancers

P59. Role of PET CT in management of recurrent or residual head and neck cancers

Evaluation of the role of second TUR for transurethral resection of bladder tumor in one piece (TURBO).

e15009 Background: Since transurethral resection of bladder tumor in one piece (TURBO) was reported in the Journal of Urology in 2000 by Ukai, some urologists carried out TURBO. We analyzed treatment results of TURBO in our hospital and examined the value of this procedure, especially for the pathological findings, recurrence and necessity of second TUR for TURBO. Methods: A total of 14 patients with bladder tumors carried out TURBO under spinal anesthesia, in some cases blocking the obtulater nerve, from April 2006 to June 2009 in our hospital. All cases were followed for over 1 year. The procedure is 1. point marking; 2. circular incision; 3. level incision; and 4. specimen retrieval using a needle electrode in accordance with the Ukai's method. We investigated pathological findings (margin situation), operation time, complications and recurrence. Results: It is possible to diagnose the precise pathlogical findings by TURBO. We judged the width and depth ew in sequential section. There were no complications during and after the operation. Operation time of TURBO (35-170 min) was longer than TUR-BT. Urethral catheter holding period and hospitalization period after TURBO was the same as TUR-BT. TURBO is a relatively safe procedure even for beginners. 5 cases had a recurrence in 13 cases. 2 cases had a recurrence in under 1 year, but the locations were other places. One case had a same place recurrence after 13 months. There were no cases of same place recurrence in under 1 year among margin-negative cases. Therfore we judged that ew-negative cases had no residual cancers. Conclusions: TURBO is a safe and useful procedure that provides precise pathological findings with minimal complications. Second TUR is not necessary for TURBO. TURBO has a possibility to be gold standard of the treatment for non-muscle invasive bladder cancer.

Use of gene expression patterns post neoadjuvant chemotherapy to identify a role for the MAPK phosphatase DUSP4 in therapeutic resistance and a stem-like phenotype in basal-like breast cancer (BLBC).

10509 Background: Neoadjuvant chemotherapy (NAC) leads to a pathological complete response in ~20% of patients with breast cancer. The remaining patients have residual disease and are more likely to develop metastasis. In post-NAC tumors, Ki67 levels, a marker of cell proliferation, have been shown to inform odds of recurrence and death. We hypothesized that post-NAC residual cancers with a high Ki67 harbor molecular alterations that are associated with drug resistance. Methods: We utilized Nanostring digital RNA quantification to measure the expression levels of 350 cancer-related genes in 49 post-NAC breast cancers. Gene expression data were tested for associations with post-NAC Ki67. The cohort was enriched for triple-negative breast cancer (24/49 tumors). The candidate gene DUSP4 was explored in in vitro models of breast cancer. Results: The post-NAC Ki67 (range 2.4 - 99%; median: 31.6%) was statistically different among the molecular subtypes of breast cancers enrolled, being highest in BLBC (p<0.0001). Gene expression signatures of KRAS activation were enriched in BLBC. Low levels of the MAPK phosphatase DUSP4 correlated with a high Ki67 in BLBCs (r=-0.54, p=8x10-4) and with Ras/MAPK pathway activation in human breast tumors and cell lines. Genome-wide studies identified preferential methylation of the DUSP4 promoter in BLBC. Among BLBC cell lines, low DUSP4 expression predicted for sensitivity to MEK inhibitors and resistance to docetaxel. Inhibition of MEK with AZD6244 or overexpression of DUSP4 in DUSP4-low BLBC cell lines improved sensitivity to docetaxel and inhibited mammosphere formation. RNAi-mediated knockdown of DUSP4 in BLBC cells enhanced ETS-1 phosphorylation, mammosphere formation, and MAPK-dependent secretion of the cancer stem cell-promoting cytokines IL6 and IL8. Conclusions: Low levels of DUSP4 correlate with high tumor cell proliferation after NAC. Downregulation of DUSP4 may underlie activation of the MAPK pathway in the absence of KRAS mutations in BLBC while conferring resistance to chemotherapy and a stem cell-like phenotype that is therapeutically targetable with MAPK inhibitors.

Lung Abnormalities at Multimodality Imaging after Radiation Therapy for Non–Small Cell Lung Cancer

Three-dimensional (3D) conformal radiation therapy (CRT) and stereotactic body radiation therapy (SBRT) are designed to deliver the maximum therapeutic radiation dose to the tumor, allowing improved local disease control, while minimizing irradiation of surrounding normal structures. The complex configuration of the multiple beams that deliver the radiation dose to the tumor in 3D CRT and SBRT produces patterns of lung injury that differ in location and extent from those seen after conventional radiation therapy. Radiation-induced changes in lung tissue after 3D CRT and SBRT occur within the radiation portals. The imaging appearance of irradiated tissues varies according to the time elapsed after the completion of therapy, with acute-phase changes of radiation pneumonitis represented by ground-glass opacities and consolidation and with late-phase changes of radiation fibrosis manifesting as volume loss, consolidation, and traction bronchiectasis. Knowledge of treatment timelines and radiation field locations, as well as familiarity with the full spectrum of possible radiation-induced lung injuries after 3D CRT and SBRT, is important to correctly interpret the abnormalities that may be seen at computed tomography (CT). Differential diagnoses in this context might include infections, lymphangitic carcinomatosis, local recurrence of malignancy, and radiation-induced tumors. The integration of morphologic information obtained at CT with metabolic information obtained at positron emission tomography is helpful in distinguishing radiation-induced parenchymal abnormalities from residual, recurrent, and new cancers. Thus, multimodality follow-up imaging may lead to substantial changes in disease management.

Abstract LB-100: The Norton-Simon hypothesis and cancer stem cells: how cancer stem cells may explain the effectiveness of dose-dense chemotherapy

Abstract Systemic therapies are effective initially in controlling and reversing tumor growth; however, residual cancers will invariably re-grow despite this initial response. We have published data from paired human breast cancer samples that standard therapy every three weeks kills dividing daughter cells but not tumor-initiating cells (TICs), so that samples obtained after therapy are enriched for CD44+/CD24−/low putative “tumor-initiating” or “cancer stem” cells. These data indicate that standard treatment regimens are missing the critical targets, TICS; thus, we are trying to identify novel therapeutic combinations and regimens that will eliminate these TICS that ultimately cause breast tumor recurrence and metastasis. Interestingly, we have recent data in human breast tumors that indicate that TICs may in fact be initially chemosensitive, with a decrease in TICs observed within three days of chemotherapy, but shortly thereafter, TICS are actually induced by chemotherapy. To evaluate TIC response to chemotherapy, mice with xenografted human breast tumors were treated with docetaxel (20 mg/kg) or vehicle, and then tumors were collected and dissociated for TICs assays either 3d or 14d after treatment. According to multiple assays for TICs, TICs were reduced compared to vehicle-treated at 3d post-docetaxel, and increased at 14d. This was evident in both primary and secondary mammosphere (MS) formation. Flow cytometric analyses for TICs markers (CD44+/CD24−/low and ALDH+) further corroborated these data. These results indicate TICs were initially responsive but became chemoresistant, or even activated by chemotherapy. To further define TICs response to chemotherapy, mice with xenografted human breast tumors were treated with vehicle, 10 mg/kg docetaxel, or 33 mg/kg docetaxel, and then tumors were collected and dissociated for TICs assays either 48h or 72h after treatment. Although BCM-2147 TICs did not significantly decrease at any time points or doses tested, they were induced by 72h post-treatment compared to control (ALDH+ 2- to 4-fold increased). In contrast, according to TIC markers and MS formation efficiency, BCM-2665 TICS were reduced by docetaxel compared to vehicle at 48h post 10 mg/kg docetaxel (4-fold decrease), and 48h and 72h post 33 mg/kg docetaxel compared to control (14- and 2-fold decrease, respectively). Furthermore, BCM-2665 ALDH+ cells showed 6-fold increased proliferation 72h post cisplatin compared to control, indicating that chemotherapy may inducing proliferation of TICs. When TICs are induced to proliferate, they may be more susceptible to traditional proliferative targets such as chemotherapy. These findings are consistent with the Norton-Simon Hypothesis in that chemotherapy regimens given more frequently may in fact eliminate TICs, thereby explaining the proven increased effectiveness of dose-dense chemotherapy. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr LB-100. doi:10.1158/1538-7445.AM2011-LB-100

Specific kinesin expression profiles associated with taxane resistance in basal-like breast cancer

Breast cancer is a genetically heterogenous disease with subtypes differing in prognosis and chemosensitivity. The basal-like breast cancer (BLBC) molecular subtype is associated with poorer outcomes, but is more responsive to taxane-based chemotherapy. Kinesins are intracellular transport proteins that interact with microtubules, which are also the mechanistic target for taxanes. We investigated the relationship between taxane resistance in BLBC and kinesins using both expression and functional studies. Kinesin (KIF) expression was evaluated in three settings in relation to taxane resistance: (i) the NCI-60 cancer cell lines, (ii) pre-treatment samples from four BLBC patient cohorts receiving neoadjuvant chemotherapy regimens with and without taxanes, and (iii) post-treatment samples from residual breast cancer following neoadjuvant taxane-containing chemotherapy. We used a novel functional approach to gene modification, validation-based insertional mutagenesis, to select kinesin-overexpressing clones of BLBC cells for evaluation of related mechanisms of taxane resistance. In the NCI-60 cell line dataset, overexpression of the kinesin KIFC3 is significantly correlated with resistance to both docetaxel (P<0.001) and paclitaxel (P<0.001), but not to platinum-based chemotherapy, including carboplatin (P=0.49) and cisplatin (P=0.10). Overexpression of KIFC3 and KIF5A in pre-chemotherapy samples similarly predicted resistance to paclitaxel in the MDACC cohorts (P=0.01); no KIF predicted resistance to fluorouracil-epirubicin-cyclophosphamide or cisplatin in BLBC patient cohorts treated without taxanes. KIF12 is the most overexpressed KIF gene in post-chemotherapy taxane-resistant residual breast cancers (2.8-fold-change). Functional studies established that overexpression of KIFC3, KIF5A, and KIF12 were specific in mediating resistance to docetaxel and not vincristine or doxorubicin. Mutation of the ATP-binding domain of a kinesin abolished its ability to mediate docetaxel resistance. Overall, kinesin overexpression correlates with specific taxane resistance in BLBC cell lines and tissues. Our results suggest a novel approach for drug development to overcome taxane resistance in breast cancer through concurrent or sequential use of kinesin inhibitors.