Abstract Proteogenomics is the field of integrating data from mass spectrometry-based shotgun proteomics, and phosphoproteomics into next-generation RNA and DNA sequencing data analysis pipelines that promises new insights into cancer biology and therapeutic targeting. As well as analyses of clinical samples for disease phenotype association analysis, the application of proteogenomics to model systems also has considerable potential. A Clinical Proteomic Tumor Analysis Consortium (CPTAC) proteogenomic analysis prioritized dihydropyrimidinase-like-3 (DPYSL3) as a multi-level (RNA/Protein/Phosphoprotein) expression outlier specific to the Claudin-Low (CLOW) subset of triple negative breast cancers. A Pubmed informatics tool indicated a paucity of data in the context of breast cancer which further prioritized DPYSL3 for study. DPYSL3 was identified as a protein that is regulated during neuronal differentiation in the cerebral cortex and in neuronal cell lines and plays a role in regulating neurite outgrowth somehow through an association with vesicles in the growth cone. In addition, DPYSL3 expression has been observed in several malignant tumors, including prostate cancer, pancreatic cancer, gastric cancer and neuroblastoma. DPYSL3 is reported to play a role in cell migration and metastasis suppression in prostate cancer. However, in pancreatic cancer, DPYSL3 is positively associated with liver metastasis and poor outcome. DPYSL3 knock-down in DPYSL3 (+) CLOW cell lines demonstrated reduced proliferation, yet enhanced motility and increased expression of Epithelial to Mesenchymal Transition (EMT) markers suggesting that DPYSL3 is a multi-functional signaling modulator. Slower proliferation in DPYSL3 (-) CLOW cells was associated with accumulation of multi-nucleated cells indicating a mitotic defect that was associated with a collapse of the vimentin (VIM) microfilament networkinduced by VIM hyperphosphorylation. On the other hand, DPYSL3 suppressed the expression of EMT regulators TWIST and SNAIL and opposed p21 activated kinase 2 (PAK2) dependent migration, but these EMT regulators in turn induced DPYSL3 expression, suggesting DPYSL3 participates in negative feedback in EMT. Cell migration in DPYSL3 (-) cells correlated with increased phosphorylation of PAK2 on Ser20 and was sensitive to PAK2 siRNA and pharmacological PAK inhibition.Immunoprecipitation and mass spectrometry-based proteomics or western blotting strongly suggests that PAKs interact such that DPYSL3 may function as a direct negative regulator of PAK family kinases. Thus, a PAK inhibitor could potentially mitigate increase migration as an adverse effect of DPYSL3 suppression. In conclusion, DPYSL3 is a remarkable multifunctional signaling scaffold that should be examined further to provide insights into the stem cell-like state of claudin-low breast cancers, particularly in terms of their cell cycle dependencies, migratory activity and capacity for EMT. Citation Format: Matsunuma R, Chan DW, Kim B-J, Singh P, Han A, Saltzman A, Cheng C, Lei JT, Sahin E, Leng M, Fan C, Perou CM, Malovannaya A, Ellis MJ. DPYSL3 modulates mitosis, migration and epithelial to mesenchymal transition in claudin-low breast cancer [abstract]. In: Proceedings of the 2018 San Antonio Breast Cancer Symposium; 2018 Dec 4-8; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2019;79(4 Suppl):Abstract nr P5-08-01.

Abstract Cancer is often driven by deregulated protein kinases pathways including breast cancer. To profile the kinome at the protein level we are developing a kinase pulldown (KiP) proteomic platform that uses multiple kinase inhibitors conjugated to sepharose beads for protein kinase profiling. Mass spectrometry-base tryptic peptide sequencing is used to identify and quantify the kinome in an unbiased manner. To date, we have profiled and analyzed the kinomes of more than 50 breast cancer PDX model systems that include all five breast cancer subtypes. In these analyses we have begun to explore the kinome interactome because we have observed that KiP also pulls down breast cancer subtype specific kinase interacting proteins that may represent new biological insights. For example we have consistently identified dihydropyrimidinase-like 3 (DPYSL3) in p21-activated kinase (PAK) inhibitor KiP interactome in the claudin-low WHIM12 PDX model. DPYSL3 is an intracellular phosphoprotein known to play a role in cell migration and metastasis. However it remains to be seen if DPYSL3 functions as a suppressor or a promoter of metastasis. We therefore generated DPYSL3 knock-down cell lines to evaluate function in claudin-low breast cancer. Proliferation levels of DPYSL3 depleted Claudin-low WHIM12 cells (DPYSL3-) were lower than those of control WHIM 12 (DPYSL3+) cells. In contrast, migration levels of DPYSL3- WHIM12 were greater than those of control cells, suggesting a pivotal role for DPYSL3 in several cellular physiologies. Additionally the activity of a PAK inhibitor (FRAX597) on migration appeared to require DPYSL3, as the inhibitory effect of FRAX579 on migration in WHIM12 was lost when DPYSL3 expression was depleted. Additionally we find that only the long isoform of DPYSL3, not a short isoform, interacts with PAK to regulate migration, suggesting that alternative splicing of DPYSL3 may an important to the control of migration. Furthermore Snail is also negatively regulated DPYSL3 with elevated expression in DYPSL3- cells. In conclusion, the study of the kinome interactome in breast cancer PDX identified DPYSL3 as regulator of the PAK family of kinases in a claudin low breast cancer. DPYSL3 is necessary for PAK to promote migration and may play a pivotal role in complex cellular transitions from a proliferative state versus an EMT/cell migratory state. Citation Format: Matsunuma R, Chan DW, Kim B-J, Singh P, Han A, Malovannaya A, Perou CM, Ellis MJ. DPYSL3 modulates proliferation and migration in claudin-low breast cancer [abstract]. In: Proceedings of the 2017 San Antonio Breast Cancer Symposium; 2017 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2018;78(4 Suppl):Abstract nr P1-01-08.

(Background) Patient-derived xenograft (PDX)s are valuable models for precision oncology as they are thought to recapitulate the biological and genomic characteristics of the human tumors they were derived from. Even though this model is widely used, it is still difficult to manipulate genes in this system and it is not as convenient as in vitro cell culture for drug sensitivity screening. On the other hand in vitro cell cultures have a highly artificial microenvironment and also have undergone selection which might generate misleading data. To address these issues cell lines from PDX tumors with different intrinsic subtypes were characterized by kinobead precipitation/mass spectrometry analysis (KiP/MS) to profile each PDX line in vivo and ex vivo for therapeutic targets. (Materials and Methods)Washington University Human in Mice (WHIM) tumors and Hutsman Cancer Institute (HCI) tumors were transplanted into mammary fat pads of female severe combined immunodeficiency/beige (SCID/beige) mice. PDXs tumors were harvested when they reached 1˜1.5cm. PDX tumor-derived ex vivo cells/organoids were routinely cultured with Rock inhibitor support. Estradiol was applied only for the tumors originating from E2 supplementation in vivo. Cells and tumors were harvested and lysed by sonication. Kinases in soluble lysates are enriched with drug-bound beads (kinobead) and digested with trypsin. Digested peptides are analyzed by mass spectrometry. (Results) 5 of WHIM tumors and 4 of HCI tumors were successfully dissociated and cultured ex vivo for further analysis and experiments. Hierarchical clustering of tumors and corresponding cells showed ex vivo cultured cells cluster together with their original PDX tumors. Tumors/ex vivo cultured cells clustered by intrinsic subtype and enrichment analysis identified specific kinases for each PDX tumor/cell line. The WHIM 4 tumor-cell pair showed high level of PIM kinase and EGFR and other PDX tumors such as WHIM18, WHIM 20 (Luminal subtypes), WHIM 35(HER2 enriched subtype) also showed model unique intrinsic kinases, such as JAK2 for WHIM18, EPHB4 for WHIM20. (Conclusion) PDX tumor-derived ex vivo lines could be routinely cultured with Rock inhibitor support. Each PDX tumor and cell line pair were cluster together in hierarchical clustering and categorized into the same intrinsic subtype based on kinome profiling, suggesting these cells maintain their tumor specific intrinsic kinase signaling. A subset of kinases exhibit activity/expression that is conserved after ex vivo culture, we hypothesize these are intrinsic kinases might be promising target for treatment, because they are tumor intrinsic, i.e. their high expression is maintained despite the strong contrast in the microenvironment of in vitro versus in vitro growth. Ongoing studies with drugs and knock down reagents are examining whether the in vivo/ex vivo comparative KIP analysis indeed identifies therapeutic targets, which will be presented at the meeting more in detail. Citation Format: Han A, Kim B-J, Chan DW, Matsunuma R, Singh P, Ellis MJ. Proteomic analysis of conserved kinases between PDX tumors and corresponding PDX-derived cell lines [abstract]. In: Proceedings of the 2017 San Antonio Breast Cancer Symposium; 2017 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2018;78(4 Suppl):Abstract nr P5-06-01.

Abstract Background. Dysregulation of the estrogen receptor gene (ESR1) is an established mechanism of inducing endocrine therapy resistance. We previously discovered a chromosomal translocation event generating an estrogen receptor gene fused in-frame to C-terminal sequences of YAP1 (ESR1-YAP1) that contributed to endocrine therapy resistance in estrogen receptor positive (ER+) breast cancer models. This study compares functional, transcriptional, and pharmacological properties of additional ESR1 gene fusion events of both early stage (ESR1-NOP2) late stage (ESR1-YAP1 and ESR1-PCDH11x) breast cancers to gain a better understanding of therapeutic resistance and metastasis. Understanding the role of ESR1 fusions in inducing metastasis is critical, since the primary cause of death in breast cancer patients is through metastasis to distant sites. Methods. RNA-seq screens identified ESR1 fusions from early and late stage, endocrine therapy resistant breast tumor samples. Functional experiments were conducted using ER+ breast cancer cell lines, xenograft, and PDX models to test the ability of ESR1 fusions to induce therapeutic resistance and metastasis. ChIP-seq and RNA-seq were performed to examine transcriptional properties and differential gene expression induced by the fusions which directed subsequent pharmacological experiments with a CDK4/6 inhibitor. Results. ESR1-YAP1 and ESR1-PCDH11x promoted estrogen-independent and fulvestrant-resistant growth in vitro and induced greater tumor growth and increased metastatic capacity to the lungs of xenografted mice. In contrast, the ESR1-NOP2 fusion was sensitive to low estrogen conditions in vitro, and did not promote tumor growth. RNA-seq profiling revealed E2F targets pathway as the most highly enriched pathway induced by the ESR1 fusions. IHC revealed higher levels of pRb in ESR1-YAP1 and ESR1-PCDH11x xenograft tumors and subsequent CDK4/6 inhibition completely blocked tumor growth in an ESR1-YAP1 PDX model. Integrating RNA-seq with ChIP-seq data, we discovered a set of EMT and metastasis genes bound by all ESR1 fusions and WT-ER, but whose expression was strongly and uniquely up-regulated only by the ESR1-YAP1 and ESR1-PCDH11x fusions. These studies also revealed gained sites bound only by the ESR1-YAP1 and ESR1-PCDH11x fusions, not bound by WT-ER nor ESR1-NOP2. Genes mapping to these sites have a role in metastatic biology and were highly up-regulated by the YAP1 and PCDH11x fusions, potentially mediated by long range transcriptional activation. Conclusion. ESR1-YAP1 and ESR1-PCDH11x are driver fusions that occur in drug-resistant, advanced stage breast cancer and are a new class of recurrent somatic mutation that can cause acquired endocrine therapy resistance, yet can be treated with CDK4/6 inhibition. These driver fusions also confer increased metastatic ability through their ability to drive expression of genes that contribute to EMT and metastasis. In contrast, ESR1-NOP2 did not produce functional protein and appears to be a passenger event. These studies may provide pre-clinical rationale for targeting ESR1 translocated breast tumors, since the presence of an ESR1 driver fusion places a patient in a therapeutic category where none of the currently available endocrine therapies are likely to be effective. Citation Format: Lei JT, Shao J, Zhang J, Iglesia M, Chan DW, Cao J, Anurag M, Singh P, Haricharan S, Kavuri SM, Matsunuma R, Schmidt C, Kosaka Y, Crowder R, Hoog J, Phommaly C, Goncalves R, Ramalho S, Rodrigues-Peres RM, Lai W-C, Hampton O, Rogers A, Tobias E, Parikh P, Davies S, Ma C, Suman V, Hunt K, Watson M, Hoadley KA, Thompson A, Perou CM, Creighton CJ, Maher C, Ellis MJ. ESR1 gene fusions drive endocrine therapy resistance and metastasis in breast cancer [abstract]. In: Proceedings of the 2017 San Antonio Breast Cancer Symposium; 2017 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2018;78(4 Suppl):Abstract nr PD8-03.

Abstract Background. We previously reported an alternative ESR1 somatic gain-of-function chromosomal translocation event in a patient presenting with aggressive, endocrine therapy resistant estrogen receptor (ER) positive disease, producing an in-frame fusion gene consisting of N-terminal ESR1 and the C-terminus of the Hippo pathway coactivator YAP1 (ESR1-YAP1). We recently identified another ESR1 fusion through RNA sequencing (RNA-seq) in advanced stage ER+ disease from a chest wall recurrence in a male patient that was refractory to multiple lines of treatment. Two examples of fusions discovered in primary breast cancer samples include ESR1 fused in-frame to C-terminal sequences from NOP2 (ESR1-NOP2), identified in a resistant cohort from a RNA-seq screen focused on 81 primary breast cancers from aromatase inhibitor clinical trials, and a second ESR1 fusion, fused in-frame to the entire coding sequence of POLH (ESR1-POLH), that was identified from RNA-seq analysis of 728 Cancer Genome Atlas breast samples. This current study extends our previous characterization of ESR1-YAP1 by comparing functional and pharmacological properties of these three additional ESR1 gene fusion events of both early stage and advanced breast cancers. Methods. In vitro and in vivo experiments were conducted to test ESR1 fusions to induce therapeutic resistance, and metastasis. The transcriptional and binding properties of each fusion was also examined. Pharmacological inhibition with Palbociclib, a cyclin-dependent kinase 4/6 inhibitor, was utilized to assess drug sensitivity in ESR1 fusion containing breast cancer cells and in a patient derived xenograft (PDX) model expressing ESR1-YAP1 (WHIM18). Results. The YAP1 and PCDH11x fusions conferred estrogen-independent and fulvestrant-resistant growth. Immunohistochemistry revealed significantly higher numbers of ER+ cells in lungs of mice xenografted with T47D cells expressing the YAP1 and PCDH11x fusions compared to YFP control, NOP2 and POLH fusions. Results from ChIP-seq and microarray studies suggest that these two fusions promote proliferation and metastasis through genomic action by binding estrogen response elements (ERE) and subsequent gene activation. We thereby define these fusions as “canonical” fusions compared to “non-canonical” NOP2 and POLH fusions, which demonstrated dramatically decreased genomic binding ability. The non-canonical fusions induced genes associated with basal-like breast cancer and promoted HER2, EGFR, and MAPK gene expression signatures in contrast to genes associated with cell cycle/proliferation induced by canonical fusions. The proliferative ability of canonical fusion-containing ER+ cells was inhibited by Palbociclib in a dose-dependent manner. In vivo WHIM18 tumors in mice fed with Palbociclib-containing chow demonstrated significantly reduced tumor volume, growth rate, and weight compared to tumors in mice on control chow. Conclusions. In-frame ERE activating canonical fusions occur in end-stage drug resistant advanced breast cancer and can be added to ESR1 point mutations as a class of recurrent somatic mutation that may cause acquired resistance. Growth induced by these fusions can be antagonized by Palbociclib and is potentially clinically helpful. Citation Format: Lei JT, Shao J, Zhang J, Iglesia M, Cao J, Chan DW, He X, Kosaka Y, Schmidt C, Matsunuma R, Haricharan S, Crowder R, Hoog J, Phommaly C, Goncalves R, Ramalho S, Lai W-C, Hampton O, Rogers A, Tobias E, Parikh P, Davies S, Ma C, Suman V, Hunt K, Watson M, Hoadley KA, Thompson A, Chen X, Perou CM, Creighton CJ, Maher C, Ellis MJ. Recurrent functionally diverse in-frame ESR1 gene fusions drive endocrine resistance in breast cancer [abstract]. In: Proceedings of the 2016 San Antonio Breast Cancer Symposium; 2016 Dec 6-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2017;77(4 Suppl):Abstract nr PD2-03.

Background: Diffuse optical spectroscopic imaging (DOSI) can be exploited as a marker of tumor blood volume quantified by tissue hemoglobin (tHb) concentration. In DOSI, frequent measurement is possible for breast cancer patients because of its non-invasiveness. The tHb concentration determined by DOSI is expected to be a new biomarker for prediction of breast cancer response to neoadjuvant chemotherapy (NAC). Purpose: Our objective is to determine whether early change of tumor tHb concentration predicts pathological complete response (pCR) to NAC in patients with operable breast cancer. Methods: In a prospective study, one hundred patients with primary breast cancer were enrolled for primary objective analysis. The regimens of NAC were according to the standard of care. Patients underwent sequential scans using DOSI at baseline, after 1st course and 2nd course of chemotherapy. The mean value of tHb (tHbmean) concentration of the targeted lesion was measured and the percentage change in tHbmean (ΔtHbmean) concentration was calculated. Receiver operating curve analysis demonstrated diagnostic performance of DOSI for predicting a pCR. Results: In interim analysis, it was regarded as a good outcome that area under the curve (AUC) for ΔtHbmean after 1nd course was 0.797 (SE 0.104, 95%CI 0.633-0.911), and after 2st course was 0.867 (SE 0.06, 95%CI 0.715-0.956). Conclusion: DOSI could predict accurately a pCR to neoadjuvant chemotherapy in patients with primary breast cancer. Citation Format: Ogura H, Yoshizawa N, Ueda S, Hosokawa Y, Matsunuma R, Tochikubo J, Nasu H, Shigekawa T, Takeuchi H, Osaki A, Saeki T, Yoshimoto K, Ohmae E, Suzuki T, Ueda Y, Yamashita Y, Sakahara H. Near-infrared diffuse optical imaging for early prediction to neoadjuvant chemotherapy in patients with primary breast cancer. [abstract]. In: Proceedings of the Thirty-Eighth Annual CTRC-AACR San Antonio Breast Cancer Symposium: 2015 Dec 8-12; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2016;76(4 Suppl):Abstract nr P4-03-06.

Background: The St.Gallen consensus guideline recommends the sequential administration chemotherapy followed by of endocrine therapy as postoperative therapy for the higher risk ER-positive breast cancer patients based on results of a single study (Albain et al, Lancet 2009). In metastatic settings however, several trials conducted in the 1980's demonstrated that tumor response rates were higher when chemotherapy and tamoxifen were concomitantly administered, than when chemotherapy and tamoxifen administered were given sequentially. In the preoperative settings, pathological complete response (pCR) rate can be used a surrogate marker to predict event-free survival or overall survival in Luminal B(HER2-negative) breast cancer. We therefore designed a prospective randomized safety and efficacy trial in order to test a hypothesis that the concomitant administration of an aromatase inhibitor and chemotherapy improves pathological complete response(pCR) rate than chemotherapy alone in the preoperative setting. Trial design: The trial is a prospective, multi-center, randomized comparison of chemotherapy alone versus concomitant chmo-endocrine therapy evaluating the efficacy in terms of pCR rate and safety in preoperative settings in patients with Luminal B (HER2-negative) breast cancer. 94 patients were to be accrued into this trial. - arm A (control): 12 cycles of weekly paclitaxel(80mg/m2) followed by 4 cycles of every 3-week AC(Doxorubicine 60mg/m2, Cyclophosphamide 600mg/m2). - arm B (experimental): The same chermotherapy as arm A and anastrozole in postmenopausal patients or anastrozole+leuprolerine in premenopansal patients. Eligibility criteria: 1)Female patients with operable and histologically confirmed invasive breast cancer; 2)HER2-negative; 3)Either ER -positive or PgR-positive; 4)Either Ki67-LI> = 14% and NG> = 2 or NG = 3 regardless of Ki67-LI. Endpoints : Primary endpoint is the pCR rate. Secondary endpoints are the clinical response rate(RECIST), the adverse events(CTC-AE ver.4.0), the breast conserving rate and the health related quality of life. Statistical Considerations : The pCR rates in the control arm and the experimental arm are expected to be 10% and 25%, respectively. In order to show the superiority of the experimental arm with an alfa error at 5% and beta error at 20%, calculated number of patients needed were 96. Present Accrual and Target Accrual: As of June 06, 2013, 18 patients were enrolled from 8 institutions. Citation Information: Cancer Res 2013;73(24 Suppl): Abstract nr OT3-2-03.

Abstract Objectives: Optical mammography (O-MMG), which utilizes near-infrared light for the detection of abnormal breast tissue, can be a potentially useful and non-invasive tool for the screening of breast cancer patients. Here we report on the preliminary results of our clinical trial using a time-resolved O-MMG system developed by Hamamatsu Photonics (Hamamatsu, Japan). In our past examination, negative results included cases with mismatch for breast cup because of small breast, lesion outside of cup because of big breast, and bad positioning. So, size and shape of a gantry was improved (3 cup sizes; S, M and L cup). We reported the initial experience of the improved O-MMG systems. Materials and Methods: In principle, the technique involves sequential irradiation of the breast tissue with a pulsed laser with a wavelength of 760, 800 and, 830nm, which is then detected by the variable capacity gantry at multiple sites after passing through the breast tissue. Measurement time has improved to about 7 minutes (a third of the conventional measurement time). The data are analyzed and tomographic images of the absorption coefficient of the breast are then reconstructed based on our tomographic reconstruction algorithm. Between October 2011 and May 2012, fifteen patients with breast cancer and one patient with fibroadenoma participated in the trial. Results: In patients with breast cancer, S cup was used for one patient, M cup for 12 patients and L cup for two patients. Five patients after neo-adjuvant chemotherapy were excluded from the analysis. The lesions were depicted as an area of high hemoglobin concentration in 7 of 10 patients with cancer. One lesion of fibroadenoma was also identified as an area of high hemoglobin concentration. Conclusion: Further examination with our new O-MMG systems will be of interest. Citation Information: Cancer Res 2012;72(24 Suppl):Abstract nr P4-03-05.

Background: Non-HIV Pneumocystis pneumonia (PCP) could occur in immunosupressed patients having malignancy or on immunosuppressive agents. A-DROP system by the Japanese Respiratory Society (JRS), CURB-65 score by British Respiratory Society (BTS) and the Pneumonia Severity Index (PSI) by the Infectious Disease Society of America (IDSA) are widely used in classifying patients with CAP in Japan. Another predictive rule, I-ROAD, is used for hospital associated pneumonia (HAP) in Japan. We previously reported that I-ROAD could evaluate correctly the severity of CAP as well. For the purpose of evaluating how correctly the I-ROAD could reflect the severity of non-HIV PCP, we retrospectively analyzed all the non-HIV PCP patients. Methods: We reviewed all non-HIV PCP patients admitted to our department from 2009 to 2011. A total of 23 non-HIV PCP patients were enrolled in this study. Patients’ characteristics, the severities of pneumonia, outcomes and the accuracy of predictive rules in each guideline were evaluated. Results: A total of 23 non-HIV PCP patients were enrolled in this study. The patients were 13 males and 10 females. Fourteen were cured and 9 were dead. Based on A-DROP, 20 of the 23 (87%) patients were classified as mild or moderate. Sixteen of 20 (80%) developed respiratory failure and 8 of the 16 (50%) died. Based on CURB-65, 21 of 23 patients (91.3%) were evaluated as mild or moderate. Seventeen of the 21 (81%) developed respiratory failure and 9 of the 17 (52.9%) died. Based on PSI, 8 of 23 patients (34.8%) were evaluated as mild or moderate. Four of the 8 (50%) developed respiratory failure and 1 of the 8 (12.5%) died. However, based on I-ROAD, they were classified as 8 mild (group A)/ 4 moderate (group B)/ 11 severe (group C). All the patients who died were classified as group C. The sensitivity of A-DROP, CURB-65, PSI and I-ROAD for death were 12.5, 0, 87.5 and 100% respectively. The positive predictive value of each guideline for death was 33.3, 0, 50 and 72.7% respectively. Conclusion: Conventional predictive rules might underestimate the severity of non-HIV PCP. I-ROAD would be useful in evaluating the severity of non-HIV PCP.

Background: Activation of lipid metabolism is an early event of carcinogenesis and a central hallmark of many cancers including breast cancer. Recent findings argue that stearoyl CoA desaturase-1 (SCD1), a key regulator of the fatty acid (FA) composition and the endoplasmic reticulum resident enzyme that converts saturated FA (SFA) into monounsaturated FA (MUFA) is a novel regulator of carcinogenesis. The distinctive lipids composition of membrane in cancer cells and the biological functions of SCD1, however, still remain uncertain. Imaging mass spectrometry (IMS) is a mass spectrometry-based analyzing technique that enables visualization of the individual molecules without requiring antibodies. It allows comprehensive detection of a wide range of biomolecules, such as lipids. We attempted to visualize the localization of lipids in breast cancer by IMS for better understanding of cancer proliferation. Materials and methods: 13 specimens were obtained from the primary breast cancer patients. All were Japanese woman and aged 41–86 years (mean 61.5y.o.). Only one patient received preoperative systematic therapy. 6 were estrogen receptor (ER) and/or progesterone receptor (PgR) positive and human epidermal growth factor receptor 2 (HER2) negative, 2 were ER and/or PgR positive and HER2 positive, 2 were both ER and PgR negative and HER2 positive and 2 were triple negative. IMS: Samples were immediately chilled in liquid Hexan and stored at −80°. All specimens were sliced into 10 mm thin sections, mounted onto one indium-tin oxide-coated glass slides (Bruker Daltonics) and then sprayed by 2,5-Dihydroxybenzoic acid. Matrix assisted laser desorption ionization (MALDI) technique was used as a soft ionization method. We used time of flight (TOF)/TOF type instrument (Ultraflex, Bruker Daltonics) and all the spectrum were acquired automatically using Fleximaging software (Bruker Daltonics). Each spectral intensity at any mass-to-charge ratio (m/z) was measured at 16 regions of interest (ROI); 13 ROI were picked up from cancerous parts and 3 were from non-cancerous parts. Spectral intensities were compared and statistical analysis was performed by Mann Whitney test. The software was also used to create two-dimensional ion-density maps. Results: In the cancerous parts of all the 13 specimens, two distinct peaks of the molecular ions were detected at m/z 798.5 and 810.5, which were not found in the non-cancerous parts. Median intensity of the molecular ions at m/z 798.5 and 810.5 were 38.9 and 3.18 in the cancerous part, while they were 0.84 and 1.02 in the non-cancerous part (p=0.010 and 0.015, respectively). Tandem mass spectrometry analysis for these two molecules revealed that they were two kinds of phosphatidylcholine (PC), PC (16:0/18:1) and PC (18:0/18:1). Localization of the individual PC was visualized by means of IMS, which showed that in cancerous part accumulation of PCs containing MUFA was more pronounced than those containing SFA only. Conclusion: Two kinds of PC containing MUFA were found to highly accumulate in cancerous parts, which may suggest involvement of SCD1 in the membrane composition regulation and cancer proliferation. Further studies may thus be warranted to explore the relation between PC localization and the SCD1 expression. Citation Information: Cancer Res 2011;71(24 Suppl):Abstract nr P4-05-05.