Kristy L. Berry

Mesd is a general inhibitor of different Wnt ligands in Wnt/LRP signaling and inhibits PC-3 tumor growth in vivo.

Mesd is a specialized chaperone for Wnt co‐receptor low‐density lipoprotein receptor‐related protein‐5 (LRP5) and LRP6, which contain four β‐propeller/epidermal growth factor modules, named E1 to E4 from N‐ to C‐terminal, in their extracellular domains. Herein, we demonstrated that recombinant Mesd protein is a general Wnt inhibitor that blocks Wnt/β‐catenin signaling induced not only by LRP6 E1‐E2‐binding Wnts but also by LRP6 E3‐E4‐binding Wnts. We also found that Mesd suppressed Wnt/β‐catenin signaling induced by Wnt1 in prostate cancer PC‐3 cells, and inhibited tumor growth in PC‐3 xenograft model. Our results indicate that Mesd is a universal inhibitor of Wnt/LRP signaling on the cell surface.

Abstract 5159: Characterization of a novel class of RAS inhibitory compounds with potent anti-tumor activity

Introduction: Over 30% of all human cancers harbor activating RAS mutations which induce deregulation of the cell cycle, uncontrolled proliferation, and decreased apoptosis. Through a phenotypic screening strategy, we have identified a series of indene derivatives which potently and selectively inhibit growth of tumor cells harboring activated RAS. A development candidate from this series, DC070-547, disrupts RAS-RAF binding, inhibits RAS signaling, causes cell cycle arrest and induces apoptosis, and exhibits strong anti-tumor activity in a mouse KRAS mutant tumor model. Methods: Viable cell number was measured using a luminescent indicator of ATP. RAS activation was measured by GST-RAF1-RBD pull-down and western blotting using an anti-RAS antibody. Disruption of RAS-RAF binding was determined by pre-incubation of GST-RAF1-RBD beads with cell lysates or recombinant RAS in the presence of test compounds for 30 min. Cell cycle distribution was measured by DNA content and immunofluorescent detection of cell cycle proteins. Antitumor activity was determined in a subcutaneous mouse tumor model involving KRAS mutant colon tumors. Results: Low nanomolar concentrations of DC070-547 selectively inhibited growth of a diverse panel of tumor cell lines harboring activated RAS relative to tumor cell lines lacking activated RAS. Transfection of HT-29 cells lacking activated RAS with mutant RAS conferred sensitivity to DC070-547. The compounds blocked binding of RAF1-RBD to recombinant RAS, RAS from cell lysates, as well as RAS in intact cells. Sustained and potent growth inhibitory effects of DC070-547 were demonstrated by colony formation assays. Immunoblotting showed that DC070-547 inhibited EGF-induced signaling in HCT-116 colon tumor cells with activated RAS at concentrations that inhibit growth. DC070-547 also induced apoptosis as evident by Annexin V labeling and M-phase arrest in HCT116 cells as shown by DNA content and immunostaining of phospho-Histone H3B and Cdc25C, an important downstream mediator of RAS signaling. DC070-547 and three analogs from this series were evaluated for antitumor activity in an athymic mouse model using subcutaneously implanted KRAS mutant colon tumor cells. Treatments either completely suppressed tumor growth or caused tumor regression with no discernible toxicity. Conclusion: While RAS is widely considered to be non-druggable, a novel compound series was identified that potently and selectivity inhibit in vitro and in vivo the growth of tumor cells harboring activated RAS by inhibiting RAS-effector binding. Together, these findings support further preclinical development of this compound class for Phase I/II clinical evaluation for the treatment of RAS-driven cancers. Citation Format: Antonio Ward, Xi Chen, Jacob Valiyaveettil, Bing Zhu, Veronica Ramirez-Alcantara, Kevin J. Lee, Ashley Lindsey, Kristy Berry, Tyler E. Mattox, Kate McConnell, Michael R. Boyd, Gary A. Piazza, Adam B. Keeton. Characterization of a novel class of RAS inhibitory compounds with potent anti-tumor activity [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 5159. doi:10.1158/1538-7445.AM2017-5159

Abstract 5174: Phosphodiesterase 10A inhibition as a novel approach to suppress β-catenin signaling in ovarian cancer cells

Canonical Wnt/β-catenin signaling is known to be associated with platinum resistance in ovarian cancer in which inhibitors hold promise for the treatment of refractory disease. Phosphodiesterase 10A (PDE10A) is a dual cyclic AMP and cyclic GMP phosphodiesterase isozyme recently implicated in colon cancer. PDE10A inhibition in colon cancer cells by siRNA or small molecule inhibitors increased cGMP levels and activated PKG to inhibit β-catenin signaling. A novel PDE10 inhibitor, ADT-061, was identified by screening a library of indene derivatives, and showed strong antineoplastic activity in the Apc+/min-FCCC mouse (Lee K et al., unpublished data). Cyclic GMP and phosphodiesterases participate in the ovarian follicular development, although little is known about PDE10A expression in ovaries, especially with regard to a potential role in ovarian tumorigenesis. PDE10A protein was found to be expressed in various established ovarian cancer cell lines at higher levels than immortalized or primary ovarian surface epithelial cell lines. Pf-2545920, a known PDE10A inhibitor, and ADT-061 inhibited the growth of multiple ovarian tumor cell lines with IC50s around 20µM and 0.5µM, respectively. Both compounds induced apoptosis after 24h treatment, as measured by PI/Annexin-V staining and PARP cleavage. Pf-2545920 and ADT-061 induced phosphorylation of VASP at Ser157 and Ser239 in various ovarian cancer cell lines, indicating activation of cyclic AMP and cyclic GMP signaling, respectively. Treatment also decreased levels of β-catenin and downstream targets of TCF-dependent transcription, including c-MYC, survivin and cyclin-D1. Homozygous knockout PDE10A clones of OV-90 ovarian cancer cells obtained using CRISPR/Cas9 showed decreased clonogenic potential, decreased Pf-2545920-mediated VASP phosphorylation and β-catenin, c-MYC and survivin expression. Ongoing efforts are focused on the development of more potent ADT-061 analogs. These observations support further study of a role of PDE10 in ovarian tumorigenesis and the development of ADT-061 or analogs for the treatment of refractory ovarian cancer as well as the prevention of malignant recurrence. Citation Format: Luciana Madeira Da Silva, Elaine Gavin, Kevin J. Lee, Veronica Ramirez-Alcantara, Kristy L. Berry, Holly T. Taylor, Alla Musiyenko, Ileana V. Aragon, Adam B. Keeton, Jennifer Scalici, Rodney P. Rocconi, Gary A. Piazza. Phosphodiesterase 10A inhibition as a novel approach to suppress β-catenin signaling in ovarian cancer cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 5174. doi:10.1158/1538-7445.AM2017-5174

Abstract 1140: Characterization of a novel PDE10 inhibitor in lung tumor cells and an orthotopic mouse model of lung cancer

BACKGROUND: Screening a focused library of indene derivatives for PDE10 inhibitory activity identified novel leads with potent and selective tumor cell growth inhibitory activity. ADT-030 emerged from lead optimization chemistry with excellent drug-like properties and oral bioavailability. Here we characterize the anti-tumor activity of ADT-030 in human lung tumor cells and an orthotopic mouse model of lung cancer. METHODS: Growth inhibitory activity of ADT-030 was measured in a panel of human lung tumor cell lines by ATP quantification following 72 h of treatment. The effect of ADT-030 on intracellular cGMP/cAMP was measured in whole cell lysates using a competitive ELISA assay. PDE inhibitory activity of ADT-030 was evaluated in lysates of human lung tumor cells and by recombinant PDE isozymes using the IMAP fluorescence polarization PDE assay. Activation of PKG signaling and suppression of β-catenin levels in response to ADT-030 treatment was evaluated by Western blot using whole cell lysates of human lung tumor cells. ADT-030 was orally administrated to C57BL/6 mice and free levels quantified in plasma and tissues by LC-MS. Anti-tumor activity of ADT-030 was evaluated in athymic nude-Foxn1nu mice after inoculating the left lung with 1x106 A549 lung tumor cells and treating once daily by oral administration at dosages ranging from 25 - 125 mg/kg. Tumor growth was monitored by in situ bioluminescence using IVIS as well as necropsy and pathological grading after 4 weeks of treatment. RESULTS: ADT-030 inhibited the growth of human lung tumor cell lines with IC50 values in the low micromolar range by inducing apoptosis, while appreciably higher concentrations were required to affect the growth of normal human airway epithelial cells. ADT-030 treatment of human lung tumor cells increased both intracellular cGMP and cAMP levels, activated PKG and suppressed β-catenin within the same concentration range as required for tumor cell growth inhibition. Pharmacokinetic studies in mice demonstrated a half-life suitable for once a day dosing. Tissue distribution studies revealed appreciably higher concentrations of ADT-030 in lungs relative to plasma and other tissues, with the highest accumulation measured in the parenchyma. ADT-030 was well tolerated in mice implanted with A549 tumor cells and displayed strong anti-tumor activity as evident by reduced luminescence, tumor grading, and double-blinded pathological evaluation. CONCLUSIONS: ADT-030 represents a prospective drug development candidate with favorable drug-like properties that concentrates in lung after oral administration exhibiting a strong anti-tumor activity in a pre-clinical mouse model. The mechanism of lung tumor cell growth inhibition involves PDE10 inhibition, elevation of cGMP, activation of PKG, and attenuation of β-catenin. Citation Format: Veronica Ramirez-Alcantara, Bing Zhu, Xi Chen, Rajkumar Savai, Prema Subbarayal, Michele A. Schuler, Kevin J. Lee, Ashley S. Lindsey, Kristy L. Berry, Dennis Otali, Joshua Canzoneri, Jacob Valiyaveettil, Adam Keeton, Lori Coward, Gregory Gorman, William Grizzle, Michael Boyd, Gary A. Piazza. Characterization of a novel PDE10 inhibitor in lung tumor cells and an orthotopic mouse model of lung cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 1140. doi:10.1158/1538-7445.AM2017-1140

Abstract 4972: Novel Ras inhibitor DC070-547 potently and selectively blocks Ras-RBD binding, EGFR binding to Ras signaling complex, EGFR activation of Ras signaling, and growth of Ras-driven lung tumor cells

A novel series of compounds that potently and selectively inhibits the growth of tumor cells harboring constitutively activated Ras relative to cells lacking activated Ras were identified by screening a proprietary library of indene derivatives in a phenotypic, cell-based assay. Lead-optimization produced a drug development candidate, DC070-547, which showed strong antitumor activity at doses not causing any discernible toxicity in preclinical mouse models. Here we characterize the underlying mechanism of growth inhibition in lung tumor cells. A panel of non-small cell lung cancer lines with constitutively activated Ras were highly sensitive to DC070-547 with IC50 values as low as 2 nM, while normal airway epithelial cells were essentially insensitive. Transfection of wild-type ras H322 bronchioalveolar tumor cells with mutant ras (G12V) confirmed that activated Ras is required for the selective growth inhibitory activity of DC070-547. Ras-RBD binding assays showed that DC070-547 disrupts Ras-RBD binding at low nanomolar concentrations that parallel those required to inhibit the growth of lung tumor cells with activated Ras. Similar concentrations of DC070-547 were found to inhibit the binding of phosphorylated EGFR (Y1068) to Ras immunoprecipitates in mutant ras transfected H322 cells, but not in control H322 cells. DC070-547 also inhibited the binding of SOS, Grb2, Gab1, S338 phosphorylated c-Raf (pc-Raf), S473 phosphorylated Akt-1 (pAkt-1) and T202/Y204 phosphorylated Erk1/2 (pERK1/2) to Ras or EGFR immunoprecipitates. To determine if DC070-547 can inhibit EGF-stimulated Ras signaling, serum-starved mutant ras transfected H322 cells were treated with EGF and probed for effects on Ras signaling components. DC070-547 caused a concentration-dependent inhibition of EGF-induced Y1068-EGFR as measured in Ras immunoprecipitates, and also reduced pc-Raf, pAkt-1, pErk1/2 and pGab1 (Y627) levels in Ras or EGFR immunoprecipitates. In addition, DC070-547 caused a concentration-dependent decrease in Erk1/2 and Akt-1-mediated phosphorylation of Bad proteins (S112, S136 and S155) to induce apoptosis. These results show that DC070-547 prevents Ras-RBD binding to block EGF-induced Raf/MAPK and Akt signaling to potently and selectively inhibit the growth of lung tumor cells harboring constitutively activated Ras. Support provided by NCI grants 1R01CA197147 and 1R21CA182941. Citation Format: Bing Zhu, Xi Chen, Jacob Valiyaveettil, Joshua Canzoneri, Kevin Lee, Kate Saville, Kristy Berry, Luciana Barnes, Tyler Maddox, Ashley Lindsey, Antonio Ward, Veronica Ramirez-Alcantara, Adam Keeton, Michael Boyd, Gary Piazza. Novel Ras inhibitor DC070-547 potently and selectively blocks Ras-RBD binding, EGFR binding to Ras signaling complex, EGFR activation of Ras signaling, and growth of Ras-driven lung tumor cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 4972. doi:10.1158/1538-7445.AM2017-4972

Abstract 3730: A quick and cost effective 12-cell line panel assay to predict drug activity in human tumor xenograft models

Proceedings: AACR Annual Meeting 2014; April 5-9, 2014; San Diego, CA The procedure to identify and develop an anti-cancer drug first involves testing drug candidates in cell lines followed by human tumor xenograft models, usually selected based upon the histotype of the cell lines in which the drug showed optimal activity. Many drugs fail at this stage, as activity in cell lines does not often correlate with activity in xenograft models. This is not surprising, as we have previously shown that gene expression in xenograft models does not necessarily correlate with the cell line from which it was derived. In an attempt to improve the success rate of drugs tested in xenograft models, we have developed a fast and cost effective 12-panel human tumor cell line assay that represents the genetic diversity of all our xenograft models and several different cancer histotypes. Affymetrix genomic analysis was performed on 100 human tumor xenograft and cell line models. The genomic profiles obtained underwent Unsupervised Hierarchical Cluster Analysis to group models with similar genetic profiles. This analysis resulted in 12 distinct clusters; a representative cell line was chosen from each cluster. Stocks of each representative cell line were frozen and tested to ensure exponential growth immediately upon thawing, resulting in no waiting time for drug testing. It follows that if a candidate drug shows activity in one or more of these representative cell lines, other cell lines and/or xenograft models in the same cluster can also be tested. As the cell lines and xenograft models within the same cluster will have a similar genetic profile, the chances of success should thus be increased. To test the effectiveness of this approach, we used our database to further develop an internal compound. SRI-20900 had been tested previously in the CCRF-CEM and CAKI-1 xenograft models. The compound showed no activity in CCRF-CEM cells, but excellent activity in CAKI-1 cells. These models were in completely different clusters. So, based on these data, we tested the compound in the SKOV-3 and IGROV-1 xenograft models, as these clustered closely to the CAKI-1 model. The compound showed excellent activity in both SKOV-3 and IGROV-1 models. Although these data provide proof of principle, further work needs to be done by testing targeted compounds in the 12-cell line panel, followed by testing in xenograft models within the same cluster as the cell lines that show optimal activity. In addition, it would follow that a xenograft model within the same cluster as an inactive cell line should also be tested. We hope to start these studies early in 2014. Citation Format: Michael J. Roberts, Tommie A. Gamble, Richard D. May, Murray Stackhouse, Kristy L. Berry, Andrew D. Penman, Robert J. Rooney, Yulia Maxuitenko, Michael S. Koratich. A quick and cost effective 12-cell line panel assay to predict drug activity in human tumor xenograft models. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 3730. doi:10.1158/1538-7445.AM2014-3730

Abstract 2778: Tumor target vs. tissue of tumor origin: cluster analysis of genomic profile of 42 human tumor in vitro and in vivo models.

Traditionally, drug development has relied upon testing cancer drug candidates in cell lines. Active drugs are then tested in human tumor xenograft models, usually selected based upon the cell lines in which the drug showed activity. The majority of drugs fail at this stage as they do not show activity in the xenograft models chosen. We performed Affymetrix genomic analysis on 42 human tumor xenograft models and the original cell lines from which they were established. The genomic profiles obtained underwent Unsupervised Hierarchical Cluster Analysis to ascertain which cell lines and xenograft models had similar genomic profiles and which did not. The analysis showed that only 24 of 42 human tumor xenograft models clustered side-by-side with the cell line from which they were established. All 6 human leukemia/lymphoma xenograft models clustered very well with the cell lines from which they were established, and they clustered perfectly according to histological class. Five out of six human colon tumor xenograft models clustered well with the cell lines from which they were established and according to histotype. Of the 18 xenograft/cell line pairs that did not cluster side-by-side, 10 pairs remained in the same general cluster, whereas the partners of 8 other pairs were dispersed across different major clusters. Ovarian, breast, melanoma, and pancreatic human tumor xenograft models did not cluster according to histotype. Our data may explain why some drugs that show in vitro activity in some cell lines are not active in other cell lines of the same histological type, and also why some drugs that show activity in vitro then fail in xenograft models. In our laboratory, the PANC-1 cell line is very often chosen as a model of pancreatic cancer. A drug showing activity in the PANC-1 cell line would next be tested in other in vitro models of pancreatic cancer (e.g., MIA PaCa-2, CFPAC-1, and BxPC-3). However, none of these other pancreatic models have a similar genetic profile to PANC-1. Based upon our data, the cell line showing most similarity to the PANC-1 cell line is the breast cancer cell line MDA-MB-231. It is our suggestion that a drug showing activity in the PANC-1 cell line should be tested in other cell lines showing similar genetic profiles, not in cell lines based on histotype. Another example from our analysis is the LOX-IMV1 melanoma cell line. Not only does this cell line not cluster with its corresponding LOX-IMV1 xenograft model, it clusters most closely with the NCI/ADR-RES ovarian cell line. In summary, the genomic profiles of approximately 57% of the tumor xenograft models analyzed closely associate with the cell line from which they were established. Some of the tumor xenograft models show very little similarity to the cell lines from which they were established. Additionally, many of the models (both xenografts and cell lines), do not cluster according to their tissue of origin. Citation Format: Michael J. Roberts, Michael S. Koratich, Murray Stackhouse, Richard D. May, Andrew D. Penman, Tommie A. Gamble, Kristy L. Berry, Joseph F. Murphy, Robert J. Rooney, Yulia Y. Maxuitenko. Tumor target vs. tissue of tumor origin: cluster analysis of genomic profile of 42 human tumor in vitro and in vivo models. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 2778. doi:10.1158/1538-7445.AM2013-2778

Abstract 3114: Effects of tumor-stromal interactions on gene expression in panel of mouse tumor models

Tumor growth is not determined solely by the tumor cells but is governed by interactions between tumor cells and host stromal cells, including endothelial cell activation and fibroblastic stroma response. Tumor stroma profoundly influences many steps of tumor progression. In many human cancers, such as breast, prostate, and colon, the stroma comprises the majority of the tumor mass, as a hallmark of the clinical feature called desmoplasia. Numerous studies have showed that tumor-stromal cell interactions play crucial roles in supporting cancer progression and in promoting anticancer drug resistance by alternating gene expression profiles in both tumor and stromal cells through network tumor-stromal interactions in the tumor microenvironments. It has been challenging to obtain separate gene profiles for tumor and stroma with human tumor samples as both tumor and stromal cells share the same genome. In xenograft mouse tumor models, human tumor cells are supported by mouse host stromal microenvironment. Therefore, effects of tumor stromal integrations on gene expression can be profiled separately by taking advantage of this heterogeneous genetic makeup. To understand the underlying biological process of stroma in cancer and select relevant in vitro and in vivo model systems for various targeted anticancer drug discovery and development projects, we selected a panel of 30 commonly used xenograft tumor mouse models that are derived from human tumor cell lines of various cancer histotypes and conducted analyses of differential gene expression in both human cancer cells and mouse host stromal cells before and after their interactions in vivo by using quantitative PCR with mouse and human specific primers. Growing the human tumors as a continuous in vivo passage subcutaneously in immunodeficient mice permits stroma infiltration over a long time. We have examined a number of anticancer target genes involved in different signaling pathways, such as tumor angiogenesis, apoptosis and survival (Akt/mTOR signaling pathway). Our study results have demonstrated that tumor stromal interactions significantly regulate expression levels of various genes important in tumor progression and development of resistance to treatment in both tumor and stromal cells. Tumor animal models play a critical role in translating the bench science to the bedside medical care of cancer patients. Decisions for moving new anticancer agents into costly clinical investigations are mostly based on the preclinical results using xenograft mouse models. The results of this gene profiling approach could provide tools for studying tumor microenvironment and tumor stromal interactions in vivo to advance anticancer drug development. 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 3114. doi:10.1158/1538-7445.AM2011-3114

Abstract 3943: Affymetrix whole genome microarray analysis of 51 human tumor cell lines representative of 16 different tissues of origin: Unsupervised hierarchical clustering

The total mRNA for each sample of 51 human tumor cell lines representative of 16 different tissues of origin was split into 3 replicates, and analyzed against the entire human genome using standard Affx WT procedures. Approximately 50% of the samples (25 out of the 51 cell lines tested) exhibited low-level clustering related to their tissue of origin. The remaining 50% (26 out of the 51 cell lines tested) did not cluster with other samples of the same tissue of origin. These data reveal the importance of testing potential anticancer agents in multiple models representative of several different tumors of origin, as there is a 50% chance that the model chosen is not actually representative of the intended tissue of origin. This analysis also showed that the pancreatic cancer cell line CFPAC-1 did not cluster with any other cell line tested, revealing the unique genetic profile of this cell line. Interestingly, the reported lung cancer cell lines NCI-H69 and NCI-H82 clustered more closely with leukemic lines than with lung or any other solid tumor. This is particularly interesting as these lines are known to grow/behave more like a suspension culture than a monolayer. The NCI recently published its genetic analysis of their 60-panel, and they revealed that the MDA-MB-435 cell line, traditionally thought to be a breast cancer cell line, more closely resembled a melanoma line; hence, it was re-classified as a melanoma (likely a metastasized melanoma that was taken from the breast site). Our analysis reveals that another traditional breast cancer cell line, UISO-BCA-1, also clusters more closely with the melanomas (including the MDA-MB-435 cell line), suggesting that this cell line also may have been misclassified. Based on these data, we suggest that any potential anticancer agent showing activity in a particular cell line should be tested in other cell lines that cluster with the active line, and not merely in other lines supposedly representative of the same tissue of origin. Furthermore, in early stage testing, it would be more prudent to test several cell lines from different clusters, rather than several cell lines from different tissues of origin. It follows that by testing orphan drugs against several cell lines from each cluster, it would be possible to significantly narrow (and possibly identify), the likely drug target. 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 3943. doi:10.1158/1538-7445.AM2011-3943

Abstract 4418: Affymetrix whole genome microarray analysis of 24 human tumor xenograft models and the cell lines from which they were developed: Clustering of targets vs tumor tissue of origin

The NCI recently performed microarray expression analysis on its 60-panel of human tumor cell lines. This revealed important information, with some cell lines shown to be from a different tissue of origin than originally believed. Traditionally, potential anti-cancer agents have been evaluated in these in vitro models, and then moved into the corresponding in vivo xenograft model(s) based on the in vitro results. It has been shown that drugs which are effective in vitro are not necessarily effective in vivo and vice versa. Systematic microarray analysis of traditional xenograft models in conjunction with their in vitro counterparts has not been performed. The development of a human tumor xenograft in a mouse might be expected to lead to changes in gene expression, and this could account, in some instances, for the disconnect in results observed between in vitro and in vivo models. Our aim was to perform a genetic analysis against the entire human genome using 24 cell lines from 11 differing tissues of origin that were implanted into immune-deficient mice to establish a xenograft model for each. Once the tumors reached approximately 1 cm3 in size, the tumors were removed, cut into approx. 2-3 mm3 fragments, and an in vivo tumor passage was established. Microarray expression in fragments of those xenografted tumors was compared to microarray expression in the cell line from which they were developed. The total mRNA for each sample was split into 3 replicates, and analyzed against the entire human genome using standard Affx WT procedures. The results showed that over 60% (15 of 24) of the xenograft samples clustered with the cell line from which it was developed, whereas approximately 40% (9 of 24) did not, revealing that major changes in gene expression had occurred in 40% of these xenograft samples. Furthermore, when analyzed alone, these particular 24 cell line samples clustered according to their tissue of origin, whereas the tumor fragment samples did not appear to cluster. On the basis of these data we are currently performing the same analysis on an additional 25 tumor fragments and their corresponding matched cell lines to allow for a more accurate, in-depth cluster analysis. These data strongly suggest that although precedent exists to select in vitro models on the basis of their tissue of origin, no such precedent exists for in vivo models. In vivo models should be more carefully selected to ensure that the model chosen is still representative of the tissue to be tested. It follows that a drug candidate effective in a particular in vitro model might be expected to show activity in other in vitro lines from the same tissue of origin. However, a drug candidate effective in a particular in vivo model representative of a tissue of origin should not be expected to show efficacy in other in vivo models representing the same tissue type. 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 4418. doi:10.1158/1538-7445.AM2011-4418