Patient Derived Xenografts (PDX) Models as an Avatar to Assess Personalized Therapy Options in Uveal Melanoma: A Feasibility Study.

Uveal melanoma is the most common primary intraocular malignancy in adults. Up to 50% of UM patients develop metastatic disease, usually in the liver. When metastatic, the prognosis is poor, and few treatment options exist. Here, we investigated the feasibility of establishing patient-derived xenografts (PDXs) from a patient's tumor in order to screen for therapies that the patient could benefit from. Samples obtained from 29 primary tumors and liver metastases of uveal melanoma were grafted into SCID mice. PDX models were successfully established for 35% of primary patient tumors and 67% of liver metastases. The tumor take rate was proportional to the risk of metastases. PDXs showed the same morphology, the same GNAQ/11, BAP1, and SF3B1 mutations, and the same chromosome 3 and 8q status as the corresponding patient samples. Six PDX models were challenged with two compounds for 4 weeks. We show that, for 31% of patients with high or intermediate risk of metastasis, the timing to obtain efficacy results on PDX models derived from their primary tumors was compatible with the selection of the therapy to treat the patient after relapse. PDXs could thus be a valid tool ("avatar") to select the best personalized therapy for one third of patients that are most at risk of relapse.

Two-photon microscopy for microrobotics: Visualization of micro-agents below fixed tissue.

Optical microscopy is frequently used to visualize microrobotic agents (i.e., micro-agents) and physical surroundings with a relatively high spatio-temporal resolution. However, the limited penetration depth of optical microscopy techniques used in microrobotics (in the order of 100 µm) reduces the capability of visualizing micro-agents below biological tissue. Two-photon microscopy is a technique that exploits the principle of two-photon absorption, permitting live tissue imaging with sub-micron resolution and optical penetration depths (over 500 µm). The two-photon absorption principle has been widely applied to fabricate sub-millimeter scale components via direct laser writing (DLW). Yet, its use as an imaging tool for microrobotics remains unexplored in the state-of-the-art. This study introduces and reports on two-photon microscopy as an alternative technique for visualizing micro-agents below biological tissue. In order to validate two-photon image acquisition for microrobotics, two-type micro-agents are fabricated and employed: (1) electrospun fibers stained with an exogenous fluorophore and (2) bio-inspired structure printed with autofluorescent resin via DLW. The experiments are devised and conducted to obtain three-dimensional reconstructions of both micro-agents, perform a qualitative study of laser-tissue interaction, and visualize micro-agents along with tissue using second-harmonic generation. We experimentally demonstrate two-photon microscopy of micro-agents below formalin-fixed tissue with a maximum penetration depth of 800 µm and continuous imaging of magnetic electrospun fibers with one frame per second acquisition rate (in a field of view of 135 × 135 µm2). Our results show that two-photon microscopy can be an alternative imaging technique for microrobotics by enabling visualization of micro-agents under in vitro and ex ovo conditions. Furthermore, bridging the gap between two-photon microscopy and the microrobotics field has the potential to facilitate in vivo visualization of micro-agents.

[Home treatment of low-risk pulmonary embolism patients : Efficacy and safety of an outpatient program including the general practitioner]. / Prise en charge ambulatoire des embolies pulmonaires à risque faible ; expérience d'une filière centrée sur le lien ville-hôpital.

AIM: Outpatient treatment (OT) of patients with low-risk pulmonary embolisms (PE) is recommended. A multidisciplinary OT program including the general practitioner (GP) has been implemented at Versailles hospital in 2019. The objectives of the study were to assess the feasibility, safety and acceptability of the program. MATERIAL AND METHODS: The feasibility of, and the inclusion criteria for OT were defined from a retrospective cohort study of PE patients carried out in 2018. In the prospective study, consecutive patients consulting in the emergency department between 2019 and 2021 with confirmed PE were eligible for OT if they had sPESI and HESTIA scores equal to 0, normal troponin and NT-pro-BNP levels, and no right ventricular dilation on imaging. PEs associated with COVID were excluded. The OT program included 4 appointments within 3 months, including 2 with the GP. Events (death, recurrence of PE or venous thromboembolism, bleeding, rehospitalisation) were collected at 3-month follow-up. RESULTS: In the retrospective study, 19% of the 138 PE patients seen in the emergency department were eligible for OT. No complication occurred at Day 90. In the prospective study, 313 consecutive patients with confirmed PE in the emergency department were included, 66 (21%) were eligible for OT. Overall, 43 patients (14%) received OT (39 eligible) and 27 patients eligible for OT were hospitalised (92% because of pulmonary infarction). At 3-month follow-up, there were no death, no recurrence of thromboembolism, and one patient has been early hospitalised for COVID; 3 female patients treated with rivaroxaban had minor bleeding (heavy menstrual bleeding). The satisfaction rate of general practitioner was 95%. CONCLUSIONS: This study confirms the feasibility and safety of our OT program for low-risk EP patients, centered on the general practitioner. It reduces the time spent in the emergency department, reduces hospitalisations and strengthens the city-hospital link for care.

In vitro bone metastasis dwelling in a 3D bioengineered niche.

Bone is the most frequent metastasis site for breast cancer. As well as dramatically increasing disease burden, bone metastases are also an indicator of poor prognosis. One of the main challenges in investigating bone metastasis in breast cancer is engineering in vitro models that replicate the features of in vivo bone environments. Such in vitro models ideally enable the biology of the metastatic cells to mimic their in vivo behavior as closely as possible. Here, taking benefit of cutting-edge technologies both in microfabrication and cancer cell biology, we have developed an in vitro breast cancer bone-metastasis model. To do so we first 3D printed a bone scaffold that reproduces the trabecular architecture and that can be conditioned with osteoblast-like cells, a collagen matrix, and mineralized calcium. We thus demonstrated that this device offers an adequate soil to seed primary breast cancer bone metastatic cells. In particular, patient-derived xenografts being considered as a better approach than cell lines to achieve clinically relevant results, we demonstrate the ability of this biomimetic bone niche model to host patient-derived xenografted metastatic breast cancer cells. These patient-derived xenograft cells show a long-term survival in the bone model and maintain their cycling propensity, and exhibit the same modulated drug response as in vivo. This experimental system enables access to the idiosyncratic features of the bone microenvironment and cancer bone metastasis, which has implications for drug testing.

A large collection of integrated genomically characterized patient-derived xenografts highlighting the heterogeneity of triple-negative breast cancer.

Triple-negative breast cancer (TNBC) represents 10% of all breast cancers and is a very heterogeneous disease. Globally, women with TNBC have a poor prognosis, and the development of effective targeted therapies remains a real challenge. Patient-derived xenografts (PDX) are clinically relevant models that have emerged as important tools for the analysis of drug activity and predictive biomarker discovery. The purpose of this work was to analyze the molecular heterogeneity of a large panel of TNBC PDX (n = 61) in order to test targeted therapies and identify biomarkers of response. At the gene expression level, TNBC PDX represent all of the various TNBC subtypes identified by the Lehmann classification except for immunomodulatory subtype, which is underrepresented in PDX. NGS and copy number data showed a similar diversity of significantly mutated gene and somatic copy number alteration in PDX and the Cancer Genome Atlas TNBC patients. The genes most commonly altered were TP53 and oncogenes and tumor suppressors of the PI3K/AKT/mTOR and MAPK pathways. PDX showed similar morphology and immunohistochemistry markers to those of the original tumors. Efficacy experiments with PI3K and MAPK inhibitor monotherapy or combination therapy showed an antitumor activity in PDX carrying genomic mutations of PIK3CA and NRAS genes. TNBC PDX reproduce the molecular heterogeneity of TNBC patients. This large collection of PDX is a clinically relevant platform for drug testing, biomarker discovery and translational research.

Primary CNS lymphoma patient-derived orthotopic xenograft model capture the biological and molecular characteristics of the disease.

Primary CNS lymphomas (PCNSL) are rare and poor prognosis diffuse large B-cell lymphomas. Because of the brain tumor environment and the restricted distribution of drugs in the CNS, specific PCNSL patient-derived orthotopic xenograft (PDOX) models are needed for preclinical research to improve the prognosis of PCNSL patients. PCNSL patient specimens (n = 6) were grafted in the caudate nucleus of immunodeficient nude mice with a 83% rate of success, while subcutaneous implantation in nude mice of human PCNSL sample did not generate lymphoma, supporting the role of the brain microenvironment in the PCNSL physiopathology. PDOXs showed diffuse infiltration of B-cell lymphoma cells in the brain parenchyma. Each model had a unique mutational signature for genes in the BCR and NF-κB pathways and retained the mutational profile of the primary tumor. The models can be stored as cryopreserved biobank. Human IL-10 levels measured in the plasma of PCNSL-PDOX mice showed to be a reliable tool to monitor the tumor burden. Treatment response could be measured after a short treatment with the targeted therapy ibrutinib. In summary, we established a panel of human PCNSL-PDOX models that capture the histological and molecular characteristics of the disease and that proved suitable for preclinical experiments. Our methods of generation and characterization will enable the generation of additional PDOX-PCNSL models, essential tools for cognitive and preclinical drug discovery.

Inhibition of mTOR downregulates expression of DNA repair proteins and is highly efficient against BRCA2-mutated breast cancer in combination to PARP inhibition.

Breast cancer is a complex disease in which each patient could present several genetic alterations that are therapeutically relevant in cancers. Here we explored the therapeutic benefit of combining PARP and mTOR inhibitors in a context of DNA repair deficiency and PI3K pathway activation. The combination of everolimus and olaparib was tested in BRCA2-mutated patient-derived xenografts (PDX) carrying alterations in the PI3K/AKT/mTOR pathway. An RPPA analysis of different signalling pathways was performed in untreated and treated xenografts. Everolimus and olaparib showed marked anti-tumor activities in the monotherapy setting and high efficacy when given in combination with 100% of mice showing tumor regressions. The fraction of P-H2AX positive cells was increased in both monotherapy arms and strongly increased in the combination setting. Everolimus given as monotherapy resulted in downregulation of different proteins involved in DNA damage repair, including FANCD2, RAD50 and SUV39H1. In the combination setting, expression of these proteins was almost completely abolished, suggesting convergence of PARP and mTOR in downregulation of DNA damage repair components. In conclusion, our results suggest that combining mTOR and DNA repair inhibition could be a successful strategy to treat a subset of breast cancer with BRCA2 mutation and alterations in the PI3K/AKT/mTOR pathway.

The iron chelator deferasirox synergises with chemotherapy to treat triple-negative breast cancers.

To ensure their high proliferation rate, tumor cells have an iron metabolic disorder causing them to have increased iron needs, making them more susceptible to iron deprivation. This vulnerability could be a therapeutic target. In breast cancers, the development of new therapeutic approaches is urgently needed for patients with triple-negative tumors, which frequently relapse after chemotherapy and suffer from a lack of targeted therapies. In this study, we demonstrated that deferasirox (DFX) synergises with standard chemotherapeutic agents such as doxorubicin, cisplatin and carboplatin to inhibit cell proliferation and induce apoptosis and autophagy in triple-negative breast cancer (TNBC) cells. Moreover, the combination of DFX with doxorubicin and cyclophosphamide delayed recurrences in breast cancer patient-derived xenografts without increasing the side-effects of chemotherapies alone or altering the global iron storage of mice. Antitumor synergy of DFX and doxorubicin seems to involve downregulation of the phosphoinositide 3-kinase and nuclear factor-κB pathways. Iron deprivation in combination with chemotherapy could thus help to improve the effectiveness of chemotherapy in TNBC patients without increasing toxicity. Copyright © 2018 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

PKD1 is a potential biomarker and therapeutic target in triple-negative breast cancer.

Protein Kinase D1 (PKD1) is a serine/threonine kinase encoded by the PRKD1 gene. PKD1 has been previously shown to be a prognostic factor in ERα+ tamoxifen-resistant breast tumors and PKD1 overexpression confers estrogen independence to ERα+ MCF7 cells. In the present study, our goal was to determine whether PKD1 is a prognostic factor and/or a relevant therapeutic target in breast cancer. We analyzed PRKD1 mRNA levels in 527 primary breast tumors. We found that high PRKD1 mRNA levels were significantly and independently associated with a low metastasis-free survival in the whole breast cancer population and in the triple-negative breast cancer (TNBC) subtype specifically. High PRKD1 mRNA levels were also associated with a low overall survival in TNBC. We identified novel PKD1 inhibitors and assessed their antitumor activity in vitro in TNBC cell lines and in vivo in a TNBC patient-derived xenograft (PDX) model. Pharmacological inhibition and siRNA-mediated depletion of PKD1 reduced colony formation in MDA-MB-436 TNBC cells. PKD1 inhibition also reduced tumor growth in vivo in a TNBC PDX model. Together, these results establish PKD1 as a poor prognostic factor and a potential therapeutic target in TNBC.

Capecitabine Efficacy Is Correlated with TYMP and RB1 Expression in PDX Established from Triple-Negative Breast Cancers.

Purpose: Triple-negative breast cancer (TNBC) patients with residual disease after neoadjuvant chemotherapy have a poor outcome. We developed patient-derived xenografts (PDX) from residual tumors to identify efficient chemotherapies and predictive biomarkers in a context of resistance to anthracyclines- and taxanes-based treatments.Experimental Design: PDX were established from residual tumors of primary breast cancer patients treated in neoadjuvant setting. TNBC PDX were treated by anthracyclines, taxanes, platins, and capecitabine. Predictive biomarkers were identified by transcriptomic and immunohistologic analysis. Downregulation of RB1 was performed by siRNA in a cell line established from a PDX.Results: Residual TNBC PDX were characterized by a high tumor take, a short latency, and a poor prognosis of the corresponding patients. With the exception of BRCA1/2-mutated models, residual PDX were resistant to anthracyclines, taxanes, and platins. Capecitabine, the oral prodrug of 5-FU, was highly efficient in 60% of PDX, with two models showing complete responses. Prior treatment of a responder PDX with 5-FU increased expression of thymidylate synthase and decreased efficacy of capecitabine. Transcriptomic and IHC analyses of 32 TNBC PDX, including both residual tumors and treatment-naïve derived tumors, identified RB1 and TYMP proteins as predictive biomarkers for capecitabine response. Finally, RB1 knockdown in a cell line established from a capecitabine-responder PDX decreased sensitivity to 5-FU treatment.Conclusions: We identified capecitabine as efficient chemotherapy in TNBC PDX models established from residual disease and resistant to anthracyclines, taxanes, and platins. RB1 positivity and high expression of TYMP were significantly associated with capecitabine response. Clin Cancer Res; 24(11); 2605-15. ©2018 AACR.

Combination of COX-2 expression and PIK3CA mutation as prognostic and predictive markers for celecoxib treatment in breast cancer.

COX-2 expression level and prognostic value are still a matter of debate in breast cancer (BC). We addressed these points in the context of PIK3CA mutational status. Based on an interesting study of aspirin efficacy in colorectal cancer, we hypothesized that celecoxib antitumoral activity may be restricted to PIK3CA mutated BC.COX-2 mRNA expression was analyzed in 446 BC samples and in 61 BC patient-derived xenografts (PDX) using quantitative RT-PCR. The prognostic impact of COX-2 expression level was assessed independently and according to PIK3CA mutational status in our cohort and in a validation set of 817 BC. The antitumoral activity of celecoxib was tested in two triple-negative (TN) PDX with a PIK3CA wild-type (wt) or mutated genotype.COX-2 mRNA was overexpressed in 2% of BC and significantly associated with TN subtype. Metastasis-free survival (MFS) was significantly better in patients with high COX-2 expression level, the prognosis of whom was similar to patients with PIK3CA mutations. TCGA validation cohort confirmed that patients with low COX-2 expression PIK3CA wt tumors had the worse disease-free survival (DFS) compared to all other subgroups. Celecoxib had a significant antitumoral effect in PIK3CA mutated PDX only. Celecoxib antitumoral activity involved S6 ribosomal protein and AKT phosphorylation.Low expression of COX-2 has a significant negative impact on the MFS/DFS of BC patients. Antitumoral effect of celecoxib is restricted to PIK3CA mutated PDX. These results suggest that PIK3CA mutation may be a new predictive biomarker for celecoxib efficacy.

Combination of Carboplatin and Bevacizumab Is an Efficient Therapeutic Approach in Retinoblastoma Patient-Derived Xenografts.

PURPOSE: Retinoblastoma (Rb) is a rare childhood cancer of the retina with a survival rate of 95% in children living in high-income countries, after appropriate therapies such as chemotherapy, local ophthalmologic treatment, and radiotherapy. However, due to inactivation of the RB1 gene, all bilateral and almost 15% of unilateral retinoblastoma patients have a higher risk of s econdary cancers, especially sarcomas. Hence, new nonmutagen treatments are warranted. Therefore, we investigated the efficacy of therapy using anti-VEGF antibody bevacizumab, either alone or with carboplatin, in well-characterized Rb patient-derived xenografts (PDXs). METHODS: Three Rb PDXs previously established and characterized, RB102, RB111, and RB200, have been treated using carboplatin, bevacizumab, or carboplatin + bevacizumab. In order to define antitumor responses, various quantitative PCR and histopathologic analyses have then been performed on tumors collected at the end of experiments. RESULTS: In all treated PDX models, we have observed a high and significant improvement of chemotherapy-induced in vivo efficacy by the antiangiogenic antibody. The overall response rate, lower than -0.5, was 48%, 27%, and 86% after carboplatin, bevacizumab, and carboplatin + bevacizumab, respectively (carboplatin versus carboplatin + bevacizumab; P < 10-2; bevacizumab versus carboplatin + bevacizumab; P < 10-3). In the Rb200 PDX, such a result was also observed when bevacizumab was combined with lower doses of carboplatin. Quantitative PCR and histopathologic analyses have been performed and confirmed the impact of the bevacizumab-based treatments on various angiogenic markers. CONCLUSIONS: Overall, our in vivo results confirm the interest in antiangiogenic therapy for the treatment of Rb in combination with carboplatin and provide a robust rationale for testing this combination in the clinical setting for Rb patients.

Targeting mTOR pathway inhibits tumor growth in different molecular subtypes of triple-negative breast cancers.

Triple-negative breast cancers (TNBC) are characterized by frequent alterations in the PI3K/AKT/mTOR signaling pathway. In this study, we analyzed PI3K pathway activation in 67 patient-derived xenografts (PDX) of breast cancer and investigated the anti-tumor activity of the mTOR inhibitor everolimus in 15 TNBC PDX with different expression and mutational status of PI3K pathway markers. Expression of the tumor suppressors PTEN and INPP4B was lost in 55% and 76% of TNBC PDX, respectively, while mutations in PIK3CA and AKT1 genes were rare. In 7 PDX treatment with everolimus resulted in a tumor growth inhibition higher than 50%, while 8 models were classified as low responder or resistant. Basal-like, LAR (Luminal AR), mesenchymal and HER2-enriched tumors were present in both responder and resistant groups, suggesting that tumor response to everolimus is not restricted to a specific TNBC subtype. Analysis of treated tumors showed a correlation between tumor response and post-treatment phosphorylation of AKT, increased in responder PDX, while PI3K pathway markers at baseline were not sufficient to predict everolimus response. In conclusion, targeting mTOR decreased tumor growth in 7 out of 15 TNBC PDX tested. Response to everolimus occurred in different TNBC subtypes and was associated with post-treatment increase of P-AKT.

Characterization of Breast Cancer Preclinical Models Reveals a Specific Pattern of Macrophage Polarization.

Drug discovery efforts have focused on the tumor microenvironment in recent years. However, few studies have characterized the stroma component in patient-derived xenografts (PDXs) and genetically engineered mouse models (GEMs). In this study, we characterized the stroma in various models of breast cancer tumors in mice. We performed transcriptomic and flow cytometry analyses on murine populations for a series of 25 PDXs and the two most commonly used GEMs (MMTV-PyMT and MMTV-erBb2). We sorted macrophages from five models. We then profiled gene expression in these cells, which were also subjected to flow cytometry for phenotypic characterization. Hematopoietic cell composition, mostly macrophages and granulocytes, differed between tumors. Macrophages had a specific polarization phenotype related to their M1/M2 classification and associated with the expression of genes involved in the recruitment, invasion and metastasis processes. The heterogeneity of the stroma component of the models studied suggests that tumor cells modify their microenvironment to satisfy their needs. Our observations suggest that such models are of relevance for preclinical studies.

Supersonic Shear Wave Elastography of Response to Anti-cancer Therapy in a Xenograft Tumor Model.

Our objective was to determine if supersonic shear wave elastography (SSWE) can detect changes in stiffness of a breast cancer model under therapy. A human invasive carcinoma was implanted in 22 mice. Eleven were treated with an anti-angiogenic therapy and 11 with glucose for 24 d. Tumor volume and stiffness were assessed during 2 wk before treatment and 0, 7, 12, 20 and 24 d after the start of therapy using SSWE. Pathology was assessed after 12 and 24 d of treatment. We found that response to therapy was associated with early softening of treated tumors only, resulting in a significant difference from non-treated tumors after 12 d of treatment (p = 0.03). On pathology, large areas of necrosis were observed at 12 d in treated tumors. Although treatment was still effective, treated tumors subsequently stiffened during a second phase of the treatment (days 12-24), with a small amount of necrosis observed on pathology on day 24. In conclusion, SSWE was able to measure changes in the stiffness of tumors in response to anti-cancer treatment. However, stiffness changes associated with good response to treatment may change over time, and increased stiffness may also reflect therapy efficacy.

HDAC inhibition does not induce estrogen receptor in human triple-negative breast cancer cell lines and patient-derived xenografts.

Several publications have suggested that histone deacetylase inhibitors (HDACis) could reverse the repression of estrogen receptor alpha (ERα) in triple-negative breast cancer (TNBC) cell lines, leading to the induction of a functional protein. Using different HDACis, vorinostat, panobinostat, and abexinostat, we therefore investigated this hypothesis in various human TNBC cell lines and patient-derived xenografts (PDXs). We used three human TNBC cell lines and three PDXs. We analyzed the in vitro toxicity of the compounds, their effects on the hormone receptors and hormone-related genes and protein expression both in vitro and in vivo models. We then explored intra-tumor histone H3 acetylation under abexinostat in xenograft models. Despite major cytotoxicity of all tested HDAC inhibitors and repression of deactylation-dependent CCND1 gene, neither ERα nor ERß, ESR1 or ESR2 genes respectively, were re-expressed in vitro. In vivo, after administration of abexinostat for three consecutive days, we did not observe any induction of ESR1 or ESR1-related genes and ERα protein expression by RT-qPCR and immunohistochemical methods in PDXs. This observation was concomitant to the fact that in vivo administration of abexinostat increased intra-tumor histone H3 acetylation. These observations do not allow us to confirm previous studies which suggested that HDACis are able to convert ER-negative (ER-) tumors to ER-positive (ER+) tumors, and that a combination of HDAC inhibitors and hormone therapy could be proposed in the management of TNBC patients.

Establishment of an Orthotopic Xenograft Mice Model of Retinoblastoma Suitable for Preclinical Testing.

Retinoblastoma is a rare cancer that occurs during childhood. The goal of current and future therapeutic strategies is to conserve the eye and visual function without using external beam radiotherapy, which is known to increase the risk of secondary cancers in genetically predisposed patients. Multimodality therapy (usually intravenous but also intra-arterial and intravitreal chemotherapy, transpupillary thermotherapy, cryotherapy, or brachytherapy) has recently improved the eye salvage rate in retinoblastoma and has led to a decreased need for external beam radiotherapy. However, the treatment of advanced intraocular retinoblastoma remains a real challenge, especially in cases of vitreous and subretinal seeding. There is a need for alternative and less toxic therapies as well as for better ways to administer the drugs. Animal models are an integral part of preclinical research in the field of oncology. This paper describes the different xenograft rodent models published in the literature so far. We will also describe a new orthotopic xenografted retinoblastoma model in immunodeficient mice, which is suitable for preclinical assays. The xenograft model was established from tumor tissue obtained directly from surgical samples and closely mimics human retinoblastoma.

Acquired resistance to endocrine treatments is associated with tumor-specific molecular changes in patient-derived luminal breast cancer xenografts.

PURPOSE: Patients with luminal breast cancer (LBC) often become endocrine resistant over time. We investigated the molecular changes associated with acquired hormonoresistances in patient-derived xenografts of LBC. EXPERIMENTAL DESIGN: Two LBC xenografts (HBCx22 and HBCx34) were treated with different endocrine treatments (ET) to obtain xenografts with acquired resistances to tamoxifen (TamR) and ovariectomy (OvaR). PI3K pathway activation was analyzed by Western blot analysis and IHC and responses to ET combined to everolimus were investigated in vivo. Gene expression analyses were performed by RT-PCR and Affymetrix arrays. RESULTS: HBCx22 TamR xenograft was cross-resistant to several hormonotherapies, whereas HBCx22 OvaR and HBCx34 TamR exhibited a treatment-specific resistance profile. PI3K pathway was similarly activated in parental and resistant xenografts but the addition of everolimus did not restore the response to tamoxifen in TamR xenografts. In contrast, the combination of fulvestrant and everolimus induced tumor regression in vivo in HBCx34 TamR, where we found a cross-talk between the estrogen receptor (ER) and PI3K pathways. Expression of several ER-controlled genes and ER coregulators was significantly changed in both TamR and OvaR tumors, indicating impaired ER transcriptional activity. Expression changes associated with hormonoresistance were both tumor and treatment specific and were enriched for genes involved in cell growth, cell death, and cell survival. CONCLUSIONS: PDX models of LBC with acquired resistance to endocrine therapies show a great diversity of resistance phenotype, associated with specific deregulations of ER-mediated gene transcription. These models offer a tool for developing anticancer therapies and to investigate the dynamics of resistance emerging during pharmacologic interventions. Clin Cancer Res; 20(16); 4314-25. ©2014 AACR.

Inhibiting aurora kinases reduces tumor growth and suppresses tumor recurrence after chemotherapy in patient-derived triple-negative breast cancer xenografts.

Triple-negative breast cancers (TNBC) have an aggressive phenotype with a relatively high rate of recurrence and poor overall survival. To date, there is no approved targeted therapy for TNBCs. Aurora kinases act as regulators of mammalian cell division. They are important for cell-cycle progression and are frequently overexpressed or mutated in human tumors, including breast cancer. In this study, we investigated the therapeutic potential of targeting Aurora kinases in preclinical models of human breast cancers using a pan-inhibitor of Aurora kinases, AS703569. In vitro, AS703569 was tested in 15 human breast cancer cell lines. TNBC cell lines were more sensitive to AS703569 than were other types of breast cancer cells. Inhibition of proliferation was associated with cell-cycle arrest, aneuploidy, and apoptosis. In vivo, AS703569 administered alone significantly inhibited tumor growth in seven of 11 patient-derived breast cancer xenografts. Treatment with AS703569 was associated with a decrease of phospho-histone H3 expression. Finally, AS703569 combined to doxorubicin-cyclophosphamide significantly inhibited in vivo tumor recurrence, suggesting that Aurora kinase inhibitors could be used both in monotherapy and in combination settings. In conclusion, these data indicate that targeting Aurora kinases could represent a new effective approach for TNBC treatment.

Pyrophosphorolysis-activated polymerization detects circulating tumor DNA in metastatic uveal melanoma.

PURPOSE: To develop a molecular tool to detect circulating tumor-derived DNA (ctDNA) in the plasma from patients with uveal melanoma as a marker of tumor burden and monitor treatment efficacy. EXPERIMENTAL DESIGN: A real-time PCR was developed on the basis of bidirectional pyrophosphorolysis-activated polymerization (bi-PAP) for the quantification of ctDNA using 3'blocked primer pairs specific for the 3 recurrent mutually exclusive mutations of Gα subunits GNAQ and GNA11. RESULTS: Sensitivity and specificity of bi-PAP were assessed on serial dilutions of tumor DNA in normal DNA for the 3 recurrent mutations. Each assay could detect a single mutated molecule per reaction, whereas 10(4) copies of normal DNA were not detected. The ctDNA was readily detected in plasma of mice bearing uveal melanoma xenografts in amounts proportional to circulating human DNA. Finally, plasma was almost always found positive (20 of 21 tested patients) in a prospective analysis of patients with metastatic uveal melanoma. CONCLUSIONS: Bi-PAP assays detect and quantify ctDNA in patients with metastatic uveal melanoma. A prospective study is ongoing to assess the clinical usefulness of ctDNA level in uveal melanoma.