Publications by authors named "Todd W Miller"

64 Publications

Pan-Cancer Transcriptional Models Predicting Chemosensitivity in Human Tumors.

Cancer Inform 2021 19;20:11769351211002494. Epub 2021 Mar 19.

Department of Molecular & Systems Biology, Norris Cotton Cancer Center, Dartmouth-Hitchcock Medical Center, Geisel School of Medicine at Dartmouth, Lebanon, NH, USA.

Motivation: Despite increasing understanding of the molecular characteristics of cancer, chemotherapy success rates remain low for many cancer types. Studies have attempted to identify patient and tumor characteristics that predict sensitivity or resistance to different types of conventional chemotherapies, yet a concise model that predicts chemosensitivity based on gene expression profiles across cancer types remains to be formulated. We attempted to generate pan-cancer models predictive of chemosensitivity and chemoresistance. Such models may increase the likelihood of identifying the type of chemotherapy most likely to be effective for a given patient based on the overall gene expression of their tumor.

Results: Gene expression and drug sensitivity data from solid tumor cell lines were used to build predictive models for 11 individual chemotherapy drugs. Models were validated using datasets from solid tumors from patients. For all drug models, accuracy ranged from 0.81 to 0.93 when applied to all relevant cancer types in the testing dataset. When considering how well the models predicted chemosensitivity or chemoresistance within individual cancer types in the testing dataset, accuracy was as high as 0.98. Cell line-derived pan-cancer models were able to statistically significantly predict sensitivity in human tumors in some instances; for example, a pan-cancer model predicting sensitivity in patients with bladder cancer treated with cisplatin was able to significantly segregate sensitive and resistant patients based on recurrence-free survival times ( = .048) and in patients with pancreatic cancer treated with gemcitabine ( = .038). These models can predict chemosensitivity and chemoresistance across cancer types with clinically useful levels of accuracy.
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http://dx.doi.org/10.1177/11769351211002494DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7983245PMC
March 2021

Changes in Peripheral and Local Tumor Immunity after Neoadjuvant Chemotherapy Reshape Clinical Outcomes in Patients with Breast Cancer.

Clin Cancer Res 2020 Nov 21;26(21):5668-5681. Epub 2020 Aug 21.

Department of Medicine, Vanderbilt University Medical Center, Nashville, Tennessee.

Purpose: The recent approval of anti-programmed death-ligand 1 immunotherapy in combination with nab-paclitaxel for metastatic triple-negative breast cancer (TNBC) highlights the need to understand the role of chemotherapy in modulating the tumor immune microenvironment (TIME).

Experimental Design: We examined immune-related gene expression patterns before and after neoadjuvant chemotherapy (NAC) in a series of 83 breast tumors, including 44 TNBCs, from patients with residual disease (RD). Changes in gene expression patterns in the TIME were tested for association with recurrence-free (RFS) and overall survival (OS). In addition, we sought to characterize the systemic effects of NAC through single-cell analysis (RNAseq and cytokine secretion) of programmed death-1-high (PD-1) CD8 peripheral T cells and examination of a cytolytic gene signature in whole blood.

Results: In non-TNBC, no change in expression of any single gene was associated with RFS or OS, while in TNBC upregulation of multiple immune-related genes and gene sets were associated with improved long-term outcome. High cytotoxic T-cell signatures present in the peripheral blood of patients with breast cancer at surgery were associated with persistent disease and recurrence, suggesting active antitumor immunity that may indicate ongoing disease burden.

Conclusions: We have characterized the effects of NAC on the TIME, finding that TNBC is uniquely sensitive to the immunologic effects of NAC, and local increases in immune genes/sets are associated with improved outcomes. However, expression of cytotoxic genes in the peripheral blood, as opposed to the TIME, may be a minimally invasive biomarker of persistent micrometastatic disease ultimately leading to recurrence.
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http://dx.doi.org/10.1158/1078-0432.CCR-19-3685DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7642197PMC
November 2020

Identification of Let-7f-5p as a novel biomarker of recurrence in non-muscle invasive bladder cancer.

Cancer Biomark 2020 ;29(1):101-110

Norris Cotton Cancer Center, Geisel School of Medicine at Dartmouth, Lebanon, NH, USA.

Background: Among patients diagnosed with non-muscle invasive bladder cancer (NMIBC), 30% to 70% experience recurrences within 6 to 12 years of diagnosis. The need to screen for these events every 3 to 6 months and ultimately annually by cystoscopy makes bladder cancer one of the most expensive malignancies to manage.

Objective: The purpose of this study was to identify reproducible prognostic microRNAs in resected non-muscle invasive bladder tumor tissue that are predictive of the recurrent tumor phenotype as potential biomarkers and molecular therapeutic targets.

Methods: Two independent cohorts of NMIBC patients were analyzed using a biomarker discovery and validation approach, respectively.

Results: miRNA Let-7f-5p showed the strongest association with recurrence across both cohorts. Let-7f-5p levels in urine and plasma were both found to be significantly correlated with levels in tumor tissue. We assessed the therapeutic potential of targeting Lin28, a negative regulator of Let-7f-5p, with small-molecule inhibitor C1632. Lin28 inhibition significantly increased levels of Let-7f-5p expression and led to significant inhibition of viability and migration of HTB-2 cells.

Conclusions: We have identified Let-7f-5p as a miRNA biomarker of recurrence in NMIBC tumors. We further demonstrate that targeting Lin28, a negative regulator of Let-7f-5p, represents a novel potential therapeutic opportunity in NMIBC.
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http://dx.doi.org/10.3233/CBM-191322DOI Listing
January 2020

Molecular matching and treatment strategies for advanced stage lung cancer at Dartmouth-Hitchcock Medical Center: A three-year review of a Molecular Tumor Board.

Pract Lab Med 2020 Aug 12;21:e00174. Epub 2020 Jun 12.

Department of Pathology and Laboratory Medicine, Geisel School of Medicine at Dartmouth, Dartmouth-Hitchcock Medical Center, Lebanon, NH, USA.

Matching of actionable tumor mutations with targeted therapy increases response rates and prolongs survival in lung cancer patients. Drug development and trials targeting genetic alterations are expanding rapidly. We describe the role of a Molecular Tumor Board (MTB) in the design of molecularly informed treatment strategies in our lung cancer patient population. Tumor DNA was sequenced using a 50-gene targeted next-generation sequencing panel. Cases were evaluated by a multidisciplinary MTB who suggested a course of treatment based on each patient's molecular findings. During a three-year period, 21 lung cancer patients were presented at the MTB. All patients lacked common activating mutations and rearrangements. One patient had Stage IIIb disease; all others were Stage IV; 18 patients had received ≥1 prior line of therapy (range 0-5). Suggestions for treatment with a targeted therapy were made for 19/21 (90.5%) patients, and four patients (21%) underwent treatment with a targeted agent, two as part of a clinical trial. Identified barriers to treatment with targeted therapy included: ineligibility for clinical trials (n ​= ​2), lack of interest in study/distance to travel (n ​= ​2), lack of disease progression (n ​= ​2), poor performance status (n ​= ​5), decision to treat next with immunotherapy (n ​= ​3), and unknown (n ​= ​1). For the majority of lung cancer patients, the MTB provided recommendations based on tumor genetic profiles. Identified barriers to treatment suggest that presentation to the MTB at earlier stages of disease may increase the number of patients eligible for treatment with a genetically informed targeted agent.
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http://dx.doi.org/10.1016/j.plabm.2020.e00174DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7322356PMC
August 2020

Plasma DNA as a "liquid biopsy" incompletely complements tumor biopsy for identification of mutations in a case series of four patients with oligometastatic breast cancer.

Breast Cancer Res Treat 2020 Aug 19;182(3):665-677. Epub 2020 Jun 19.

Comprehensive Breast Program, Norris Cotton Cancer Center, Geisel School of Medicine at Dartmouth, Lebanon, NH, USA.

Purpose: Circulating tumor DNA in plasma may present a minimally invasive opportunity to identify tumor-derived mutations to inform selection of targeted therapies for individual patients, particularly in cases of oligometastatic disease where biopsy of multiple tumors is impractical. To assess the utility of plasma DNA as a "liquid biopsy" for precision oncology, we tested whether sequencing of plasma DNA is a reliable surrogate for sequencing of tumor DNA to identify targetable genetic alterations.

Methods: Blood and biopsies of 1-3 tumors were obtained from 4 evaluable patients with advanced breast cancer. One patient provided samples from an additional 7 tumors post-mortem. DNA extracted from plasma, tumor tissues, and buffy coat of blood were used for probe-directed capture of all exons in 149 cancer-related genes and massively parallel sequencing. Somatic mutations in DNA from plasma and tumors were identified by comparison to buffy coat DNA.

Results: Sequencing of plasma DNA identified 27.94 ± 11.81% (mean ± SD) of mutations detected in a tumor(s) from the same patient; such mutations tended to be present at high allelic frequency. The majority of mutations found in plasma DNA were not found in tumor samples. Mutations were also found in plasma that matched clinically undetectable tumors found post-mortem.

Conclusions: The incomplete overlap of genetic alteration profiles of plasma and tumors warrants caution in the sole reliance of plasma DNA to identify therapeutically targetable alterations in patients and indicates that analysis of plasma DNA complements, but does not replace, tumor DNA profiling.

Trial Registration: Subjects were prospectively enrolled in trial NCT01836640 (registered April 22, 2013).
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http://dx.doi.org/10.1007/s10549-020-05714-2DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7375191PMC
August 2020

AMPK Activation by Metformin Promotes Survival of Dormant ER Breast Cancer Cells.

Clin Cancer Res 2020 07 22;26(14):3707-3719. Epub 2020 Apr 22.

Department of Molecular & Systems Biology, Norris Cotton Cancer Center, Geisel School of Medicine at Dartmouth, Lebanon, New Hampshire.

Purpose: Despite adjuvant endocrine therapy for patients with estrogen receptor alpha (ER)-positive breast cancer, dormant residual disease can persist for years and eventually cause tumor recurrence. We sought to deduce mechanisms underlying the persistence of dormant cancer cells to identify therapeutic strategies.

Experimental Design: Mimicking the aromatase inhibitor-induced depletion of estrogen levels used to treat patients, we developed preclinical models of dormancy in ER breast cancer induced by estrogen withdrawal in mice. We analyzed tumor xenografts and cultured cancer cells for molecular and cellular responses to estrogen withdrawal and drug treatments. Publicly available clinical breast tumor gene expression datasets were analyzed for responses to neoadjuvant endocrine therapy.

Results: Dormant breast cancer cells exhibited upregulated 5' adenosine monophosphate-activated protein kinase (AMPK) levels and activity, and upregulated fatty acid oxidation. While the antidiabetes AMPK-activating drug metformin slowed the estrogen-driven growth of cells and tumors, metformin promoted the persistence of estrogen-deprived cells and tumors through increased mitochondrial respiration driven by fatty acid oxidation. Pharmacologic or genetic inhibition of AMPK or fatty acid oxidation promoted clearance of dormant residual disease, while dietary fat increased tumor cell survival.

Conclusions: AMPK has context-dependent effects in cancer, cautioning against the widespread use of an AMPK activator across disease settings. The development of therapeutics targeting fat metabolism is warranted in ER breast cancer.
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http://dx.doi.org/10.1158/1078-0432.CCR-20-0269DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7367755PMC
July 2020

Endocytosis of very low-density lipoproteins: an unexpected mechanism for lipid acquisition by breast cancer cells.

J Lipid Res 2020 02 5;61(2):205-218. Epub 2019 Dec 5.

Norris Cotton Cancer Center, Dartmouth-Hitchcock Medical Center, Lebanon, NH

We previously described the expression of CD36 and LPL by breast cancer (BC) cells and tissues and the growth-promoting effect of VLDL observed only in the presence of LPL. We now report a model in which LPL is bound to a heparan sulfate proteoglycan motif on the BC cell surface and acts in concert with the VLDL receptor to internalize VLDLs via receptor-mediated endocytosis. We also demonstrate that gene-expression programs for lipid synthesis versus uptake respond robustly to triglyceride-rich lipoprotein availability. The literature emphasizes de novo FA synthesis and exogenous free FA uptake using CD36 as paramount mechanisms for lipid acquisition by cancer cells. We find that the uptake of intact lipoproteins is also an important mechanism for lipid acquisition and that the relative reliance on lipid synthesis versus uptake varies among BC cell lines and in response to VLDL availability. This metabolic plasticity has important implications for the development of therapies aimed at the lipid dependence of many types of cancer, in that the inhibition of FA synthesis may elicit compensatory upregulation of lipid uptake. Moreover, the mechanism that we have elucidated provides a direct connection between dietary fat and tumor biology.-.
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http://dx.doi.org/10.1194/jlr.RA119000327DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6997602PMC
February 2020

Statistical determination of synergy based on Bliss definition of drugs independence.

PLoS One 2019 25;14(11):e0224137. Epub 2019 Nov 25.

Molecular & Systems Biology, Geisel School of Medicine at Dartmouth, Hanover, New Hampshire, United States of America.

Although synergy is a pillar of modern pharmacology, toxicology, and medicine, there is no consensus on its definition despite its nearly one hundred-year history. Moreover, methods for statistical determination of synergy that account for variation of response to treatment are underdeveloped and if exist are reduced to the traditional t-test, but do not comply with the normal distribution assumption. We offer statistical models for estimation of synergy using an established definition of Bliss drugs' independence. Although Bliss definition is well-known, it remains a theoretical concept and has never been applied for statistical determination of synergy with various forms of treatment outcome. We rigorously and consistently extend the Bliss definition to detect statistically significant synergy under various designs: (1) in vitro, when the outcome of a cell culture experiment with replicates is the proportion of surviving cells for a single dose or multiple doses, (2) dose-response methodology, (3) in vivo studies in organisms, when the outcome is a longitudinal measurement such as tumor volume, and (4) clinical studies, when the outcome of treatment is measured by survival. For each design, we developed a specific statistical model and demonstrated how to test for independence, synergy, and antagonism, and compute the associated p-value.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0224137PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6876842PMC
April 2020

Integrated pan-cancer gene expression and drug sensitivity analysis reveals SLFN11 mRNA as a solid tumor biomarker predictive of sensitivity to DNA-damaging chemotherapy.

PLoS One 2019 4;14(11):e0224267. Epub 2019 Nov 4.

Department of Molecular & Systems Biology, Geisel School of Medicine at Dartmouth, Lebanon, New Hampshire, United States of America.

Background: Precision oncology seeks to integrate multiple layers of data from a patient's cancer to effectively tailor therapy. Conventional chemotherapies are sometimes effective but accompanied by adverse events, warranting the identification of a biomarker of chemosensitivity.

Objective: Identify an mRNA biomarker that predicts chemosensitivity across solid tumor subtypes.

Methods: We performed a pan-solid tumor analysis integrating gene expression and drug sensitivity profiles from 3 cancer cell line datasets to identify transcripts correlated with sensitivity to a panel of chemotherapeutics. We then tested the ability of an mRNA biomarker to predictive clinical outcomes in cohorts of patients with breast, lung, or ovarian cancer.

Results: Expression levels of several mRNA transcripts were significantly correlated with sensitivity or resistance chemotherapeutics in cancer cell line datasets. The only mRNA transcript significantly correlated with sensitization to multiple classes of DNA-damaging chemotherapeutics in all 3 cell line datasets was encoded by Schlafen Family Member 11 (SLFN11). Analyses of multiple breast, lung, and ovarian cancer patient cohorts treated with chemotherapy confirmed SLFN11 mRNA expression as a predictive biomarker of longer overall survival and improved tumor response.

Conclusions: Tumor SLFN11 mRNA expression is a biomarker of sensitivity to an array of DNA-damaging chemotherapeutics across solid tumor subtypes.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0224267PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6827986PMC
April 2020

A Transcriptionally Definable Subgroup of Triple-Negative Breast and Ovarian Cancer Samples Shows Sensitivity to HSP90 Inhibition.

Clin Cancer Res 2020 01 26;26(1):159-170. Epub 2019 Sep 26.

Department of Molecular & Systems Biology, Norris Cotton Cancer Center, Geisel School of Medicine at Dartmouth, Lebanon, New Hampshire.

Purpose: We hypothesized that integrated analysis of cancer types from different lineages would reveal novel molecularly defined subgroups with unique therapeutic vulnerabilities. On the basis of the molecular similarities between subgroups of breast and ovarian cancers, we analyzed these cancers as a single cohort to test our hypothesis.

Experimental Design: Identification of transcriptional subgroups of cancers and drug sensitivity analyses were performed using mined data. Cell line sensitivity to Hsp90 inhibitors (Hsp90i) was tested . The ability of a transcriptional signature to predict Hsp90i sensitivity was validated using cell lines, and cell line- and patient-derived xenograft (PDX) models. Mechanisms of Hsp90i sensitivity were uncovered using immunoblot and RNAi.

Results: Transcriptomic analyses of breast and ovarian cancer cell lines uncovered two mixed subgroups comprised primarily of triple-negative breast and multiple ovarian cancer subtypes. Drug sensitivity analyses revealed that cells of one mixed subgroup are significantly more sensitive to Hsp90i compared with cells from all other cancer lineages evaluated. A gene expression classifier was generated that predicted Hsp90i sensitivity , and in cell line- and PDXs. Cells from the Hsp90i-sensitive subgroup underwent apoptosis mediated by Hsp90i-induced upregulation of the proapoptotic proteins Bim and PUMA.

Conclusions: Our findings identify Hsp90i as a potential therapeutic strategy for a transcriptionally defined subgroup of ovarian and breast cancers. This study demonstrates that gene expression profiles may be useful to identify therapeutic vulnerabilities in tumor types with limited targetable genetic alterations, and to identify molecularly definable cancer subgroups that transcend lineage.
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http://dx.doi.org/10.1158/1078-0432.CCR-18-2213DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6942625PMC
January 2020

Tumor pharmacokinetics and pharmacodynamics of the CDK4/6 inhibitor ribociclib in patients with recurrent glioblastoma.

J Neurooncol 2019 Sep 9;144(3):563-572. Epub 2019 Aug 9.

Department of Neurology, Division of Neuro-Oncology, University of Virginia Health System, P.O. Box 800432, Charlottesville, VA, 22908, USA.

Introduction: We conducted a phase Ib study (NCT02345824) to determine whether ribociclib, an inhibitor of cyclin-dependent kinases 4 and 6 (CDK4/6), penetrates tumor tissue and modulates downstream signaling pathways including retinoblastoma protein (Rb) in patients with recurrent glioblastoma (GBM).

Methods: Study participants received ribociclib (600 mg QD) for 8-21 days before surgical resection of their recurrent GBM. Total and unbound concentrations of ribociclib were measured in samples of tumor tissue, plasma, and cerebrospinal fluid (CSF). We analyzed tumor specimens obtained from the first (initial/pre-study) and second (recurrent/on-study) surgery by immunohistochemistry for Rb status and downstream signaling of CDK4/6 inhibition. Participants with Rb-positive recurrent tumors continued ribociclib treatment on a 21-day-on, 7-day-off schedule after surgery, and were monitored for toxicity and disease progression.

Results: Three participants with recurrent Rb-positive GBM participated in this study. Mean unbound (pharmacologically active) ribociclib concentrations in plasma, CSF, MRI-enhancing, MRI-non-enhancing, and tumor core regions were 0.337 μM, 0.632 μM, 1.242 nmol/g, 0.484 nmol/g, and 1.526 nmol/g, respectively, which exceeded the in vitro IC (0.04 μM) for inhibition of CDK4/6 in cell-free assay. Modulation of pharmacodynamic markers of ribociclib CDK 4/6 inhibition in tumor tissues were inconsistent between study participants. No participants experienced serious adverse events, but all experienced early disease progression.

Conclusions: This study suggests that ribociclib penetrated recurrent GBM tissue at concentrations predicted to be therapeutically beneficial. Our study was unable to demonstrate tumor pharmacodynamic correlates of drug activity. Although well tolerated, ribociclib monotherapy seemed ineffective for the treatment of recurrent GBM.
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http://dx.doi.org/10.1007/s11060-019-03258-0DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6863163PMC
September 2019

Estrogen therapy induces an unfolded protein response to drive cell death in ER+ breast cancer.

Mol Oncol 2019 08 9;13(8):1778-1794. Epub 2019 Jul 9.

Department of Molecular & Systems Biology, Norris Cotton Cancer Center, Geisel School of Medicine at Dartmouth, Lebanon, NH, USA.

Estrogens have been shown to elicit anticancer effects against estrogen receptor α (ER)-positive breast cancer. We sought to determine the mechanism underlying the therapeutic response. Response to 17β-estradiol was assessed in ER+ breast cancer models with resistance to estrogen deprivation: WHIM16 patient-derived xenografts, C7-2-HI and C4-HI murine mammary adenocarcinomas, and long-term estrogen-deprived MCF-7 cells. As another means to reactivate ER, the anti-estrogen fulvestrant was withdrawn from fulvestrant-resistant MCF-7 cells. Transcriptional, growth, apoptosis, and molecular alterations in response to ER reactivation were measured. 17β-estradiol treatment and fulvestrant withdrawal induced transcriptional activation of ER, and cells adapted to estrogen deprivation or fulvestrant were hypersensitive to 17β-estradiol. ER transcriptional response was followed by an unfolded protein response and apoptosis. Such apoptosis was dependent upon the unfolded protein response, p53, and JNK signaling. Anticancer effects were most pronounced in models exhibiting genomic amplification of the gene encoding ER (ESR1), suggesting that engagement of ER at high levels is cytotoxic. These data indicate that long-term adaptation to estrogen deprivation or ER inhibition alters sensitivity to ER reactivation. In such adapted cells, 17β-estradiol treatment and anti-estrogen withdrawal hyperactivate ER, which drives an unfolded protein response and subsequent growth inhibition and apoptosis. 17β-estradiol treatment should be considered as a therapeutic option for anti-estrogen-resistant disease, particularly in patients with tumors harboring ESR1 amplification or ER overexpression. Furthermore, therapeutic strategies that enhance an unfolded protein response may increase the therapeutic effects of ER reactivation.
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http://dx.doi.org/10.1002/1878-0261.12528DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6670014PMC
August 2019

Molecular and epigenetic profiles of BRCA1-like hormone-receptor-positive breast tumors identified with development and application of a copy-number-based classifier.

Breast Cancer Res 2019 01 25;21(1):14. Epub 2019 Jan 25.

Department of Epidemiology, Lebanon, USA.

Background: BRCA1-mutated cancers exhibit deficient homologous recombination (HR) DNA repair, resulting in extensive copy number alterations and genome instability. HR deficiency can also arise in tumors without a BRCA1 mutation. Compared with other breast tumors, HR-deficient, BRCA1-like tumors exhibit worse prognosis but selective chemotherapeutic sensitivity. Presently, patients with triple negative breast cancer (TNBC) who do not respond to hormone endocrine-targeting therapy are given cytotoxic chemotherapy. However, more recent evidence showed a similar genomic profile between BRCA1-deficient TNBCs and hormone-receptor-positive tumors. Characterization of the somatic alterations of BRCA1-like hormone-receptor-positive breast tumors as a group, which is currently lacking, can potentially help develop biomarkers for identifying additional patients who might respond to chemotherapy.

Methods: We retrained and validated a copy-number-based support vector machine (SVM) classifier to identify HR-deficient, BRCA1-like breast tumors. We applied this classifier to The Cancer Genome Atlas (TCGA) and Molecular Taxonomy of Breast Cancer International Consortium (METABRIC) breast tumors. We assessed mutational profiles and proliferative capacity by covariate-adjusted linear models and identified differentially methylated regions using DMRcate in BRCA1-like hormone-receptor-positive tumors.

Results: Of the breast tumors in TCGA and METABRIC, 22% (651/2925) were BRCA1-like. Stratifying on hormone-receptor status, 13% (302/2405) receptor-positive and 69% (288/417) triple-negative tumors were BRCA1-like. Among the hormone-receptor-positive subgroup, BRCA1-like tumors showed significantly increased mutational burden and proliferative capacity (both P < 0.05). Genome-scale DNA methylation analysis of BRCA1-like tumors identified 202 differentially methylated gene regions, including hypermethylated BRCA1. Individually significant CpGs were enriched for enhancer regions (P < 0.05). The hypermethylated gene sets were enriched for DNA and chromatin conformation (all Bonferroni P < 0.05).

Conclusions: To provide insights into alternative classification and potential therapeutic targeting strategies of BRCA1-like hormone-receptor-positive tumors we developed and applied a novel copy number classifier to identify BRCA1-like hormone-receptor-positive tumors and their characteristic somatic alteration profiles.
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http://dx.doi.org/10.1186/s13058-018-1090-zDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6347811PMC
January 2019

Ductal Carcinoma in Situ Biomarkers in a Precision Medicine Era: Current and Future Molecular-Based Testing.

Am J Pathol 2019 05 29;189(5):956-965. Epub 2018 Oct 29.

Department of Pathology and Laboratory Medicine, Dartmouth Hitchcock Medical Center, Lebanon, New Hampshire. Electronic address:

Historically, ductal carcinoma in situ (DCIS) of the breast has been managed aggressively with surgery and radiotherapy because of a risk of progression to invasive ductal carcinoma. However, this treatment paradigm has been challenged by overtreatment concerns and evidence that suggests that DCIS can be stratified according to risk of recurrence or risk of progression to invasive disease. Traditional methods of risk stratification include histologic grade and hormone receptor status. Recent technological advancements have enabled an era of precision medicine, where DCIS can be molecularly analyzed by tools, such as next-generation DNA and RNA sequencing, to identify molecular biomarkers for risk stratification. These findings have led to the development of tools such as the Oncotype DX Breast DCIS Score, a gene expression-based assay with the potential to prevent overtreatment in low-risk disease.
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http://dx.doi.org/10.1016/j.ajpath.2018.08.020DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6521891PMC
May 2019

Cytokine sensitivity screening highlights BMP4 pathway signaling as a therapeutic opportunity in ER breast cancer.

FASEB J 2019 02 30;33(2):1644-1657. Epub 2018 Aug 30.

Department of Molecular and Systems Biology, Norris Cotton Cancer Center, Geisel School of Medicine at Dartmouth, Lebanon, New Hampshire, USA.

Despite the success of approved systemic therapies for estrogen receptor α (ER)-positive breast cancer, drug resistance remains common. We hypothesized that secreted factors from the human tumor microenvironment could modulate drug resistance. We previously screened a library of 297 recombinant-secreted microenvironmental proteins for the ability to confer resistance to the anti-estrogen fulvestrant in 2 ER breast cancer cell lines. Herein, we considered whether factors that enhanced drug sensitivity could be repurposed as therapeutics and provide leads for drug development. Screening data revealed bone morphogenic protein (BMP)4 as a factor that inhibited cell growth and synergized with approved anti-estrogens and cyclin-dependent kinase 4/6 inhibitors (CDK4/6). BMP4-mediated growth inhibition was dependent on type I receptor activin receptor-like kinase (ALK)3-dependent phosphorylation (P) of mothers against decapentaplegic homolog (SMAD/P-SMAD)1 and 5, which could be reversed by BMP receptor inhibitors and ALK3 knockdown. The primary effect of BMP4 on cell fate was cell-cycle arrest, in which RNA sequencing, immunoblot analysis, and RNA interference revealed to be dependent on p21 upregulation. BMP4 also enhanced sensitivity to approved inhibitors of mammalian target of rapamycin complex 1 and CDK4/6 via ALK3-mediated P-SMAD1/5 and p21 upregulation in anti-estrogen-resistant cells. Patients bearing primary ER breast tumors, exhibiting a transcriptomic signature of BMP4 signaling, had improved disease outcome following adjuvant treatment with anti-estrogen therapy, independently of age, tumor grade, and tumor stage. Furthermore, a transcriptomic signature of BMP4 signaling was predictive of an improved biologic response to the CDK4/6 palbociclib, in combination with an aromatase inhibitor in primary tumors. These findings highlight BMP4 and its downstream pathway activation as a therapeutic opportunity in ER breast cancer.-Shee, K., Jiang, A., Varn, F. S., Liu, S., Traphagen, N. A., Owens, P., Ma, C. X., Hoog, J., Cheng, C., Golub, T. R., Straussman, R., Miller, T. W. Cytokine sensitivity screening highlights BMP4 pathway signaling as a therapeutic opportunity in ER breast cancer.
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http://dx.doi.org/10.1096/fj.201801241RDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6338642PMC
February 2019

Estrogen receptor alpha drives mTORC1 inhibitor-induced feedback activation of PI3K/AKT in ER+ breast cancer.

Oncotarget 2018 Feb 15;9(10):8810-8822. Epub 2018 Jan 15.

Department of Molecular and Systems Biology, Norris Cotton Cancer Center, Dartmouth-Hitchcock Medical Center, Geisel School of Medicine at Dartmouth, Lebanon, NH, USA.

The mTORC1 inhibitor RAD001 (everolimus) is approved for treatment of recurrent/metastatic estrogen receptor (ER)-positive breast cancer in combination with the aromatase inhibitor (AI) exemestane. The benefits of A) continued anti-estrogen therapy for anti-estrogen-resistant disease in the context of mTORC1 inhibition, and B) adjuvant everolimus in combination with anti-estrogen therapy for early-stage disease are being tested clinically, but molecular rationale remains unclear. We hypothesized that mTORC1 inhibition activates the IGF-1R/InsR/IRS-1/2 axis in an ER-dependent manner to drive PI3K/AKT and promote cancer cell survival, implicating ER in survival signaling induced by mTORC1 inhibition. Anti-estrogen treatment synergized with RAD001 to inhibit ER+ breast cancer cell growth. Inhibition of ER, IGF-1R/InsR, or IRS-1/2 suppressed AKT activation induced by mTORC1 inhibition. RAD001 primed IGF-1R/InsR for activation, which was enhanced by ER signaling. Post-menopausal patients with early-stage ER+ breast cancer were treated presurgically +/- the AI letrozole. Viable tumor fragments from surgical specimens were treated with RAD001 and/or OSI-906 ; RAD001 increased AKT activation, which was abrogated by presurgical letrozole. Letrozole decreased IGF-1R and IRS-1/2 tumor levels. These data suggest that ER drives PI3K/AKT activation in response to mTORC1 inhibition, providing molecular rationale for therapeutic combinations of anti-estrogens and mTORC1 inhibitors in endocrine-sensitive disease.
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http://dx.doi.org/10.18632/oncotarget.24256DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5823630PMC
February 2018

Coamplification of protects -amplified breast cancers from targeted therapy.

Proc Natl Acad Sci U S A 2018 03 23;115(11):E2594-E2603. Epub 2018 Feb 23.

Department of Oral and Craniofacial Molecular Biology, Philips Institute for Oral Health Research, VCU School of Dentistry and Massey Cancer Center, Virginia Commonwealth University, Richmond, VA 23298;

() amplification is a driving oncogenic event in breast cancer. Clinical trials have consistently shown the benefit of HER2 inhibitors (HER2i) in treating patients with both local and advanced HER2+ breast cancer. Despite this benefit, their efficacy as single agents is limited, unlike the robust responses to other receptor tyrosine kinase inhibitors like EGFR inhibitors in -mutant lung cancer. Interestingly, the lack of HER2i efficacy occurs despite sufficient intracellular signaling shutdown following HER2i treatment. Exploring possible intrinsic causes for this lack of response, we uncovered remarkably depressed levels of NOXA, an endogenous inhibitor of the antiapoptotic MCL-1, in -amplified breast cancer. Upon investigation of the mechanism leading to low NOXA, we identified a micro-RNA encoded in an intron of , termed , that targets the mRNA of the Estrogen Receptor α (). Reduced ESR1 expression in turn prevents ERα-mediated transcription of , mitigating apoptosis following treatment with the HER2i lapatinib. Importantly, resistance can be overcome with pharmacological inhibition of MCL-1. More generally, while many cancers like -mutant lung cancer are driven by activated kinases that when drugged lead to robust monotherapeutic responses, we demonstrate that the efficacy of targeted therapies directed against oncogenes active through focal amplification may be mitigated by coamplified genes.
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http://dx.doi.org/10.1073/pnas.1717820115DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5856537PMC
March 2018

Therapeutically targeting tumor microenvironment-mediated drug resistance in estrogen receptor-positive breast cancer.

J Exp Med 2018 03 7;215(3):895-910. Epub 2018 Feb 7.

Department of Molecular and Systems Biology, Norris Cotton Cancer Center, Geisel School of Medicine at Dartmouth, Lebanon, NH

Drug resistance to approved systemic therapies in estrogen receptor-positive (ER+) breast cancer remains common. We hypothesized that factors present in the human tumor microenvironment (TME) drive drug resistance. Screening of a library of recombinant secreted microenvironmental proteins revealed fibroblast growth factor 2 (FGF2) as a potent mediator of resistance to anti-estrogens, mTORC1 inhibition, and phosphatidylinositol 3-kinase inhibition in ER+ breast cancer. Phosphoproteomic analyses identified ERK1/2 as a major output of FGF2 signaling via FGF receptors (FGFRs), with consequent up-regulation of Cyclin D1 and down-regulation of Bim as mediators of drug resistance. FGF2-driven drug resistance in anti-estrogen-sensitive and -resistant models, including patient-derived xenografts, was reverted by neutralizing FGF2 or FGFRs. A transcriptomic signature of FGF2 signaling in primary tumors predicted shorter recurrence-free survival independently of age, grade, stage, and FGFR amplification status. These findings delineate FGF2 signaling as a ligand-based drug resistance mechanism and highlights an underdeveloped aspect of precision oncology: characterizing and treating patients according to their TME constitution.
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http://dx.doi.org/10.1084/jem.20171818DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5839765PMC
March 2018

Trailblazing Precision Oncology for Rare Tumor Subtypes.

Oncologist 2018 02 20;23(2):143-144. Epub 2017 Nov 20.

Department of Molecular & Systems Biology, Lebanon, New Hampshire, USA

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http://dx.doi.org/10.1634/theoncologist.2017-0494DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5813755PMC
February 2018

Autophagy promotes escape from phosphatidylinositol 3-kinase inhibition in estrogen receptor-positive breast cancer.

FASEB J 2018 03 3;32(3):1222-1235. Epub 2018 Jan 3.

Department of Molecular and Systems Biology, Norris Cotton Cancer Center, Geisel School of Medicine at Dartmouth, Lebanon, New Hampshire, USA.

Hyperactivation of the PI3K pathway has been implicated in resistance to antiestrogen therapies in estrogen receptor α (ER)-positive breast cancer, prompting the development of therapeutic strategies to inhibit this pathway. Autophagy has tumor-promoting and -suppressing roles and has been broadly implicated in resistance to anticancer therapies, including antiestrogens. Chloroquine (CQ) is an antimalarial and amebicidal drug that inhibits autophagy in mammalian cells and human tumors. Herein, we observed that CQ inhibited proliferation and autophagy in ER breast cancer cells. PI3K inhibition with GDC-0941 (pictilisib) induced autophagy. Inhibition of autophagy using CQ or RNA interference potentiated PI3K inhibitor-induced apoptosis. Combined inhibition of PI3K and autophagy effectively induced mitochondrial membrane depolarization, which required the BH3-only proapoptotic proteins Bim and PUMA. Treatment with GDC-0941, CQ, or the combination, significantly suppressed the growth of ER breast cancer xenografts in mice. In an antiestrogen-resistant xenograft model, GDC-0941 synergized with CQ to provide partial, but durable, tumor regression. These findings warrant clinical evaluation of therapeutic strategies to target ER, PI3K, and autophagy for the treatment of ER breast cancer.-Yang, W., Hosford, S. R., Traphagen, N. A., Shee, K., Demidenko, E., Liu, S., Miller, T. W. Autophagy promotes escape from phosphatidylinositol 3-kinase inhibition in estrogen receptor-positive breast cancer.
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http://dx.doi.org/10.1096/fj.201700477RDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5892720PMC
March 2018

P-Rex1 Expression in Invasive Breast Cancer in relation to Receptor Status and Distant Metastatic Site.

Int J Breast Cancer 2017 15;2017:4537532. Epub 2017 Jun 15.

Comprehensive Breast Program, Dartmouth-Hitchcock Medical Center, Lebanon, NH, USA.

Background: Phosphatidylinositol-3,4,5-trisphosphate-dependent Rac exchange factor 1 (P-Rex1) has been implicated in cancer growth, metastasis, and response to phosphatidylinositol 3-kinase (PI3K) inhibitor therapy. The aim of this study was to determine whether P-Rex1 expression differs between primary and metastatic human breast tumors and between breast cancer subtypes.

Design: P-Rex1 expression was measured in 133 specimens by immunohistochemistry: 40 and 42 primary breast tumors from patients who did versus did not develop metastasis, respectively, and 51 breast-derived tumors from metastatic sites (36 of which had matching primary tumors available for analysis).

Results: Primary breast tumors showed significant differences in P-Rex1 expression based on receptor subtype. ER+ and HER2+ primary tumors showed higher P-Rex1 expression than primary triple-negative tumors. HER2+ metastases from all sites showed significantly higher P-Rex1 expression compared to other metastatic receptor subtypes. Solid organ (i.e., brain, lung, and liver) metastases showed higher P-Rex1 expression compared to bone metastases.

Conclusions: P-Rex1 expression is increased in ER+ and HER2+ breast cancers compared to triple-negative tumors. P-Rex1 may be differentially expressed in metastatic tumors based on site and receptor status. The role of P-Rex1 in the development of breast cancer metastases and as a predictive biomarker of therapeutic response warrants further investigation.
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http://dx.doi.org/10.1155/2017/4537532DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5494073PMC
June 2017

Therapeutic sensitivity to Rac GTPase inhibition requires consequential suppression of mTORC1, AKT, and MEK signaling in breast cancer.

Oncotarget 2017 Mar;8(13):21806-21817

Department of Molecular & Systems Biology, Norris Cotton Cancer Center, Geisel School of Medicine at Dartmouth, Lebanon, NH, USA.

Rac GTPases have oncogenic roles in cell growth, survival, and migration. We tested response to the Rac inhibitor EHT1864 in a panel of breast cancer cell lines. EHT1864-induced growth inhibition was associated with dual inhibition of the PI3K/AKT/mTORC1 and MEK/ERK pathways. Breast cancer cells harboring PIK3CA mutations or HER2 overexpression were most sensitive to Rac inhibition, suggesting that such oncogenic alterations link Rac activation with PI3K/AKT/mTORC1 and MEK/ERK signaling. Interestingly, EHT1864 decreased activation of the mTORC1 substrate p70S6K earlier than AKT inhibition, suggesting that Rac may activate mTORC1/p70S6K independently of AKT. Comparison of the growth-inhibitory profile of EHT1864 to 137 other anti-cancer drugs across 656 cancer cell lines revealed significant correlation with the p70S6K inhibitor PF-4708671. We confirmed that Rac complexes contain MEK1/2 and ERK1/2, but also contain p70S6K; these interactions were disrupted by EHT1864. Pharmacokinetic profiles revealed that EHT1864 was present in mouse plasma at concentrations effective in vitro for approximately 1 h after intraperitoneal administration. EHT1864 suppressed growth of HER2+ tumors, and enhanced response to anti-estrogen treatment in ER+ tumors. Further therapeutic development of Rac inhibitors for HER2+ and PIK3CA-mutant cancers is warranted.
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http://dx.doi.org/10.18632/oncotarget.15586DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5400625PMC
March 2017

Maximizing the Benefit-Cost Ratio of Anthracyclines in Metastatic Breast Cancer: Case Report of a Patient with a Complete Response to High-Dose Doxorubicin.

Case Rep Oncol 2016 Sep-Dec;9(3):840-846. Epub 2016 Dec 8.

Norris Cotton Cancer Center, Geisel School of Medicine at Dartmouth, Lebanon, NH, USA; Department of Hematology/Oncology, Dartmouth Hitchcock Medical Center, Lebanon, NH, USA.

Despite the clinical efficacy of anthracycline agents such as doxorubicin, dose-limiting cardiac toxicities significantly limit their long-term use. Here, we present the case of a 33-year-old female patient with extensive metastatic ER+/PR+/HER2- mucinous adenocarcinoma of the breast, who was started on doxorubicin/cyclophosphamide therapy after progressing on paclitaxel and ovarian suppressor goserelin with aromatase inhibitor exemestane. The patient was comanaged by cardiology, who carefully monitored measures of cardiac function, including EKGs, serial echocardiograms, and profiling of lipids, troponin, and pro-BNP every 2 months. The patient was treated with the cardioprotective agent dexrazoxane, and changes in cardiac markers [e.g. decreases in ejection fraction (EF)] were immediately addressed by therapeutic intervention with the ACE inhibitor lisinopril and beta-blocker metoprolol. The patient had a complete response to doxorubicin therapy, with a cumulative dose of 1,350 mg/m, which is significantly above the recommended limits, and to our knowledge, the highest dose reported in literature. Two and a half years after the last doxorubicin cycle, the patient is asymptomatic with no cardiotoxicity and an excellent quality of life. This case highlights the importance of careful monitoring and management of doxorubicin-mediated cardiotoxicity, and that higher cumulative doses of anthracyclines can be considered in patients with ongoing clinical benefit.
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http://dx.doi.org/10.1159/000453608DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5216250PMC
December 2016

Combined Inhibition of Both p110α and p110β Isoforms of Phosphatidylinositol 3-Kinase Is Required for Sustained Therapeutic Effect in PTEN-Deficient, ER Breast Cancer.

Clin Cancer Res 2017 Jun 30;23(11):2795-2805. Epub 2016 Nov 30.

Depts. of Molecular & Systems Biology, Norris Cotton Cancer Center, Geisel School of Medicine at Dartmouth, Lebanon, NH.

Determine the roles of the PI3K isoforms p110α and p110β in PTEN-deficient, estrogen receptor α (ER)-positive breast cancer, and the therapeutic potential of isoform-selective inhibitors. Anti-estrogen-sensitive and -resistant PTEN-deficient, ER human breast cancer cell lines, and mice bearing anti-estrogen-resistant xenografts were treated with the anti-estrogen fulvestrant, the p110α inhibitor BYL719, the p110β inhibitor GSK2636771, or combinations. Temporal response to growth factor receptor-initiated signaling, growth, apoptosis, predictive biomarkers, and tumor volumes were measured. p110β primed cells for response to growth factor stimulation. Although p110β inhibition suppressed cell and tumor growth, dual targeting of p110α/β enhanced apoptosis and provided sustained tumor response. The growth of anti-estrogen-sensitive cells was inhibited by fulvestrant, but fulvestrant inconsistently provided additional therapeutic effects beyond PI3K inhibition alone. Treatment-induced decreases in phosphorylation of AKT and Rb were predictive of therapeutic response. Short-term drug treatment induced tumor cell apoptosis and proliferative arrest to induce tumor regression, whereas long-term treatment only suppressed proliferation to provide durable regression. p110β is the dominant PI3K isoform in PTEN-deficient, ER breast cancer cells. Upon p110β inhibition, p110α did not induce significant reactivation of AKT, but combined targeting of p110α/β most effectively induced apoptosis and and provided durable tumor regression. Because apoptosis and tumor regression occurred early but not late in the treatment course, and proliferative arrest was maintained throughout treatment, p110α/β inhibitors may be considered short-term cytotoxic agents and long-term cytostatic agents. .
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http://dx.doi.org/10.1158/1078-0432.CCR-15-2764DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5449270PMC
June 2017

The PREX1/Rac signaling axis: Potential as a biomarker and therapeutic target in breast cancer.

Mol Cell Oncol 2015 Jul-Sep;2(3):e996016. Epub 2015 Apr 30.

Department of Pharmacology and Toxicology; Norris Cotton Cancer Center; Geisel School of Medicine at Dartmouth; Dartmouth-Hitchcock Medical Center; Lebanon, NH, USA; Comprehensive Breast Program; Norris Cotton Cancer Center; Geisel School of Medicine at Dartmouth; Dartmouth-Hitchcock Medical Center; Lebanon, NH, USA.

PREX1 is a Rac guanine exchange factor that coordinates signaling inputs from G protein-coupled receptors and receptor tyrosine kinases (RTKs). PREX1 creates a positive feedback loop to drive RTK, phosphatidylinositol 3-kinase (PI3K)/AKT, and MEK/ERK signaling. High PREX1 levels predict sensitivity to PI3K inhibitors in breast cancer cells.
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http://dx.doi.org/10.1080/23723556.2014.996016DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4905322PMC
June 2016

Clinical Implementation of Novel Targeted Therapeutics in Advanced Breast Cancer.

J Cell Biochem 2016 11 3;117(11):2454-63. Epub 2016 Jun 3.

Comprehensive Breast Program, Norris Cotton Cancer Center, Geisel School of Medicine at Dartmouth, Lebanon, New Hampshire.

The majority of advanced breast cancers have genetic alterations that are potentially targetable with drugs. Through initiatives such as The Cancer Genome Atlas (TCGA) and the International Cancer Genome Consortium (ICGC), data can be mined to provide context for next-generation sequencing (NGS) results in the landscape of advanced breast cancer. Therapies for targets other than estrogen receptor alpha (ER) and HER2, such as cyclin-dependent kinases CDK4 and CDK6, were recently approved based on efficacy in patient subpopulations, but no predictive biomarkers have been found, leaving clinicians to continue a trial-and-error approach with each patient. Next-generation sequencing identifies potentially actionable alterations in genes thought to be drivers in the cancerous process including phosphatidylinositol 3-kinase (PI3K), AKT, fibroblast growth factor receptors (FGFRs), and mutant HER2. Epigenetically directed and immunologic therapies have also shown promise for the treatment of breast cancer via histone deacetylases (HDAC) 1 and 3, programmed T cell death 1 (PD-1), and programmed T cell death ligand 1 (PD-L1). Identifying biomarkers to predict primary resistance in breast cancer will ultimately affect clinical decisions regarding adjuvant therapy in the first-line setting. However, the bulk of medical decision-making is currently made in the secondary resistance setting. Herein, we review the clinical potential of PI3K, AKT, FGFRs, mutant HER2, HDAC1/3, PD-1, and PD-L1 as therapeutic targets in breast cancer, focusing on the rationale for therapeutic development and the status of clinical testing. J. Cell. Biochem. 117: 2454-2463, 2016. © 2016 Wiley Periodicals, Inc.
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http://dx.doi.org/10.1002/jcb.25590DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6010350PMC
November 2016

Targeted next-generation sequencing detects a high frequency of potentially actionable mutations in metastatic breast cancers.

Exp Mol Pathol 2016 06 16;100(3):421-5. Epub 2016 Apr 16.

Department of Pathology and Laboratory Medicine, Dartmouth-Hitchcock Medical Center, 1 Medical Center Drive, Lebanon, NH 03766, United States. Electronic address:

Background: Metastatic breast cancer is a genetically heterogeneous disease and effective therapies for advanced stage disease are limited.

Methods: In this study, distant metastases of 22 formalin-fixed, paraffin-embedded (FFPE) breast cancer samples were sequenced using the Ion Torrent PGM and the 50 gene AmpliSeq Cancer Hotspot Panel v2 from 10ng of extracted DNA using 318 chips. Data analysis was performed with the Ion Torrent Variant Caller Plugin (hg19) and Golden Helix's SVS software for annotation and prediction of the significance of the variants.

Results: All patients were female with a median age of 61years (range 37-85years). Metastatic sites included liver (n=6, 27%), skin (n=5, 23%), brain (n=4, 18%), lymph node (n=3, 14%), lung (n=2, 9%), retroperitoneum (n=1, 4.5%), and colon (n=1, 4.5%). Overall, 28 variants in 11 genes were observed. Five (23%) samples showed no alterations and 17 (77%) showed at least one potentially biologically significant variant (BSV) defined as having FDA-approved drugs or clinical trials evaluating their significance. BSVs included mutations in the following genes: TP53 (n=8), APC (n=4), PIK3CA (n=5), MET (n=2), ERBB2 (n=2), AKT1 (n=1), CDKN2A (n=1), KRAS (n=1), and FGFR3 (n=1).

Conclusions: Potentially actionable mutations were identified in the majority of breast cancer metastases. Evaluating metastatic breast tumors using a NGS approach provides a better understanding of the mechanisms behind tumor progression and evolution and also identifies additional potentially beneficial therapeutic targets for patient management or eligibility for clinical trials.
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http://dx.doi.org/10.1016/j.yexmp.2016.04.002DOI Listing
June 2016

Strategically Timing Inhibition of Phosphatidylinositol 3-Kinase to Maximize Therapeutic Index in Estrogen Receptor Alpha-Positive, PIK3CA-Mutant Breast Cancer.

Clin Cancer Res 2016 05 5;22(9):2250-60. Epub 2016 Jan 5.

Department of Pharmacology and Toxicology, Norris Cotton Cancer Center, Geisel School of Medicine at Dartmouth, Lebanon, New Hampshire. Comprehensive Breast Program, Norris Cotton Cancer Center, Geisel School of Medicine at Dartmouth, Lebanon, New Hampshire.

Purpose: Phosphatidylinositol 3-kinase (PI3K) inhibitors are being developed for the treatment of estrogen receptor α (ER)-positive breast cancer in combination with antiestrogens. Understanding the temporal response and pharmacodynamic effects of PI3K inhibition in ER(+) breast cancer will provide a rationale for treatment scheduling to maximize therapeutic index.

Experimental Design: Antiestrogen-sensitive and antiestrogen-resistant ER(+) human breast cancer cell lines and mice bearing PIK3CA-mutant xenografts were treated with the antiestrogen fulvestrant, the PI3K inhibitor GDC-0941 (pictilisib; varied doses/schedules that provided similar amounts of drug each week), or combinations. Cell viability, signaling pathway inhibition, proliferation, apoptosis, tumor volume, and GDC-0941 concentrations in plasma and tumors were temporally measured.

Results: Treatment with the combination of fulvestrant and GDC-0941, regardless of dose/schedule, was significantly more effective than that with single-agent treatments in fulvestrant-resistant tumors. Short-term, complete PI3K inhibition blocked cell growth in vitro more effectively than chronic, incomplete inhibition. Longer-term PI3K inhibition hypersensitized cells to growth factor signaling upon drug withdrawal. Different schedules of GDC-0941 elicited similar tumor responses. While weekly high-dose GDC-0941 with fulvestrant continuously suppressed PI3K signaling for 72 hours, inducing a bolus of apoptosis and inhibiting proliferation, PI3K reactivation upon GDC-0941 washout induced a proliferative burst. Fulvestrant with daily low-dose GDC-0941 metronomically suppressed PI3K for 6 to 9 hours/day, repeatedly inducing small amounts of apoptosis and temporarily inhibiting proliferation, followed by proliferative rebound compared with fulvestrant alone.

Conclusions: Continuous and metronomic PI3K inhibition elicits robust anticancer effects in ER(+), PIK3CA-mutant breast cancer. Clinical exploration of alternate treatment schedules of PI3K inhibitors with antiestrogens is warranted. Clin Cancer Res; 22(9); 2250-60. ©2016 AACRSee related commentary by Toska and Baselga, p. 2099.
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http://dx.doi.org/10.1158/1078-0432.CCR-15-2276DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4854767PMC
May 2016

Implementation of a Molecular Tumor Board: The Impact on Treatment Decisions for 35 Patients Evaluated at Dartmouth-Hitchcock Medical Center.

Oncologist 2015 Sep 23;20(9):1011-8. Epub 2015 Jul 23.

Departments of Pathology, Community and Family Medicine, Medicine, and Pharmacology & Toxicology, Comprehensive Breast Program, and Familial Cancer Program, Norris Cotton Cancer Center, Geisel School of Medicine at Dartmouth, Dartmouth-Hitchcock Medical Center, Lebanon, New Hampshire, USA

Background: Although genetic profiling of tumors is a potentially powerful tool to predict drug sensitivity and resistance, its routine use has been limited because clinicians are often unfamiliar with interpretation and incorporation of the information into practice. We established a Molecular Tumor Board (MTB) to interpret individual patients' tumor genetic profiles and provide treatment recommendations.

Patients And Methods: DNA from tumor specimens was sequenced in a Clinical Laboratory Improvement Amendments-certified laboratory to identify coding mutations in a 50-gene panel (n = 34) or a 255-gene panel (n = 1). Cases were evaluated by a multidisciplinary MTB that included pathologists, oncologists, hematologists, basic scientists, and genetic counselors.

Results: During the first year, 35 cases were evaluated by the MTB, with 32 presented for recommendations on targeted therapies, and 3 referred for potential germline mutations. In 56.3% of cases, MTB recommended treatment with a targeted agent based on evaluation of tumor genetic profile and treatment history. Four patients (12.5%) were subsequently treated with a MTB-recommended targeted therapy; 3 of the 4 patients remain on therapy, 2 of whom experienced clinical benefit lasting >10 months.

Conclusion: For the majority of cases evaluated, the MTB was able to provide treatment recommendations based on targetable genetic alterations. The most common reasons that MTB-recommended therapy was not administered stemmed from patient preferences and genetic profiling at either very early or very late stages of disease; lack of drug access was rarely encountered. Increasing awareness of molecular profiling and targeted therapies by both clinicians and patients will improve acceptance and adherence to treatments that could significantly improve outcomes.

Implications For Practice: Case evaluation by a multidisciplinary Molecular Tumor Board (MTB) is critical to benefit from individualized genetic data and maximize clinical impact. MTB recommendations shaped treatment options for the majority of cases evaluated. In the few patients treated with MTB-recommended therapy, disease outcomes were positive and support genetically informed treatment.
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http://dx.doi.org/10.1634/theoncologist.2015-0097DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4571816PMC
September 2015

The PI3K/mTOR dual inhibitor P7170 demonstrates potent activity against endocrine-sensitive and endocrine-resistant ER+ breast cancer.

Breast Cancer Res Treat 2015 Jan 10;149(1):69-79. Epub 2014 Dec 10.

Department of Pharmacology & Toxicology, Norris Cotton Cancer Center, Geisel School of Medicine at Dartmouth, One Medical Center Dr., Lebanon, NH, 03756, USA,

Activation of the phosphatidylinositol 3-kinase (PI3K)/AKT/mTOR pathway has been implicated in anti-estrogen resistance in breast cancer. We tested the therapeutic potential of the novel PI3K/mTOR dual inhibitor P7170 in a panel of anti-estrogen-sensitive and anti-estrogen-resistant models of ER+ breast cancer. Estrogen receptor-positive (ER+) breast cancer cells were treated ±P7170. Fresh cores from primary ER+/HER2- tumors from two patients were treated ±P7170 ex vivo. Mice bearing breast cancer xenografts were randomized to treatment with vehicle, fulvestrant, P7170, or combinations, and tumor volumes were measured. Tissues and cells were analyzed for markers of pathway activity, cell viability, and apoptosis. In cell lines, P7170 exhibited IC50 values in the range of 0.9-7 nM and induced apoptosis. P7170 potently inhibited mTOR activity (≤ 25 nM) and inhibited PI3K at higher concentrations (≥ 200 nM). P7170 completely inhibited MCF-7 tumor growth, significantly inhibited growth of fulvestrant-resistant T47D tumors, and suppressed tumor cell proliferation but did not induce apoptosis. While P7170 inhibits PI3K and mTOR in ER+/HER2- human breast cancer cells and tumors ex vivo, in vivo data indicate that the primary mechanism of P7170 anti-tumor action is inhibition of mTOR and cell proliferation. P7170 is a novel agent worthy of further investigation for the treatment of ER+ breast cancer.
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http://dx.doi.org/10.1007/s10549-014-3201-6DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4302040PMC
January 2015