Exploitation of Sulfated Glycosaminoglycan Status for Precision Medicine of Triplatin in Triple-Negative Breast Cancer.

Triple-negative breast cancer (TNBC) is a subtype of breast cancer lacking targetable biomarkers. TNBC is known to be most aggressive and when metastatic is often drug-resistant and uncurable. Biomarkers predicting response to therapy improve treatment decisions and allow personalized approaches for patients with TNBC. This study explores sulfated glycosaminoglycan (sGAG) levels as a predictor of TNBC response to platinum therapy. sGAG levels were quantified in three distinct TNBC tumor models, including cell line-derived, patient-derived xenograft (PDX) tumors, and isogenic models deficient in sGAG biosynthesis. The in vivo antitumor efficacy of Triplatin, a sGAG-directed platinum agent, was compared in these models with the clinical platinum agent, carboplatin. We determined that >40% of TNBC PDX tissue microarray samples have high levels of sGAGs. The in vivo accumulation of Triplatin in tumors as well as antitumor efficacy of Triplatin positively correlated with sGAG levels on tumor cells, whereas carboplatin followed the opposite trend. In carboplatin-resistant tumor models expressing high levels of sGAGs, Triplatin decreased primary tumor growth, reduced lung metastases, and inhibited metastatic growth in lungs, liver, and ovaries. sGAG levels served as a predictor of Triplatin sensitivity in TNBC. Triplatin may be particularly beneficial in treating patients with chemotherapy-resistant tumors who have evidence of residual disease after standard neoadjuvant chemotherapy. More effective neoadjuvant and adjuvant treatment will likely improve clinical outcome of TNBC.

Genomic screening reveals ubiquitin-like modifier activating enzyme 1 as a potent and druggable target in c-MYC-high triple negative breast cancer models.

Triple negative breast cancer (TNBC) accounts for over 30% of all breast cancer (BC)-related deaths, despite accounting for only 10% to 15% of total BC cases. Targeted therapy development has largely stalled in TNBC, underlined by a lack of traditionally druggable addictions like receptor tyrosine kinases (RTKs). Here, through full genome CRISPR/Cas9 screening of TNBC models, we have uncovered the sensitivity of TNBCs to the depletion of the ubiquitin-like modifier activating enzyme 1 (UBA1). Targeting UBA1 with the first-in-class UBA1 inhibitor TAK-243 induced unresolvable endoplasmic reticulum (ER)-stress and activating transcription factor 4 (ATF4)-mediated upregulation of proapoptotic NOXA, leading to cell death. c-MYC expression correlates with TAK-243 sensitivity and cooperates with TAK-243 to induce a stress response and cell death. Importantly, there was an order of magnitude greater sensitivity of TNBC lines to TAK-243 compared to normal tissue-derived cells. In five patient derived xenograft models (PDXs) of TNBC, TAK-243 therapy led to tumor inhibition or frank tumor regression. Moreover, in an intracardiac metastatic model of TNBC, TAK-243 markedly reduced metastatic burden, indicating UBA1 is a potential new target in TNBC expressing high levels of c-MYC.

Prolactin Drives a Dynamic STAT5A/HDAC6/HMGN2 Cis-Regulatory Landscape Exploitable in ER+ Breast Cancer.

The hormone prolactin has been implicated in breast cancer pathogenesis and regulates chromatin engagement by the transcription factor, STAT5A. STAT5A is known to inducibly bind promoters and cis-regulatory elements genome-wide, though the mechanisms by which it exerts specificity and regulation of target gene expression remain enigmatic. We previously identified HDAC6 and HMGN2 as cofactors that facilitate prolactin-induced, STAT5A-mediated gene expression. Here, multicondition STAT5A, HDAC6, and HMGN2 chromatin immunoprecipitation and sequencing with parallel condition RNA-seq are utilized to reveal the cis-regulatory landscape and cofactor dynamics underlying prolactin-stimulated gene expression in breast cancer. We find that prolactin-regulated genes are significantly enriched for cis-regulatory elements bound by HDAC6 and HMGN2, and that inducible STAT5A binding at enhancers, rather than promoters, conveys specificity for prolactin-regulated genes. The selective HDAC6 inhibitor, ACY-241, blocks prolactin-induced STAT5A chromatin engagement at cis-regulatory elements as well as a significant proportion of prolactin-stimulated gene expression. We identify functional pathways known to contribute to the development and/or progression of breast cancer that are activated by prolactin and inhibited by ACY-241. Additionally, we find that the DNA sequences underlying shared STAT5A and HDAC6 binding sites at enhancers are differentially enriched for estrogen response elements (ESR1 and ESR2 motifs) relative to enhancers bound by STAT5A alone. Gene set enrichment analysis identifies significant overlap of ERα-regulated genes with genes regulated by prolactin, particularly prolactin-regulated genes with promoters or enhancers co-occupied by both STAT5A and HDAC6. Lastly, the therapeutic efficacy of ACY-241 is demonstrated in in vitro and in vivo breast cancer models, where we identify synergistic ACY-241 drug combinations and observe differential sensitivity of ER+ models relative to ER- models.

Identification of synergistic drug combinations using breast cancer patient-derived xenografts.

Compared with other breast cancer subtypes, triple-negative breast cancer (TNBC) is associated with relatively poor outcomes due to its metastatic propensity, frequent failure to respond to chemotherapy, and lack of alternative, targeted treatment options, despite decades of major research efforts. Our studies sought to identify promising targeted therapeutic candidates for TNBC through in vitro screening of 1,363 drugs in patient-derived xenograft (PDX) models. Using this approach, we generated a dataset that can be used to assess and compare responses of various breast cancer PDXs to many different drugs. Through a series of further drug screening assays and two-drug combination testing, we identified that the combination of afatinib (epidermal growth factor receptor (EGFR) inhibitor) and YM155 (inhibitor of baculoviral inhibitor of apoptosis repeat-containing 5 (BIRC5; survivin) expression) is synergistically cytotoxic across multiple models of basal-like TNBC and reduces PDX mammary tumor growth in vivo. We found that YM155 reduces EGFR expression in TNBC cells, shedding light on its potential mechanism of synergism with afatinib. Both EGFR and BIRC5 are highly expressed in basal-like PDXs, cell lines, and patients, and high expression of both genes reduces metastasis-free survival, suggesting that co-targeting of these proteins holds promise for potential clinical success in TNBC.

Quantitative assessment of breast cancer liver metastasis expansion with patient-derived xenografts.

Advanced breast cancer often spreads to the bone, brain, liver, and lungs. The survival time of a patient with breast cancer liver metastasis is often less than 9 months without treatment. Experimental model systems often focus on the lung as a site of metastatic relapse, and therefore, there is less of an understanding of the biological processes that occur during expansive liver metastasis growth. In these studies, 14 genetically distinct breast cancer patient-derived xenografts (PDXs) were characterized for growth in the liver after portal vein injection of cancer cells. Growth in the liver occurred in 12 of 14 models, and the relative growth rate across the PDXs was overall similar to growth in the mammary gland. Pathological and immunohistochemical analyses revealed that the proliferation rates of metastases were relatively similar as the metastases expanded until the tumors became necrotic, and then slightly lower proliferation rates were observed. There were influxes of macrophages and neutrophils as the metastases increased in size, suggesting these innate immune cells may result in differential responses to therapeutics in micrometastases compared to macrometastases. The development and characterization of these models is important as future studies can utilize this information to determine if targeted therapies can slow the progression of metastatic disease at different stages in the liver.

Separation of breast cancer and organ microenvironment transcriptomes in metastases.

BACKGROUND: The seed and soil hypothesis was proposed over a century ago to describe why cancer cells (seeds) grow in certain organs (soil). Since then, the genetic properties that define the cancer cells have been heavily investigated; however, genomic mediators within the organ microenvironment that mediate successful metastatic growth are less understood. These studies sought to identify cancer- and organ-specific genomic programs that mediate metastasis. METHODS: In these studies, a set of 14 human breast cancer patient-derived xenograft (PDX) metastasis models was developed and then tested for metastatic tropism with two approaches: spontaneous metastases from mammary tumors and intravenous injection of PDX cells. The transcriptomes of the cancer cells when growing as tumors or metastases were separated from the transcriptomes of the microenvironment via species-specific separation of the genomes. Drug treatment of PDX spheroids was performed to determine if genes activated in metastases may identify targetable mediators of viability. RESULTS: The experimental approaches that generated metastases in PDX models were identified. RNA sequencing of 134 tumors, metastases, and normal non-metastatic organs identified cancer- and organ-specific genomic properties that mediated metastasis. A common genomic response of the liver microenvironment was found to occur in reaction to the invading PDX cells. Genes within the cancer cells were found to be either transiently regulated by the microenvironment or permanently altered due to clonal selection of metastatic sublines. Gene Set Enrichment Analyses identified more than 400 gene signatures that were commonly activated in metastases across basal-like PDXs. A Src signaling signature was found to be extensively upregulated in metastases, and Src inhibitors were found to be cytotoxic to PDX spheroids. CONCLUSIONS: These studies identified that during the growth of breast cancer metastases, there were genomic changes that occurred within both the cancer cells and the organ microenvironment. We hypothesize that pathways upregulated in metastases are mediators of viability and that simultaneously targeting changes within different cancer cell pathways and/or different tissue compartments may be needed for inhibition of disease progression.

Sphingosine kinase 1 activation by estrogen receptor α36 contributes to tamoxifen resistance in breast cancer.

In breast cancer, 17ß-estradiol (E2) plays critical roles mainly by binding to its canonical receptor, estrogen receptor (ER) α66, and eliciting genomic effects. E2 also triggers rapid, nongenomic responses. E2 activates sphingosine kinase 1 (SphK1), increasing sphingosine-1-phosphate (S1P) that binds to its receptors, leading to important breast cancer signaling. However, the E2 receptor responsible for SphK1 activation has not yet been identified. Here, we demonstrate in triple-negative breast cancer cells, which lack the canonical ERα66 but express the novel splice variant ERα36, that ERα36 is the receptor responsible for E2-induced activation of SphK1 and formation and secretion of S1P and dihydro-S1P, the ligands for S1PRs. Tamoxifen, the first-line endocrine therapy for breast cancer, is an antagonist of ERα66, but an agonist of ERα36, and, like E2, activates SphK1 and markedly increases secretion of S1P. A major problem with tamoxifen therapy is development of acquired resistance. We found that tamoxifen resistance correlated with increased SphK1 and ERα36 expression in tamoxifen-resistant breast cancer cells, in patient-derived xenografts, and in endocrine-resistant breast cancer patients. Our data also indicate that targeting this ERα36 and SphK1 axis may be a therapeutic option to circumvent endocrine resistance and improve patient outcome.

Ablation of the CD9 receptor in human lung cancer cells using CRISPR/Cas alters migration to chemoattractants including IL-16.

CD9, a member of the tetraspanin superfamily, has been implicated in regulating various physiological processes, including cell motility, adhesion, apoptosis and metastasis. Recently, interleukin-16 (IL-16), a pro-inflammatory cytokine released by normal airway and alveolar epithelial cells, has been implicated as a possible ligand for CD9 as an alternative receptor. In this study, using A549 cells as a model of human alveolar epithelium, CD9 expression was ablated using CRISPR/Cas technology. Decreased expression of CD9 mRNA and protein levels was confirmed through RT-qPCR and flow cytometry, respectively. Individual clones were generated that expressed high levels of CD9 (wild-type, WT) or significantly less CD9 (knockdown, KD). Both wild-type and knockdown A549 cell migration was quantified using a FluoroBloc transwell chemotaxis assay. Our results indicate that wild-type A549 cells migrated towards chemoattractants. Moreover, CD9 expression was required in this process since the CD9 knockdown cells had a significantly reduced migration towards growth serum and IL-16. These results support the migratory properties for CD9 in human lung cells and support the hypothesis that CD9 serves as an alternative receptor for IL-16.

Characterizing the efficacy of cancer therapeutics in patient-derived xenograft models of metastatic breast cancer.

PURPOSE: Basal-like breast cancers are aggressive and often metastasize to vital organs. Treatment is largely limited to chemotherapy. This study aims to characterize the efficacy of cancer therapeutics in vitro and in vivo within the primary tumor and metastatic setting, using patient-derived xenograft (PDX) models. METHODS: We employed two basal-like, triple-negative PDX models, WHIM2 and WHIM30. PDX cells, obtained from mammary tumors grown in mice, were treated with twelve cancer therapeutics to evaluate their cytotoxicity in vitro. Four of the effective drugs-carboplatin, cyclophosphamide, bortezomib, and dacarbazine-were tested in vivo for their efficacy in treating mammary tumors, and metastases generated by intracardiac injection of tumor cells. RESULTS: RNA sequencing showed that global gene expression of PDX cells grown in the mammary gland was similar to those tested in culture. In vitro, carboplatin was cytotoxic to WHIM30 but not WHIM2, whereas bortezomib, dacarbazine, and cyclophosphamide were cytotoxic to both lines. Yet, these drugs were ineffective in treating both primary and metastatic WHIM2 tumors in vivo. Carboplatin and cyclophosphamide were effective in treating WHIM30 mammary tumors and reducing metastatic burden in the brain, liver, and lungs. WHIM2 and WHIM30 metastases showed distinct patterns of cytokeratin and vimentin expression, regardless of treatment, suggesting that different tumor cell subpopulations may preferentially seed in different organs. CONCLUSIONS: This study highlights the utility of PDX models for studying the efficacy of therapeutics in reducing metastatic burden in specific organs. The differential treatment responses between two PDX models of the same intrinsic subtype, in both the primary and metastatic setting, recapitulates the challenges faced in treating cancer patients and highlights the need for combination therapies and predictive biomarkers.

Live Cell Mass Accumulation Measurement Non-Invasively Predicts Carboplatin Sensitivity in Triple-Negative Breast Cancer Patient-Derived Xenografts.

Prompt and repeated assessments of tumor sensitivity to available therapeutics could reduce patient morbidity and mortality by quickly identifying therapeutic resistance and optimizing treatment regimens. Analysis of changes in cancer cell biomass has shown promise in assessing drug sensitivity and fulfilling these requirements. However, a major limitation of previous studies in solid tumors, which comprise 90% of cancers, is the use of cancer cell lines rather than freshly isolated tumor material. As a result, existing biomass protocols are not obviously extensible to real patient tumors owing to potential artifacts that would be generated by the removal of cells from their microenvironment and the deleterious effects of excision and purification. In this present work, we show that simple excision of human triple-negative breast cancer (TNBC) tumors growing in immunodeficient mouse, patient-derived xenograft (PDX) models, followed by enzymatic disaggregation into single cell suspension, is enabling for rapid and accurate biomass accumulation-based predictions of in vivo sensitivity to the chemotherapeutic drug carboplatin. We successfully correlate in vitro biomass results with in vivo treatment results in three TNBC PDX models that have differential sensitivity to this drug. With a maximum turnaround time of 40 h from tumor excision to useable results and a fully-automated analysis pipeline, the assay described here has significant potential for translation to clinical practice.

Epidemiology of community-onset bloodstream infections in Bouaké, central Côte d'Ivoire.

Bacterial bloodstream infections (BSI) account for considerable morbidity worldwide, but epidemiological data from resource-constrained tropical settings are scarce. We analysed 293 blood cultures from patients presenting to a regional referral hospital in Bouaké, central Côte d'Ivoire, to determine the aetiology of community-onset BSI. The prevalence of bacteraemia was 22.5%, with children being most commonly affected. Enterobacteriaceae (predominantly Klebsiella pneumoniae and Salmonella enterica) accounted for 94% of BSI. Staphylococcus aureus was the only relevant Gram-positive pathogen. Clinical signs and symptoms were not significantly associated with blood culture positivity after controlling for malaria.

Semapimod sensitizes glioblastoma tumors to ionizing radiation by targeting microglia.

Glioblastoma is the most malignant and lethal form of astrocytoma, with patients having a median survival time of approximately 15 months with current therapeutic modalities. It is therefore important to identify novel therapeutics. There is mounting evidence that microglia (specialized brain-resident macrophages) play a significant role in the development and progression of glioblastoma tumors. In this paper we show that microglia, in addition to stimulating glioblastoma cell invasion, also promote glioblastoma cell proliferation and resistance to ionizing radiation in vitro. We found that semapimod, a drug that selectively interferes with the function of macrophages and microglia, potently inhibits microglia-stimulated GL261 invasion, without affecting serum-stimulated glioblastoma cell invasion. Semapimod also inhibits microglia-stimulated resistance of glioblastoma cells to radiation, but has no significant effect on microglia-stimulated glioblastoma cell proliferation. We also found that intracranially administered semapimod strongly increases the survival of GL261 tumor-bearing animals in combination with radiation, but has no significant benefit in the absence of radiation. In conclusion, our observations indicate that semapimod sensitizes glioblastoma tumors to ionizing radiation by targeting microglia and/or infiltrating macrophages.

[Sensitivity to penicillin G of isolates of pneumococci meningitis and therapeutic implication in CHU of Treichville, Abidjan]. / Sensibilité à la pénicilline G des pneumocoques isolés des méningites et implication thérapeutique au CHU de Treichville - Abidjan.

To determine the frequency of pneumococci meningitis with reduced sensitivity to penicillin G isolated from cerebrospinal fluid, 539 strains were studied between 1996 to 2000. All strains were analysed with oxacilline containing 5 mcg / standard antibiogram for determination of minimal inhibitory concentration (MIC) for penicillin G by E-test. Our results do not show any significant variation from 1996 to 2000. Generally as regard the penicillin G sensitivity we observed that strains were classified as: sensitive strains (CMI < or = 0.06 mg/l): 22.5%, less sensitive strains (0.6 mg/l < CMI < or = 2 mg/l): 58.2%, resistance (CMI > 2 mg/l): 19.2%. The pneumococci rate with reduced sensitivity observed in our study is high and should be taken into consideration in the therapeutic choices.

[Epidemiology and prevention of Staphylococcus aureus nasal carriage in patients and staff at the Cococy Hemodialysis Center in Abidjan, Ivory Coast]. / Epidémiologie et prévention du portage nasal de Staphylococcus aureus chez les malades et le personnel du centre d'hémodialyse de Cocody-Abidjan.

The purpose of this study was to improve prevention of nosocomial infection due to Staphylococcus aureus in patients treated at the Cocody Hemodialysis Center in Abidjan, Cote d'Ivoire. Samples were collected from 56 dialysis patients and 8 staff members. Findings showed that there were 36 transient carriers (64.3%) and 12 permanent carriers (21.4%) in the patient group and 6 transient carriers (75%) and 1 permanent carrier (12.5%) in the staff group. A total of 181 strains of Staphylococcus aureus were isolated. Forty strains (22%) were resistant to meticillin and 179 (98.9%) were sensitive to mupirocin.

[Bacteriologic aspects of Haemophilus influenzae meningitis in children at the Central University Hospital of Treichville]. / Aspects bactériologiques des méningites dues à Haemophilus influenzae chez l'enfant au CHU de Treichville.

The object of our study was to contribute to the knowledge of the epidemiology and bacteriology of Haemophilus influenzae so as to ensure better treatment of patients consulting the Treichville Teaching Hospital. In the bacteriology laboratory of that hospital, between January 1996 and December 1999, we diagnosed by culture and/or soluble antigen, 203 cases of Haemophilus influenzae in children. We observed that biovar IV was dominant (49.7) amongst the 163 (80.3%) positive cultures. Serovar b was the most frequent capsular type found 189/203 (93%). Of the 203 specimens, the frequency of other capsular forms was e 9/203 (4.5%), f 3/203 (1.5%), c 2/203 (1%). Resistance to ampicillin is currently estimated at 39.9%, whereas resistance to chloramphenicol is estimated at 5.5%.