A case study on the implementation of quality and comprehensive indicators set in routine diabetes care.

INTRODUCTION: Type 2 diabetes is a major public health issue in Mexico due to its high prevalence and its projection for the coming years for this disease. Findings on multidisciplinary care related to chronic diseases have proven effective, based on measurement of patient-centered outcomes, The Center of Comprehensive Care for Patients with Diabetes (CAIPaDi) is a multidisciplinary program focused on reducing diabetes complications. This case study aims to illustrate the results of implementing health outcomes measurements and demonstrate the beneficial effects of establishing a comprehensive model of care through a patient-centered approach. METHODS: A descriptive analysis of the comprehensive care indicators of patients with type 2 diabetes treated in the CAIPaDi program between 2013 and 2023 was conducted. The results were structured according to the standard set of outcomes for diabetes proposed by the International Consortium for Health Outcomes Measurements (ICHOM). RESULTS: The baseline and prospective registration of consultations was completed for five years, complying with 25 of the 26 indicators of the ICHOM set. In diabetes control, 56.5% of patients had A1c ≤ 7%, 87.9% had BP ≤ 130/80 mmHg, 60.9% had LDL-cholesterol < 100 mg/dl, and obesity rates decreased from 42.19% to 30.6% during annual consultations. Fewer years of diagnosis before the first visit is key to overall improvement in program adherence (P = 0.02). In acute events, a hyperglycemic crisis occurred in only two cases and severe hypoglycemia episodes in 8 patients. For chronic complications, no lower limb amputations occurred. Cardiovascular outcomes occurred in < 1%. Periodontal disease was analyzed, and periodontitis decreased from 82.9% to 78.7%. Mortality reports were low, with COVID-19 being the main cause of death. Patient-reported outcomes demonstrated reductions in anxiety, depression, and diabetes distress during follow-up. CONCLUSION: Registering quality-of-care indicators is feasible in a comprehensive care program. It allows improving the medical, mental health, and lifestyle outcomes of patients with type 2 diabetes and provides relevant data for planning health programs. A quick diagnosis before program adherence is crucial for overall improvement in patients.

Differential DNA methylation and CTCF binding between the ESR1 promoter a of MCF-7 and MDA-MB-231 breast cancer cells.

BACKGROUND: ESR1 is expressed by 60-70% of breast tumours. it's a good prognosis factor and the target of hormone therapy. Optimization of ESR1 reactivation therapy is currently ongoing. Here we probe if the transcription factor CTCF plays a role in the differential expression of ESR1 in the breast cancer cell lines MCF-7 (ESR1+) and MDA-MB-231 (ESR1-). METHODS AND RESULTS: Knockdown of CTCF in MCF-7 resulted in decreased ESR1 gene expression. CTCF binds to the promoter of ESR1 in MCF-7 but not in MDA-MB-231 cells. CTCF ESR1 binding sites are unmethylated in MCF7 but methylated in MDA-MB-231 cells. CONCLUSION: ESR1 expression in MCF7 cells is dependent on CTCF expression. CTCF can bind to specific regions of the promotor of ESR1 gene in MCF-7 cells but not in MDA-MB-231 cells, this correlates with the methylation status of these regions and could be involved in the transcriptional regulation of ESR1.

Evaluations of Anticancer Effects of Combinations of Cisplatin and Tirucallane-Type Triterpenes Isolated from Amphipterygium adstringens (Schltdl).

The cytotoxic activity of combinations of masticadienonic (AMD) or 3αOH-hydroxy-masticadienonic (3αOH-AMD) acids with cisplatin (CDDP) was evaluated against PC3 prostate and HCT116 colon cancer cell lines. Combinations A (half the IC50 value), B (IC50 value), and C (twice the IC50 value) were tested at a 1 : 1 ratio. All AMD plus CDDP combinations demonstrated increased cytotoxic effect, as determined by the sulforhodamine B test, in both cell types. The best combination was B, which showed 93 % and 91 % inhibition of the proliferation of PC3 and HCT116 cells, respectively. It also increased apoptosis in the PC3 cell lines, as evaluated by flow cytometry. However, in vivo tests showed no additional activity from the AMD plus CDDP combinations. These results showed that the increased cytotoxic activity of the combinations in vitro did not reflect in vivo tests. All combinations of 3αOH-AMD plus CDDP exerted antagonistic effects in both cell types.

Differential histological features and myogenic protein levels in distinct muscles of d-sarcoglycan null muscular dystrophy mouse model.

Skeletal muscle (SkM) comprises slow and fast-twitch fibers, which differ in molecular composition, function, and systemic energy consumption. In addition, muscular dystrophies (DM), a group of diverse hereditary diseases, present different patterns of muscle involvement, progression, and severity, suggesting that the regeneration-degeneration process may differ depending on the muscle type. Therefore, the study aimed to explore the expression of proteins involved in the repair process in different muscles at an early stage of muscular dystrophy in the δ-sarcoglycan null mice (Sgcd-null), a limb-girdle muscular dystrophy 2 F model. Hematoxylin & Eosin (H&E) Staining showed a high number of central nuclei in soleus (Sol), tibialis (Ta), gastrocnemius (Gas), and extensor digitorum longus (Edl) from four months Sgcd-null mice. However, fibrosis, determined by trichrome of Gomori modified staining, was only observed in Sgcd-null Sol. In addition, the number of Type I and II fibers variated differentially in the Sgcd-null muscles vs. wild-type muscles. Besides, the protein expression level of ß-catenin, myomaker, MyoD, and myogenin also presented different expression levels in all the Sgcd-null muscles studied. In summary, our study reveals that muscles with different metabolic characteristics showed distinct expression patterns of proteins involved in the muscle regeneration process. These results could be relevant in designing therapies for genetic and acquired myopathy.

Development of a Redox-Polymer-Based Electrochemical Glucose Biosensor Suitable for Integration in Microfluidic 3D Cell Culture Systems.

The inclusion of online, in situ biosensors in microfluidic cell cultures is important to monitor and characterize a physiologically mimicking environment. This work presents the performance of second-generation electrochemical enzymatic biosensors to detect glucose in cell culture media. Glutaraldehyde and ethylene glycol diglycidyl ether (EGDGE) were tested as cross-linkers to immobilize glucose oxidase and an osmium-modified redox polymer on the surface of carbon electrodes. Tests employing screen printed electrodes showed adequate performance in a Roswell Park Memorial Institute (RPMI-1640) media spiked with fetal bovine serum (FBS). Comparable first-generation sensors were shown to be heavily affected by complex biological media. This difference is explained in terms of the respective charge transfer mechanisms. Under the tested conditions, electron hopping between Os redox centers was less vulnerable than H2O2 diffusion to biofouling by the substances present in the cell culture matrix. By employing pencil leads as electrodes, the incorporation of these electrodes in a polydimethylsiloxane (PDMS) microfluidic channel was achieved simply and at a low cost. Under flow conditions, electrodes fabricated using EGDGE presented the best performance with a limit of detection of 0.5 mM, a linear range up to 10 mM, and a sensitivity of 4.69 µA mM-1 cm-2.

The Protective Role of Cranberries and Blueberries in Oral Cancer.

BACKGROUND: Oral cancer has a high prevalence worldwide, and this disease is caused by genetic, immunological, and environmental factors. The main risk factors associated with oral cancer are smoking and alcohol. RESULTS: There are various strategies to reduce risk factors, including prevention programs as well as the consumption of an adequate diet that includes phytochemical compounds derived from cranberries (Vaccinium macrocarpon A.) and blueberries (Vaccinium corymbosum L.); these compounds exhibit antitumor properties. RESULTS: The main outcome of this review is as follows: the properties of phytochemicals derived from cranberries were evaluated for protection against risk factors associated with oral cancer. CONCLUSIONS: The secondary metabolites of cranberries promote biological effects that provide protection against smoking and alcoholism. An alternative for the prevention of oral cancer can be the consumption of these cranberries and blueberries.

The effect of cilostazol on the platelet-derived growth factor-beta/beta isoform reduction on venous hyperplasia in an experimental balloon-induced injury model.

BACKGROUND: Intimal hyperplasia is the response to endothelial injury. Platelet-derived growth factor is released early and favors the formation of intimal hyperplasia. Although multiple treatments, from open surgery to endovascular techniques, have been used they remain controversial. There is currently interest in developing pharmacological strategies to address this pathology. Local vascular inflammation induced by vessel barotrauma generates intimal hyperplasia due to mechanical stress over the venous endothelium. Cilostazol is a selective phosphodiesterase type 3 (PDE3) selective inhibitor with a regulatory effect over intimal hyperplasia. The objective was to investigate cilostazol's role in inhibiting smooth muscle cell proliferation due to changes in the expression and release of PDGF-BB isoform and the effect on developing IH using an experimental model of vascular barotrauma (balloon-induced injury model). METHODS: We included 12 New Zealand rabbits. The balloon-induced injury model (BIIM) and experimental group cilostazol (20 mg/kg/day) included 6 rabbits each. Contralateral veins from 6 rabbits used in BIIM model has been taken as control group. We measured and compared the expression of PDGF-BB and the development of IH. A pathologist board chooses a PDGFRα antibody to localized its expression by immunohistochemistry analysis. Subsequently, using an automated immunohistochemical staining machine, the PDGFR expression was evaluated using a Zeiss Primo Star 4 light microscope. RESULTS: The measurement obtained in the intimal layer was: 126.12 µm2 in the CG, 232 µm2 in the BIIM group, and 178 µm2 in the EG. A statistically significant difference was observed. Baseline serum concentrations of PDGF-BB in the BIIM group were 0.22 pg/mL. At 12 h 0.42 pg/mL, and 0.17 pg/mL at seven days. In the experimental group, the basal levels were 0.33 pg/mL. With the use of cilostazol, a lower peak was obtained at 12 h (0.08 pg/mL). This difference was statistically significant. CONCLUSIONS: Cilostazol induced a significant reduction of IH caused by barotrauma in the venous endothelium, which correlates with decrease in the PDGF-BB in serum. This could be attributed to the pharmacologic effect on PDGFR expression.

A novel nonsense mutation in the insulin receptor gene in a patient with HAIR-AN syndrome and endometrial cancer.

Severe insulin resistance can be caused by rare genetic defects in the insulin receptor known as insulin receptoropathies. These genetic defects cause a wide spectrum of clinical manifestations ranging from mild syndromes to lethal disorders. Among those is the HAIR-AN an extreme subtype of polycystic ovary syndrome (PCOS). We present a case of a 29-year-old woman with amenorrhea, severe insulin resistance, hirsutism, and acanthosis nigricans who also developed endometrial cancer. She was found to carry a novel heterozygous nonsense mutation insulin receptor gene (INSR). The mutation was inherited from the mother. Levels of insulin receptor and AKT were measured using Western-Blot from peripheral blood mononuclear cells and were both decreased. Thus, we conclude that the identified mutation in the insulin receptor gene and lead to decreased activity of the downstream signaling of the insulin pathway.

Endothelium-Dependent Induction of Vasculogenic Mimicry in Human Triple-Negative Breast Cancer Cells Is Inhibited by Calcitriol and Curcumin.

In highly aggressive tumors, cancer cells may form channel-like structures through a process known as vasculogenic mimicry (VM). VM is generally associated with metastasis, mesenchymal phenotype, and treatment resistance. VM can be driven by antiangiogenic treatments and/or tumor microenvironment-derived factors, including those from the endothelium. Curcumin, a turmeric product, inhibits VM in some tumors, while calcitriol, the most active vitamin D metabolite, exerts potent antineoplastic effects. However, the effect of these natural products on VM in breast cancer remains unknown. Herein, we studied the effect of both compounds on triple-negative breast cancer (TNBC) VM-capacity in a co-culture model. The process of endothelial cell-induced VM in two human TNBC cell lines was robustly inhibited by calcitriol and partially by curcumin. Calcitriol promoted TNBC cells' morphological change from spindle-like to cobblestone-shape, while curcumin diminished VM 3D-structure. Notably, the treatments dephosphorylated several active kinases, especially those involved in the PI3K/Akt pathway. In summary, calcitriol and curcumin disrupted endothelium-induced VM in TNBC cells partially by PI3K/Akt inactivation and mesenchymal phenotype inhibition. Our results support the possible use of these natural compounds as adjuvants for VM inactivation in patients with malignant tumors inherently capable of forming VM, or those with antiangiogenic therapy, warranting further in vivo studies.

(-) - Epicatechin improves Tibialis anterior muscle repair in CD1 mice with BaCl2-induced damage.

Several studies have shown the beneficial effect that Epicatechin (Epi) has on the skeletal muscle of murine models and patients with muscular dystrophy and in the muscles of patients with diabetes or murine sarcopenia models. This flavanol has been shown to enhance antioxidant pathways and improve muscle architecture. However, the repair process during muscle regeneration has not been analyzed. To address this, we characterize the effect of Epi in the repair process of the Tibialis anterior in a murine model with BaCl2-induced damage. CD1 mice of 10 weeks of age were randomly selected and injured with BaCl2. One hour later, they were divided into four groups (n=6 for histology groups and n=12 for western blot groups). Epi was administered every 12h, until the time of sacrifice. Histological and morphological analysis showed that Epi significantly reduced the area of damage and hypertrophy at 15 days in the damaged muscle. Furthermore, western blot assays showed that the treatment increases ß-catenin (active) and myogenic proteins such as MyoD and Myogenin. These results show that Epi exerts therapeutic effects accelerating skeletal muscle repair after induced damage chemically, thus highlighting the therapeutic potential of this flavanol in different myopathies.

Gene Promoter-Methylation Signature as Biomarker to Predict Cisplatin-Radiotherapy Sensitivity in Locally Advanced Cervical Cancer.

Despite efforts to promote health policies focused on screening and early detection, cervical cancer continues to be one of the leading causes of mortality in women; in 2020, estimated 30,000 deaths in Latin America were reported for this type of tumor. While the therapies used to treat cervical cancer have excellent results in tumors identified in early stages, those women who are diagnosed in locally advanced and advanced stages show survival rates at 5 years of <50%. Molecular patterns associated with clinical response have been studied in patients who present resistance to treatment; none of them have reached clinical practice. It is therefore necessary to continue analyzing molecular patterns that allow us to identify patients at risk of developing resistance to conventional therapy. In this study, we analyzed the global methylation profile of 22 patients diagnosed with locally advanced cervical cancer and validated the genomic results in an independent cohort of 70 patients. We showed that BRD9 promoter region methylation and CTU1 demethylation were associated with a higher overall survival (p = 0.06) and progression-free survival (p = 0.0001), whereas DOCK8 demethylation was associated with therapy-resistant patients and a lower overall survival and progression-free survival (p = 0.025 and p = 0.0001, respectively). Our results suggest that methylation of promoter regions in specific genes may provide molecular markers associated with response to treatment in cancer; further investigation is needed.

Sera from women with different metabolic and menopause states differentially regulate cell viability and Akt activation in a breast cancer in-vitro model.

Obesity is associated with an increased incidence and aggressiveness of breast cancer and is estimated to increment the development of this tumor by 50 to 86%. These associations are driven, in part, by changes in the serum molecules. Epidemiological studies have reported that Metformin reduces the incidence of obesity-associated cancer, probably by regulating the metabolic state. In this study, we evaluated in a breast cancer in-vitro model the activation of the IR-ß/Akt/p70S6K pathway by exposure to human sera with different metabolic and hormonal characteristics. Furthermore, we evaluated the effect of brief Metformin treatment on sera of obese postmenopausal women and its impact on Akt and NF-κB activation. We demonstrated that MCF-7 cells represent a robust cellular model to differentiate Akt pathway activation influenced by the stimulation with sera from obese women, resulting in increased cell viability rates compared to cells stimulated with sera from normal-weight women. In particular, stimulation with sera from postmenopausal obese women showed an increase in the phosphorylation of IR-ß and Akt proteins. These effects were reversed after exposure of MCF-7 cells to sera from postmenopausal obese women with insulin resistance with Metformin treatment. Whereas sera from women without insulin resistance affected NF-κB regulation. We further demonstrated that sera from post-Metformin obese women induced an increase in p38 phosphorylation, independent of insulin resistance. Our results suggest a possible mechanism in which obesity-mediated serum molecules could enhance the development of luminal A-breast cancer by increasing Akt activation. Further, we provided evidence that the phenomenon was reversed by Metformin treatment in a subgroup of women.

Establishment of triple-negative breast cancer cells based on BMI: A novel model in the correlation between obesity and breast cancer.

Introduction: Obesity has been associated with an increased risk of biologically aggressive variants in breast cancer. Women with obesity often have tumors diagnosed at later stages of the disease, associated with a poorer prognosis and a different response to treatment. Human cell lines have been derived from specific subtypes of breast cancer and have served to define the cell physiology of corresponding breast cancer subtypes. However, there are no current cell lines for breast cancer specifically derived from patients with different BMIs. The availability of those breast cancer cell lines should allow to describe and unravel functional alterations linked to these comorbidities. Methods: Cell cultures were established from tumor explants. Once generated, the triple negative subtype in a patient with obesity and a patient with a normal BMI were chosen for comparison. For cellular characterization, the following assays were conducted: proliferation assays, chemo - sensitivity assays for doxorubicin and paclitaxel, wound healing motility assays, matrix invasion assays, breast cancer cell growth to estradiol by chronic exposure to leptin, induction of endothelial permeability and tumorigenic potential in athymic mice with normo - versus hypercaloric diets with an evaluation of the epithelium - mesenchymal transformation proteins. Results: Two different cell lines, were established from patients with breast cancer: DSG-BC1, with a BMI of 21.9 kg/m2 and DSG-BC2, with a BMI of 31.5 kg/m2. In vitro, these two cell lines show differential growth rates, motility, chemosensitivity, vascular permeability, response to leptin with an activation of the JAK2/STAT3/AKT signaling pathway. In vivo, they displayed distinct tumorigenic potential. In particular, DSG-BC2, presented higher tumorigenicity when implanted in mice fed with a hypercaloric diet. Discussion: To our knowledge, these primary cultures are the first in vitro representation of both breast cancer and obesity. DSG - BC2 presented a more aggressive in vivo and in vitro phenotype. These results support the hypothesis that breast cancer generated in an obese metabolic state may represent a contrasting variant within the same disease. This new model will allow both further comprehension, functional studies and the analysis of altered molecular mechanisms under the comorbidity of obesity and breast cancer.

Low-Density Granulocytes and Neutrophil Extracellular Traps as Biomarkers of Disease Activity in Adult Inflammatory Myopathies.

BACKGROUND/OBJECTIVE: Biomarkers for disease activity and damage accrual in idiopathic inflammatory myopathies (IIMs) are currently lacking. The purpose of this cross-sectional study is to analyze the relationship among low-density granulocytes (LDGs), neutrophil extracellular traps (NETs), and clinical and immunological features of patients with IIM. METHODS: We assessed disease activity, damage accrual, amount of LDGs, NETs, expression of LL-37, and serum cytokines in 65 adult patients with IIM. Differences between groups and correlations were assessed by Kruskal-Wallis, Mann-Whitney U, and Spearman ρ tests. The association between LDGs, NETs, disease activity, calcinosis, and cutaneous ulcers was assessed by logistic regression. To address the capacity of LDGs and NETs to diagnose disease activity, we used receiving operating characteristic curves. RESULTS: Low-density granulocytes were higher in patients with active disease, ulcers, calcinosis, and anti-MDA5 antibodies, which correlated with serum levels of IL-17A and IL-18. Neutrophil extracellular traps were higher in patients with calcinosis, elevated titers of antinuclear antibodies, and positive anti-PM/Scl75 tests. The combination of a high proportion of both total LDGs and NETs was associated with the presence of calcinosis and cutaneous ulcers. LL-37 was higher in NETs originating from LDGs. Normal-density neutrophils were elevated in patients with active dermatomyositis. CONCLUSIONS: Low-density granulocytes and NETs containing LL-37 are increased in patients with IIM and active disease, and correlate with proinflammatory cytokines. Both total and CD10+ LDGs are potential biomarkers for disease activity and, in combination with NETs, have the potential to detect patients who are at risk for cutaneous ulcers and calcinosis.

The Effects of COVID-19 on Healthcare Workers and Non-Healthcare Workers in Mexico: 14 Months into the Pandemic.

Background and Objectives: Healthcare workers (HCWs) play important roles in mitigating the COVID-19 pandemic and are more likely to become infected with COVID-19. Mexico, among other countries, had a high incidence and prevalence of cases and deaths from this disease. Material and Methods: This retrospective study evaluated the clinical characteristics as well as the geographical distribution of cases, deaths, and active cases of COVID-19 in HCWs and non-HCWs using official information from the Ministry of Health of Mexico. Results: A total of 235,343 cases of COVID-19 were reported in healthcare workers, and 2,094,191 cases were reported in non-healthcare workers. A total of 76.0% of cases in healthcare workers occurred in those who were between 25 and 50 years of age, and 71.4% of deaths occurred in those who were 50 to 69 years of age. Among healthcare workers, the most frequent comorbidities were obesity (15.2%), hypertension (10.9%), and diabetes (6.8%). Nurses were the group with the most cases (39.7%), followed by other healthcare workers (30.6%), physicians (26%), and dentists (1.6%). Physicians were the group with the most deaths (46%), followed by other professionals (30%), nurses (19%), and dentists (3%). Conclusion: These findings are likely the result of healthcare workers in Mexico being at a greater risk of exposure to SARS-CoV-2.

Increased FNDC5/IRISIN protein expression in breast cancer tissue is associated with obesity in postmenopausal women.

AIM: To analyse the fibronectin type III domain containing 5 (FNDC5)/irisin expression in tumour tissue of postmenopausal women presenting breast cancer and different body mass indexes (BMIs), proposing that obesity deregulates the expression of FNDC5/irisin at the breast tumour level. In addition, we investigated if different breast cancer cell lines are capable to synthesise this protein. METHODS: A total of 150 postmenopausal women (50 with a normal BMI, 50 presenting overweight and 50 having obesity) diagnosed with operable breast cancer were included. FNDC5/irisin expression was determined by immunohistochemistry or by immunocytochemistry. Qualitative analysis of protein expression was performed by the H-Score method, through ImageJ's IHC Profiler software. Statistical analyses were carried out using STATA V.14.0 (Texas, USA); p value<0.05 was accepted as statistically significant. Statistical power of the study was >80% with a p<0.05. RESULTS: FNDC5/irisin expression in breast cancer tissue of postmenopausal women with obesity was significantly increased when compared with FNDC5/irisin expression in women with a normal BMI (p=0.001). Furthermore, three breast cancer cell lines studied were capable to synthesise and express FNDC5/irisin, being the BT-474 cell line the one that exhibited the highest intensity of expression. CONCLUSIONS: Our results confirm that women with breast cancer and obesity exhibit an increased irisin expression in their tumorous tissue compared with women with breast cancer and normal BMI. Likewise, in vitro breast cancer cell lines have the capacity to synthesise and express FNDC5/irisin, without any extracellular stimuli, however the microenvironment surrounding these cells in vivo participates in its regulation.

Mesenchymal Stem Cells for the Compassionate Treatment of Severe Acute Respiratory Distress Syndrome Due to COVID 19.

Mesenchymal stem cells (MSC) have received particular attention due to their ability to inhibit inflammation caused by cytokine storm induced by COVID-19. In this way some patients have been treated successfully. The aim of this study was to evaluate the safety and describe the clinical changes after IV administration of allogeneic human umbilical cord MSC (ahUCMSC), in patients with bilateral pneumonia caused by COVID-19, complicated with severe ARDS, as compassionate treatment. This was a pilot, open-label, prospective, longitudinal study. Five patients that did not improve in their clinical conditions after 48 hours of receiving the standard medical management used by the Medical Center and with persistent PaO2/FiO2 less than 100 mmHg were enrolled. ahUCMSC were infused IV, at dose of 1x106 per Kg of body weight over 15 minutes. Patients were monitored after the infusion to detect adverse event. Pa02/FiO2, vital signs, D-dimer, C reactive protein and total lymphocytes were monitored for 21 days after the infusion or until the patient was discharged from the hospital. Descriptive statistics were used with means or medians and standard deviation or interquartile range according to the type of variable. The Wilcoxon's rank-sum was used for stationary samples. Adverse events occurred in three patients and were easily and quickly controlled. Immediately after the infusion of ahUCMSC, constant rise of PaO2/FiO2 was observed in all patients during the first 7 days, with statistical significance. Three patients survived and were extubated on the ninth day post-infusion. Two patients died at 13 and 15 days after infusion. The infusion of ahUCMSC in patients with severe ARDS caused by COVID-19, was safe, and demonstrated its anti-inflammatory capacity in the lungs, by improving the respiratory function expressed by PaO2 / FiO2, which allowed the survival of 3 patients, with extubation at 9 days.

p21-Activated Kinase 1 Promotes Breast Tumorigenesis via Phosphorylation and Activation of the Calcium/Calmodulin-Dependent Protein Kinase II.

p21-Activated kinase-1 (Pak1) is frequently overexpressed and/or amplified in human breast cancer and is necessary for transformation of mammary epithelial cells. Here, we show that Pak1 interacts with and phosphorylates the Calcium/Calmodulin-dependent Protein Kinase II (CaMKII), and that pharmacological inhibition or depletion of Pak1 leads to diminished activity of CaMKII. We found a strong correlation between Pak1 and CaMKII expression in human breast cancer samples, and combined inhibition of Pak1 and CaMKII with small-molecule inhibitors was synergistic and induced apoptosis more potently in Her2 positive and triple negative breast cancer (TNBC) cells. Co-adminstration of Pak and CaMKII small-molecule inhibitors resulted in a dramatic reduction of proliferation and an increase in apoptosis in a 3D cell culture setting, as well as an impairment in migration and invasion of TNBC cells. Finally, mice bearing xenografts of TNBC cells showed a significant delay in tumor growth when treated with small-molecule inhibitors of Pak and CaMKII. These data delineate a signaling pathway from Pak1 to CaMKII that is required for efficient proliferation, migration and invasion of mammary epithelial cells, and suggest new therapeutic strategies in breast cancer.

IFI27/ISG12 Downregulates Estrogen Receptor α Transactivation by Facilitating Its Interaction With CRM1/XPO1 in Breast Cancer Cells.

The estrogen receptor alpha (ERα) is a ligand-activated transcription factor whose activity is modulated by its interaction with multiple protein complexes. In this work, we have identified the protein interferon alpha inducible protein 27 (IFI27/ISG12) as a novel ERα-associated protein. IFI27/ISG12 transcription is regulated by interferon and estradiol and its overexpression is associated to reduced overall survival in ER+ breast cancer patients but its function in mammary gland tissue remains elusive. In this study we showed that overexpression of IFI27/ISG12 in breast cancer cells attenuates ERα transactivation activity and the expression of ERα-dependent genes. Our results demonstrated that IFI27/ISG12 overexpression in MCF-7 cells reduced their proliferation rate in 2-D and 3-D cell culture assays and impaired their ability to migrate in a wound-healing assay. We show that IFI27/ISG12 downregulation of ERα transactivation activity is mediated by its ability to facilitate the interaction between ERα and CRM1/XPO1 that mediates the nuclear export of large macromolecules to the cytoplasm. IFI27/ISG12 overexpression was shown to impair the estradiol-dependent proliferation and tamoxifen-induced apoptosis in breast cancer cells. Our results suggest that IFI27/ISG12 may be an important factor in regulating ERα activity in breast cancer cells by modifying its nuclear versus cytoplasmic protein levels. We propose that IFI27/ISG12 may be a potential target of future strategies to control the growth and proliferation of ERα-positive breast cancer tumors.

Biological Landscape of Triple Negative Breast Cancers Expressing CTLA-4.

Patients with triple-negative breast cancer (TNBC) have a poor prognosis, partly because of the absence of targeted therapies. Recognition of the key role of immune responses against cancer has allowed the advent of immunotherapy, focused on the inhibition of negative immune checkpoints, such as CTLA-4. CTLA-4 is also expressed in some cancer cells, but its activity in tumor cells is not completely understood. Thus, the aim of the present work was to determine the biological landscape and functions of CTLA-4 expressed in TNBC cells through preclinical and in silico analysis. Exploration of CTLA-4 by immunohistochemistry in 50 TNBC tumors revealed membrane and cytoplasmic expression at different intensities. Preclinical experiments, using TNBC cell lines, showed that stimulation of CTLA-4 with CD80 enhances activation of the ERK1/2 signaling pathway, while CTLA-4 blockade by Ipilimumab induces the activation of AKT and reduces cell proliferation in vitro. We then developed an analytic pipeline to define the effects of CTLA-4 in available public data that allowed us to identify four distinct tumor clusters associated with CTLA-4 activation, which are characterized by enrichment of distinctive pathways associated with cell adhesion, MAPK signaling, TGF-ß, VEGF, TNF-α, drug metabolism, ion and amino acid transport, and KRAS signaling, among others. In addition, blockade of CTLA-4 induced increased secretion of IL-2 by tumor cells, suggesting that the receptor regulates cellular functions that may impact the immune microenvironment. This is relevant because a deep characterization of immune infiltrate, conducted using public data to estimate the abundancies of immune-cell types, showed that CTLA-4-activated-like tumors present a conditional immune state similar to an escape phenotype exploited by cancer cells. Finally, by interrogating transcriptional predictors of immunotherapy response, we defined that CTLA-4 activation correlates with high immune scores related to good clinical predicted responses to anti-CTLA-4 therapy. This work sheds new light on the roles of activated CLTA-4 in the tumor compartment and suggests an important interplay between tumor CLTA-4-activated portraits and immune-infiltrating cell populations.