Conventional natural killer cells control vascular remodeling in the uterus during pregnancy by acidifying the extracellular matrix with a2V.

Vascular remodeling within the uterus immediately before and during early pregnancy increases blood flow in the fetus and prevents the development of gestational hypertension. Tissue-resident natural killer (trNK) cells secrete pro-angiogenic growth factors but are insufficient for uterine artery (UtA) remodeling in the absence of conventional natural killer (cNK) cells. Matrix metalloproteinase-9 (MMP9) is activated in acidic environments to promote UtA remodeling. We have previously shown that ATPase a2V plays a role in regulating the function of cNK cells during pregnancy. We studied the effect of a2V deletion on uterine cNK cell populations and pregnancy outcomes in VavCrea2Vfl/fl mice, where a2V is conditionally deleted in hematopoietic stem cells. Conventional NKcells were reduced but trNK cells were retained in implantation sites at gestational day 9.5, and UtA remodeling was inhibited despite no differences in concentrations of pro-angiogenic growth factors. The ratio of pro-MMP9 to total was significantly elevated in VavCrea2Vfl/fl mice, and MMP9 activity was significantly reduced. The pH of implantation sites was significantly elevated in VavCrea2Vfl/fl mice. We concluded that the role of cNK cells in the uterus is to acidify the extracellular matrix (ECM) using a2V, which activates MMP9 to degrade the ECM, release bound pro-angiogenic growth factors, and contribute to UtA remodeling. Our results are significant for the understanding of the development of gestational hypertension.

An Ultrasound Scoring Model for the Prediction of Intrapartum Morbidly Adherent Placenta and Maternal Morbidity: A Cross-Sectional Study.

OBJECTIVE: To design an ultrasound scoring model for the prediction of the intrapartum morbidly adherent placenta (MAP) and maternal morbidity. PATIENTS AND METHODS: 114 females with singleton pregnancies ≥ 28 weeks of gestation referred for suspicion of MAP were included. All patients underwent examination by two-dimensional ultrasound with the color Doppler setting. Five signs were evaluated: the retroplacental echolucent space, placental lacunae, the hyperechoic uterine-bladder interface, retroplacental myometrium thickness, and subplacental, uterine serosa-bladder wall, intraplacental and bladder wall vascularity. We designed a score ranging from 0-8.5 points, including the five signs according to their odds ratios and evaluated its prediction for MAP and maternal morbidity. RESULTS: Using multivariate logistic regression, all ultrasound signs were significant dependent predictors for both MAP and maternal morbidity (myometrium thickness < 1 mm followed by lacunae ≥ 4 and lost retroplacental echolucent space). The only independent predictors for MAP were myometrium thickness < 1 mm and lacunae ≥ 4, while myometrium thickness < 1 mm and lost retroplacental echolucent space were predictive for maternal morbidity. The score showed a perfect agreement with MAP and a good one for maternal morbidity. CONCLUSION: Application of the score we designed can improve the ultrasound diagnosis of MAP and the maternal outcome.

Validity and Reproducibility of the ADNEX MR Scoring System in the Diagnosis of Sonographically Indeterminate Adnexal Masses.

BACKGROUND: The diagnosis of sonographically indeterminate adnexal masses (AM) signifies a major challenge in clinical practice. Early detection and characterization have increased the need for accurate imaging evaluation before treatment. PURPOSE: To assess the validity and reproducibility of the ADNEX MR Scoring system in the diagnosis of sonographically indeterminate AM. STUDY TYPE: A prospective multicenter study. POPULATION: In all, 531 women (mean age, 44 ± 11.2 years; range, 21-79 years) with 572 sonographically indeterminate AM. FIELD STRENGTH/SEQUENCE: 1.5T/precontrast T1 -weighted imaging (WI) fast spin echo (FSE) (in-phase and out-of-phase, with and without fat suppression); T2 -WI FSE; diffusion-WI single-shot echo planner with b-values of 0 and 1000 s/mm2 ; and dynamic contrast-enhanced perfusion T1 -WI liver acquisition with volume acceleration (LAVA). ASSESSMENT: All MRI examinations were evaluated by three radiologists, and the AM were categorized into five scores based on the ADNEX MR Scoring system. Score 1: no AM; 2: benign AM; 3: probably benign AM; 4: indeterminate AM; 5: probably malignant AM. Histopathology and imaging follow-up were used as the standard references for evaluating the validity of the ADNEX MR Scoring system for detecting ovarian malignancy. STATISTICAL TESTS: Four-fold table test, kappa statistics (κ), and receiver operating characteristic (ROC) curve. RESULTS: In all, 136 (23.8%) AM were malignant, and 436 (76.2%) were benign. Of the 350 AM classified as score 2, one (0.3%) was malignant; of the 62 AM classified as score 3, six (9.7%) were malignant; of the 73 AM classified as score 4, 43 (58.9%) were malignant; and of the 87 AM categorized as score 5, 86 (98.9%) were malignant. The best cutoff value for predicting malignant AM was score >3 with sensitivity and specificity of 92.9% and 94.9%, respectively. The interreader agreement of the ADNEX MR Scoring was very good (κ = 0.861). DATA CONCLUSION: The current study supports the high validity and reproducibility of the ADNEX MR Scoring system for the diagnosis of sonographically indeterminate AM. LEVEL OF EVIDENCE: 1 TECHNICAL EFFICACY STAGE: 2.

Comparison of O-RADS, GI-RADS, and IOTA simple rules regarding malignancy rate, validity, and reliability for diagnosis of adnexal masses.

OBJECTIVE: The American College of Radiology (ACR) recently published the ovarian-adnexal reporting and data system (O-RADS) to provide guidelines to physicians who interpret ultrasound (US) examinations of adnexal masses (AM). This study aimed to compare the O-RADS with two other well-established US classification systems for diagnosis of AM. METHODS: This retrospective multicenter study between May 2016 and December 2019 assessed consecutive women with AM detected by the US. Five experienced consultant radiologists independently categorized each AM according to O-RADS, gynecologic imaging reporting and data system (GI-RADS), and international ovarian tumor analysis (IOTA) simple rules. Pathology and adequate follow-up were used as reference standards for calculating the validity of three US classification systems for diagnosis of AM. Kappa statistics were used to assess the inter-reviewer agreement (IRA). RESULTS: A total of 609 women (mean age, 48 ± 13.7 years; range, 18-72 years) with 647 AM were included. Of the 647 AM, 178 were malignant and 469 were benign. Malignancy rates were comparable to recommended rates by previous literature in O-RADS and IOTA, but higher in GI-RADS. O-RADS had significantly higher sensitivity for malignancy than GI-RAD and IOTA (p = 0.003 and 0.0007, respectively), but non-significant slightly lower specificity (p > 0.05). O-RADS, GI-RADS, and IOTA showed similar overall IRA (κ = 0.77, 0.69, and 0.63, respectively) with a tendency toward higher IRA with O-RADS than with GI-RADS and IOTA. CONCLUSIONS: O-RADS compares favorably with GI-RADS and IOTA. O-RADS had higher sensitivity than GI-RADS and IOTA simple rules with relatively similar specificity and reliability. KEY POINTS: • The malignancy rates were comparable to recommended rates by previous literature in O-RADS and IOTA, but higher in GI-RADS. • The O-RADS had significantly higher sensitivity for malignancy than GI-RADS and IOTA (96.8% vs 92.7% and 92.1%; p = 0.003 and 0.0007, respectively), but non-significant slightly lower specificity (92.8% vs 93.6% and 93.2%, respectively; p > 0.05). • The O-RADS, GI-RADS, and IOTA showed similar overall inter-reviewer agreement (IRA) (κ = 0.77, 0.69, and 0.63, respectively), with a tendency toward higher IRA with O-RADS than with GI-RADS and IOTA.

In vivo anti-V-ATPase antibody treatment delays ovarian tumor growth by increasing antitumor immune responses.

Tumor acidity is the key metabolic feature promoting cancer progression and is modulated by pH regulators on a cancer cell's surface that pump out excess protons/lactic acid for cancer cell survival. Neutralizing tumor acidity improves the therapeutic efficacy of current treatments including immunotherapies. Vacuolar-ATPase (V-ATPase) proton pumps encompass unique plasma membrane-associated subunit isoforms, making this molecule an important target for anticancer therapy. Here, we examined the in vivo therapeutic efficacy of an antibody (a2v-mAB) targeting specific V-ATPase-'V0a2' surface isoform in controlling ovarian tumor growth. In vitro a2v-mAb treatment inhibited the proton pump activity in ovarian cancer (OVCA) cells. In vivo intraperitoneal a2v-mAb treatment drastically delayed ovarian tumor growth with no measurable in vivo toxicity in a transplant tumor model. To explore the possible mechanism causing delayed tumor growth, histochemical analysis of the a2v-mAb-treated tumor tissues displayed high immune cell infiltration (M1-macrophages, neutrophils, CD103+ cells, and NK cells) and an enhanced antitumor response (iNOS, IFN-y, IL-1α) compared to control. There was marked decrease in CA-125-positive cancer cells and an enhanced active caspase-3 expression in a2v-mAb-treated tumors. RNA-seq analysis of a2v-mAb tumor tissues further revealed upregulation of apoptosis-related and toll-like receptor pathway-related genes. Indirect coculture of a2v-mAb-treated OVCA cells with human PBMCs in an unbuffered medium led to an enhanced gene expression of antitumor molecules IFN-y, IL-17, and IL-12-A in PBMCs, further validating the in vivo antitumor responses. In conclusion, V-ATPase inhibition using a monoclonal antibody directed against the V0a2 isoform increases antitumor immune responses and could therefore constitute an effective treatment strategy in OVCA.

Cancer-associated V-ATPase induces delayed apoptosis of protumorigenic neutrophils.

Tumors and neutrophils undergo an unexpected interaction, in which products released by tumor cells interact to support neutrophils that in turn support cancer growth, angiogenesis, and metastasis. A key protein that is highly expressed by cancer cells in tumors is the a2 isoform V-ATPase (a2V). A peptide from a2V (a2NTD) is secreted specifically by cancer cells, but not normal cells, into the tumor microenvironment. This peptide reprograms neutrophils to promote angiogenesis, cancer cell invasiveness, and neutrophil recruitment. Here, we provide evidence that cancer-associated a2V regulates the life span of protumorigenic neutrophils by influencing the intrinsic pathway of apoptosis. Immunohistochemical analysis of human cancer tissue sections collected from four different organs shows that levels of a2NTD and neutrophil counts are increased in cancer compared with normal tissues. Significant increases in neutrophil counts were present in both poorly and moderately differentiated tumors. In addition, there is a positive correlation between the number of neutrophils and a2NTD expression. Human neutrophils treated with recombinant a2NTD show significantly delayed apoptosis, and such prolonged survival was dependent on NF-κB activation and ROS generation. Induction of antiapoptotic protein expression (Bcl-xL and Bcl-2A1) and decreased expression of proapoptotic proteins (Bax, Apaf-1, caspase-3, caspase-6, and caspase-7) were a hallmark of these treated neutrophils. Autocrine secretion of prosurvival cytokines of TNF-α and IL-8 by treated neutrophils prolongs their survival. Our findings highlight the important role of cancer-associated a2V in regulating protumorigenic innate immunity, identifying a2V as a potential important target for cancer therapy.

Targeting V-ATPase Isoform Restores Cisplatin Activity in Resistant Ovarian Cancer: Inhibition of Autophagy, Endosome Function, and ERK/MEK Pathway.

Ovarian cancer (OVCA) patients often develop tolerance to standard platinum therapy that accounts for extensive treatment failures. Cisplatin resistant OVCA cells (cis-R) display enhanced survival mechanisms to cope with therapeutic stress. In these cells, increased autophagy process assists in chemoresistance by boosting the nutrient pool under stress. To improve the treatment response, both protective autophagy inhibition and its overactivation are showing efficacy in chemosensitization. Autophagy requires a tightly regulated intracellular pH. Vacuolar ATPases (V-ATPases) are proton extruding nanomotors present on cellular/vesicular membranes where they act as primary pH regulators. V-ATPase 'a2' isoform (V0a2), the major pH sensing unit, is markedly overexpressed on the plasma membrane and the early endosomes of OVCA cells. Previously, V0a2 inhibition sensitized cis-R cells to platinum drugs by acidifying cytosolic pH that elevated DNA damage. Here, we examined how V0a2 inhibition affected endosomal function and the autophagy process as a possible factor for cisplatin sensitization. Clinically, V0a2 expression was significantly higher in tissues from drug nonresponder OVCA patients compared to treatment responders. In vitro V0a2 knockdown in cis-R cells (sh-V0a2-cisR) significantly reduced the tumor sphere-forming ability and caused complete disintegration of the spheres upon cisplatin treatment. The apoptotic capacity of sh-V0a2-cisR improved substantially with potentiation of both intrinsic and extrinsic apoptotic pathway when treated with cisplatin. Unlike the chemical V-ATPase inhibitors that acutely induce autophagy, here, the stable V0a2 inhibition dampened the protective autophagy process in sh-V0a2-cisR cells with downregulated expression of proteins beclin-1, ATG-7, and LC3B and low autophagosome numbers compared to control cis-R cells. These cells showed downregulated ERK/MEK pathway that is known to repress autophagy. Interestingly, upon cisplatin treatment of sh-V0a2-cisR, the autophagy initiation proteins (LC3B, ATG7, and Beclin 1) were found upregulated as a stress response compared to the untreated cells. However, there was a concomitant downstream autophagosome accumulation and an enhanced P62 protein levels indicating the overall block in autophagy flux. Mechanistically, V0a2 knockdown caused defects in early endosome function as the transferrin internalization was impaired. Taken together, this study provides a novel insight into the mechanism by which V-ATPase-isoform regulates autophagy that assists in chemoresistance in ovarian cancer. We conclude that V-ATPase-V0a2 is a potent target for developing an effective treatment to enhance patient survival rates in ovarian cancer.

Gynecology Imaging Reporting and Data System (GI-RADS): diagnostic performance and inter-reviewer agreement.

OBJECTIVE: To evaluate diagnostic performance and inter-reviewer agreement (IRA) of the Gynecologic Imaging Reporting and Data System (GI-RADS) for diagnosis of adnexal masses (AMs) by pelvic ultrasound (US). PATIENTS AND METHODS: A prospective multicenter study included 308 women (mean age, 41 ± 12.5 years; range, 15-73 years) with 325 AMs detected by US. All US examinations were analyzed, and AMs were categorized into five categories according to the GI-RADS classification. We used histopathology and US follow-up as the reference standards for calculating diagnostic performance of GI-RADS for detecting malignant AMs. The Fleiss kappa (κ) tests were applied to evaluate the IRA of GI-RADS scoring results for predicting malignant AMs. RESULTS: A total of 325 AMs were evaluated: 127 (39.1%) were malignant and 198 (60.9%) were benign. Of 95 AMs categorized as GI-RADS 2 (GR2), none was malignant; of 94 AMs categorized as GR3, three were malignant; of 13 AMs categorized as GR4, six were malignant; and of 123 AMs categorized as GR5, 118 were malignant. On a lesion-based analysis, the GI-RADS had a sensitivity, a specificity, and an accuracy of 92.9%, 97.5%, and 95.7%, respectively, when regarding only those AMs classified as GR5 for predicting malignancy. Considering combined GR4 and GR5 as a predictor for malignancy, the sensitivity, specificity, and accuracy of GI-RADS were 97.6%, 93.9%, and 95.4%, respectively. The IRA of the GI-RADS category was very good (κ = 0.896). The best cutoff value for predicting malignant AMs was >GR3. CONCLUSIONS: The GI-RADS is very valuable for improving US structural reports. KEY POINTS: • There is still a lack of a standard in the assessment of AMs. • GI-RADS is very valuable for improving US structural reports of AMs. • GI-RADS criteria are easy and work at least as well as IOTA.

Role of Multidetector Computed Tomography (CT) Virtual Hysteroscopy in the Evaluation of Abnormal Uterine Bleeding in Reproductive Age.

BACKGROUND: Abnormal uterine bleeding (AUB) may be acute or chronic and is defined as bleeding from the uterine corpus that is abnormal in regularity, volume, frequency, or duration and occurs in the absence of pregnancy. It is a widespread complaint in the primary care units. The prevalence of abnormal bleeding is up to 30% among women of reproductive age. OBJECTIVE: To assess the role of CT virtual hysteroscopy in the evaluation of the uterine cavity in cases with abnormal uterine bleeding in reproductive age. METHODS: Cross sectional study was performed at Obstetrics and Gynecology Department and Radiology Department, Zagazig University hospitals, Egypt, on 124 women with abnormal uterine bleeding in reproductive age, and their uterine cavity was evaluated by both row multidetector computed tomography (MDCT) scanner and Office hysteroscopy. RESULTS: Mean age of studied group was 28.54 ± 5.99 years, and virtual hysteroscopy showed sensitivity 91.1% and specificity 85.3% in detection of abnormalities within uterine cavity. It showed sensitivity 91.1% and specificity 85.3% in cases of endometrial polyps. It yielded 88.5 % sensitivity and 100 % specificity in cases with submucous fibroids, while it yielded only 57.9 % sensitivity and 82.9% specificity in cases of thick endometrium. CONCLUSION: Virtual CT hysteroscopy is a good negative test in cases of abnormal uterine bleeding but has some limitations that decrease its sensitivity.

Hematopoietic stem cell specific V-ATPase controls breast cancer progression and metastasis via cytotoxic T cells.

The interaction of recruited immune effector cells and cancer cells within tumor microenvironment (TME) shapes the fate of cancer progression and metastasis. Many cancers including breast cancer, express a specific vacuolar ATPase (a2V) on their cell surface which acidifies the extracellular milieu helping cancer cell proliferation and metastasis. To understand the role of immune cell-associated-a2V during breast tumor pathogenesis, we knocked-out a2V (KO) from the hematopoietic stem cells (HSC) and generated breast tumors in mice. The a2V-KO mice developed faster growing, larger, and metastatic breast tumors compared to control mice. Further investigation of the TME revealed a significant reduction in the presence of CD4+ and CD8+ T cells in the a2V-KO tumors. Targeted RNA-Seq of the cells of the TME demonstrated that pro-inflammatory cytokines, death receptors, death receptor ligands, and cytotoxic effectors were significantly down-regulated within the a2V-KO TME. Interestingly, analysis of immune cells in the blood, spleen, and thymus of the non-tumor bearing a2V-KO mice revealed a significant decrease in CD4+ and CD8+ T cell populations. For the first time, this study demonstrates that inhibition of V-ATPase expression in HSC leads to a decrease in CD4+ and CD8+ T cell populations and thus promotes breast tumor growth and metastasis.

Microtubule inhibitor, SP-6-27 inhibits angiogenesis and induces apoptosis in ovarian cancer cells.

In ovarian cancer (OVCA), treatment failure due to chemo-resistance is a serious challenge. It is therefore critical to identify new therapies that are effective against resistant tumors and have reduced side effects. We recently identified 4-H-chromenes as tubulin depolymerizing agents that bind to colchicine site of beta-tubulin. Here, we screened a chemical library of substituted 4-H-chromenes and identified SP-6-27 to exhibit most potent anti-proliferative activity towards a panel of human cisplatin sensitive and resistant OVCA cell lines with 50% inhibitory concentration (IC50; mean ± SD) ranging from 0.10 ± 0.01 to 0.84 ± 0.20 µM. SP-6-27 exhibited minimum cytotoxicity to normal ovarian epithelia. A pronounced decrease in microtubule density as well as G2/M cell cycle arrest was observed in SP-6-27 treated cisplatin sensitive/resistant OVCA cells. The molecular mechanism of SP-6-27 induced cell death revealed modulation in cell-cycle regulation by upregulation of growth arrest and DNA damage inducible alpha transcripts (GADD45). An enhanced intrinsic apoptosis was observed in OVCA cells through upregulation of Bax, Apaf-1, caspase-6, -9, and caspase-3. In vitro wound healing assay revealed reduced OVCA cell migration upon SP-6-27 treatment. Additionally, SP-6-27 and cisplatin combinatorial treatment showed enhanced cytotoxicity in chemo-sensitive/resistant OVCA cells. Besides effect on cancer cells, SP-6-27 further restrained angiogenesis by inhibiting capillary tube formation by human umbilical vein endothelial cells (HUVEC). Together, these findings show that the chromene analog SP-6-27 is a novel chemotherapeutic agent that offers important advantages for the treatment of ovarian cancer.

Cancer derived peptide of vacuolar ATPase 'a2' isoform promotes neutrophil migration by autocrine secretion of IL-8.

Neutrophils play significant regulatory roles within the tumor microenvironment by directly promoting tumor progression that leads to poor clinical outcomes. Identifying the tumor associated molecules that regulate neutrophil infiltration into tumors may provide new and specific therapeutic targets for cancer treatment. The a2-isoform of vacuolar ATPase (a2V) is uniquely and highly expressed on cancer cell plasma membrane. Cancer cells secrete a peptide from a2V (a2NTD) that promotes the pro-tumorigenic properties of neutrophils. This provides a2V the propensity to control neutrophil migration. Here, we report that the treatment of human neutrophils with recombinant a2NTD leads to neutrophil adherence and polarization. Moreover, a2NTD treatment activates surface adhesion receptors, as well as FAK and Src kinases that are essential regulators of the migration process in neutrophils. Functional analysis reveals that a2NTD can act as a chemo-attractant and promotes neutrophil migration. In addition, a2Neuɸ secrete high levels of IL-8 via NF-κB pathway activation. Confirmatory assays demonstrate that the promoted migration of a2Neuɸ was dependent on the autocrine secretion of IL-8 from a2Neuɸ. These findings demonstrate for the first time the direct regulatory role of cancer associated a2-isoform V-ATPase on neutrophil migration, suggesting a2V as a potential target for cancer therapy.

Hormonal Changes After Laparoscopic Ovarian Diathermy in Patients with Polycystic Ovarian Syndrome.

AIM OF THE PRESENT STUDY: To assess the changes in hormonal profile (serum FSH, LH, prolactin and total testosterone) following laparoscopic ovarian drilling (LOD) in patients with polycystic ovarian syndrome. MATERIALS AND METHODS: Fifty patients with PCOS have been included in this study. Serum prolactin, total testosterone, follicular-stimulating hormone (FSH) and luteinizing hormone (LH) levels have been used as biochemical markers, before and after procedures. RESULTS: Laparoscopic ovarian drilling was successfully employed without any surgical complications and on an average follow-up time of 24 weeks after the procedure. During the follow-up serum values for prolactin, total testosterone and LH have decreased significantly and FSH levels remained unchanged after the procedure. CONCLUSIONS: The LOD in patients with PCOS may avoid or reduce the risk of OHSS and the multiple pregnancy rate induced by gonadotropin therapy. The high pregnancy rate and the economic aspect of the procedure offer an attractive management for patients with PCOS. However, LOD can be considered as second-line treatment after clomiphene citrate treatment failure and/or resistance.

Selective inhibition of tumor cell associated Vacuolar-ATPase 'a2' isoform overcomes cisplatin resistance in ovarian cancer cells.

Development of resistance to platinum compounds significantly hinders successful ovarian cancer (OVCA) treatment. In tumor cells, dysregulated pH gradient across cell membranes is a key physiological mechanism of metastasis/chemo-resistance. These pH alterations are mediated by aberrant activation of key multi-subunit proton pumps, Vacuolar-ATPases (V-ATPases). In tumor cells, its 'a2' isoform (V-ATPase-V0a2) is a component of functional plasma-membrane complex and promotes tumor invasion through tumor-acidification and immuno-modulation. Its involvement in chemo-resistance has not been studied. Here, we show that V-ATPase-V0a2 is over-expressed in acquired-cisplatin resistant OVCA cells (cis-A2780/cis-TOV112D). Of all the 'a' subunit isoforms, V-ATPase-V0a2 exhibited an elevated expression on plasma membrane of cisplatin-resistant cells compared to sensitive counterparts. Immuno-histochemistry revealed V-ATPase-V0a2 expression in both low grade (highly drug-resistant) and high grade (highly recurrent) human OVCA tissues indicating its role in a centralized mechanism of tumor resistance. In cisplatin resistant cells, shRNA mediated inhibition of V-ATPase-V0a2 enhanced sensitivity towards both cisplatin and carboplatin. This improved cytotoxicity was mediated by enhanced cisplatin-DNA-adduct formation and suppressed DNA-repair pathway, leading to enhanced apoptosis. Suppression of V0a2 activity strongly reduced cytosolic pH in resistant tumor cells, which is known to enhance platinum-associated DNA-damage. As an indicator of reduced metastasis and chemo-resistance, in contrast to plasma membrane localization, a diffused cytoplasmic localization of acidic vacuoles was observed in V0a2-knockdown resistant cells. Interestingly, pre-treatment with monoclonal V0a2-inhibitory antibody enhanced cisplatin cytotoxicity in resistant cells. Taken together, our findings suggest that the isoform specific inhibition of V-ATPase-V0a2 could serve as a therapeutic strategy for chemo-resistant ovarian carcinoma and improve efficacy of platinum drugs.

Breast cancer associated a2 isoform vacuolar ATPase immunomodulates neutrophils: potential role in tumor progression.

In invasive breast cancer, tumor associated neutrophils (TAN) represent a significant portion of the tumor mass and are associated with increased angiogenesis and metastasis. Identifying the regulatory factors that control TAN behavior will help in developing ideal immunotherapies. Vacuolar ATPases (V-ATPases), multi-subunit proton pumps, are highly expressed in metastatic breast cancer cells. A cleaved peptide from a2 isoform V-ATPase (a2NTD) has immunomodulatory role in tumor microenvironment. Here, we report for the first time the role of V-ATPase in neutrophils modulation. In invasive breast cancer cells, a2NTD was detected and a2V was highly expressed on the surface. Immunohistochemical analysis of invasive breast cancer tissues revealed that increased neutrophil recruitment and blood vessel density correlated with increased a2NTD levels. In order to determine the direct regulatory role of a2NTD on neutrophils, recombinant a2NTD was used for the treatment of neutrophils isolated from the peripheral blood of healthy volunteers. Neutrophils treated with a2NTD (a2Neuɸ) showed increased secretion of IL-1RA, IL-10, CCL-2 and IL-6 that are important mediators in cancer related inflammation. Moreover, a2Neuɸ exhibited an increased production of protumorigenic factors including IL-8, matrix metaloprotinase-9 and vascular endothelial growth factor. Further, functional characterization of a2Neuɸ revealed that a2Neuɸ derived products induce in vitro angiogenesis as well as increase the invasiveness of breast cancer cells. This study establishes the modulatory effect of breast cancer associated a2V on neutrophils, by the action of a2NTD, which has a positive impact on tumor progression, supporting that a2V can be a potential selective target for breast cancer therapy.

Monte Carlo simulation analysis of ceftobiprole, dalbavancin, daptomycin, tigecycline, linezolid and vancomycin pharmacodynamics against intensive care unit-isolated methicillin-resistant Staphylococcus aureus.

The aim of the present study was to compare the potential of ceftobiprole, dalbavancin, daptomycin, tigecycline, linezolid and vancomycin to achieve their requisite pharmacokinetic/pharmacodynamic (PK/PD) targets against methicillin-resistant Staphylococcus aureus isolates collected from intensive care unit (ICU) settings. Monte Carlo simulations were carried out to simulate the PK/PD indices of the investigated antimicrobials. The probability of target attainment (PTA) was estimated at minimum inhibitory concentration values ranging from 0.03 to 32 µg/mL to define the PK/PD susceptibility breakpoints. The cumulative fraction of response (CFR) was computed using minimum inhibitory concentration data from the Canadian National Intensive Care Unit study. Analysis of the simulation results suggested the breakpoints of 4 µg/mL for ceftobiprole (500 mg/2 h t.i.d.), 0.25 µg/mL for dalbavancin (1000 mg), 0.12 µg/mL for daptomycin (4 mg/kg q.d. and 6 mg/kg q.d.) and tigecycline (50 mg b.i.d.), and 2 µg/mL for linezolid (600 mg b.i.d.) and vancomycin (1 g b.i.d. and 1.5 g b.i.d.). The estimated CFR were 100, 100, 70.6, 88.8, 96.5, 82.4, 89.4, and 98.3% for ceftobiprole, dalbavancin, daptomycin (4 mg/kg/day), daptomycin (6 mg/kg/day), linezolid, tigecycline, vancomycin (1 g b.i.d.) and vancomycin (1.5 g b.i.d.), respectively. In conclusion, ceftobiprole and dalbavancin have the highest probability of achieving their requisite PK/PD targets against methicillin-resistant Staphylococcus aureus isolated from ICU settings. The susceptibility predictions suggested a reduction of the vancomycin breakpoint to 1 µg/mL.