Publications by authors named "Rakesh K Singh"

230 Publications

Polycation fluorination improves intraperitoneal siRNA delivery in metastatic pancreatic cancer.

J Control Release 2021 Mar 25;333:139-150. Epub 2021 Mar 25.

Center for Drug Delivery and Nanomedicine, Department of Pharmaceutical Sciences, University of Nebraska Medical Center, Omaha, NE 68198, USA. Electronic address:

Pancreatic ductal adenocarcinoma (PDAC) is a growing medical problem associated with extensive metastasis and high mortality. Intraperitoneal (IP) administration of therapeutics promises to help the treatment of cancers originated from organs in the peritoneal cavity. In this study, we evaluated how physicochemical properties of self-assembled polycation/siRNA nanoparticles affect their IP delivery efficacy in an orthotopic PDAC model. We have examined the effect of covalent polycation modification with lipophobic and hydrophobic tetrafluoro-p-toluic acid (TFTA), hydrophobic cholesterol, and hydrophilic poly(ethylene glycol) respectively. The surface charge of the three different nanoparticles was also modulated by coating the surface with serum albumin. We found that positively charged fluorine-containing particles with lipophobic properties based on a mixture of positively charged polymeric AMD3100 CXCR4 antagonist (PAMD) and PAMD modified with TFTA (mPAMD-TFTA)/siRNA displayed the best cell uptake and transfection efficacy in vitro. Biodistribution evaluation of the nanoparticles in a syngeneic orthotopic PDAC model revealed that the fluorine-containing formulation also achieved the highest PDAC tumor accumulation after IP administration. With a combination of CXCR4 inhibition by PAMD and PLK1 downregulation by siRNA, the treatment with mPAMD-TFTA/siPLK1 showed significant inhibition of both primary and metastatic PDAC tumors. Overall, our study provides insights into and guides the design of the nanoparticles for improved IP delivery of siRNA in PDAC.
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http://dx.doi.org/10.1016/j.jconrel.2021.03.028DOI Listing
March 2021

Nusinersen and Spinal Muscular Atrophies: Where are we in 2020?

Ann Indian Acad Neurol 2020 Nov-Dec;23(6):743-744. Epub 2020 Dec 18.

Department of Neurology, Grant Medical College and Sir J J Group of Hospitals, Mumbai, Maharashtra, India.

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http://dx.doi.org/10.4103/aian.AIAN_887_20DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7900727PMC
December 2020

Plexin-B3 Regulates Cellular Motility, Invasiveness, and Metastasis in Pancreatic Cancer.

Cancers (Basel) 2021 Feb 16;13(4). Epub 2021 Feb 16.

Department of Pathology and Microbiology, Nebraska Medical Center, Omaha, NE 68198, USA.

The Plexins family of proteins are well-characterized transmembrane receptors of semaphorins, axon guidance cue molecules, that mediate the cell attraction or repelling effects for such cues. Plexins and their ligands are involved in numerous cellular activities, such as motility, invasion, and adhesion to the basement membrane. The detachment of cells and the gain in motility and invasion are hallmarks of the cancer metastasis cascade, thus generating interest in exploring the role of plexins in cancer metastasis. Semaphorin-plexin complexes can act as tumor promoters or suppressors, depending upon the cancer type, and are under investigation for therapeutic purposes. Our group has identified Semaphorin-5A (SEMA5A)/Plexin-B3 as an attractive targetable complex for pancreatic cancer (PC) metastasis. However, our understanding of the Plexin-B3 function and pathological expression in PC is limited, and our present study delineates the role of Plexin-B3 in PC malignancy. We examined the pathological expression of Plexin-B3 in PC tumors and metastasis using a human tissue microarray, disease progression model of PDX-Cre-Kras (KC) mice, and different metastatic sites obtained from the Kras; Trp53; Pdx1-Cre (KPC) mice model. We observed a higher Plexin-B3 expression in PC tumor cores than the normal pancreas, and different metastatic sites were positive for Plexin-B3 expression. However, in the KC mice model, the Plexin-B3 expression increased initially and then decreased with the disease progression. Next, to evaluate the functional role of Plexin-B3, we utilized T3M-4- and CD18/HPAF-Control and -Plexin B3 knockdown cells for different in vivo and in vitro studies. The knockdown of Plexin-B3 enhanced the in vitro cellular migration, invasiveness, and impaired colony formation in three-dimensional culture, along with an increase in cellular spread and remodeling of the actin filaments. We also observed a higher metastasis in nude mice injected with T3M-4- and CD18/HPAF-shPlexin-B3 cells compared to their respective control cells. Furthermore, we observed a lower number of proliferating Ki-67-positive cells and higher ALDH1-A1-positive cells in the tumors formed by Plexin-B3 knockdown cells compared to tumors formed by the control cells. Together, our data suggest that the loss of Plexin-B3 is associated with the interference of cell division machinery and the induction of stem cell-like characteristics in PC cells.
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http://dx.doi.org/10.3390/cancers13040818DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7919786PMC
February 2021

Preliminary preclinical study of Chol-DsiRNA polyplexes formed with PLL[30]-PEG[5K] for the RNAi-based therapy of breast cancer.

Nanomedicine 2021 Feb 3;33:102363. Epub 2021 Feb 3.

Center for Drug Delivery and Nanomedicine, College of Pharmacy, University of Nebraska Medical Center, Omaha, NE, USA; Department of Pharmaceutical Sciences, College of Pharmacy, University of Nebraska Medical Center, Omaha, NE, USA. Electronic address:

RNA interference molecules have tremendous potential for cancer therapy but are limited by insufficient potency after i.v. administration. We previously found that Chol-DsiRNA polyplexes formed between cholesterol-modified dicer-substrate siRNA (Chol-DsiRNA) and the cationic diblock copolymer PLL[30]-PEG[5K] greatly increase the activity of Chol-DsiRNA against a stably expressed reporter mRNA in primary murine syngeneic breast tumors after daily i.v. dosing. Here, we provide a more thorough preliminary preclinical study of Chol-DsiRNA polyplexes against the therapeutically relevant target protein, STAT3. We found that Chol-DsiSTAT3 polyplexes greatly increase plasma exposure, distribution, potency, and therapeutic activity of Chol-DsiSTAT3 in primary murine syngeneic 4T1 breast tumors after i.v. administration. Furthermore, inactive Chol-DsiCTRL polyplexes are well tolerated by healthy female BALB/c mice after chronic i.v. administration at 50 mg Chol-DsiCTRL/kg over 28 days. Thus, Chol-DsiRNA polyplexes may be a good candidate for Phase I clinical trials to improve the treatment of breast cancer and other solid tumors.
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http://dx.doi.org/10.1016/j.nano.2021.102363DOI Listing
February 2021

Improving the survival of Lactobacillus plantarum NRRL B-1927 during microencapsulation with ultra-high-pressure-homogenized soymilk as a wall material.

Food Res Int 2021 Jan 26;139:109831. Epub 2020 Oct 26.

Department of Food Science and Technology, The University of Georgia, Griffin, GA 30223, USA. Electronic address:

Probiotic foods and supplements have been shown to offer multiple potential health benefits to consumers. Dried probiotic cultures are increasingly used by the food industry because they are easily handled, transported, stored, and used in different applications. However, drying technologies often expose probiotic cells to extreme environmental conditions that reduces cell viability. Hence, this study aimed to evaluate the effect of using ultra high-pressure homogenization (UHPH) on soymilk's microencapsulating ability, and the resultant effect on the survivability of probiotic Lactobacillus plantarum NRRL B-1927 (LP) during drying. Liquid suspensions containing LP (~10 CFU/g of solids) were prepared by suspending LP cultures in soymilk which had been either treated with UHPH at 150 MPa or 300 MPa or left untreated. LP suspensions were then dried by concurrent spray drying (CCSD), mixed-flow spray drying (MXSD) or freeze-drying (FD). Cell counts of LP were determined before and after microencapsulation. Moisture, water activity, particle size and morphology of LP powders were also characterized. LP powders produced with 300 MPa treated soymilk had 8.7, 6.4, and 2 times more cell counts than those produced with non-UHPH treated soymilk during CCSD, MXSD, and FD, respectively. In the 300 MPa treated samples, cell survival (%) of LP during drying was the highest in MXSD (83.72) followed by FD (76.31) and CCSD (34.01). Using soymilk treated at higher UHPH pressures resulted in LP powders with lower moisture content, smaller particle sizes and higher agglomeration. LP powders produced via MXSD showed higher agglomeration and fewer signs of thermal damage than powders produced via CCSD. This study demonstrates that UHPH improves the effectiveness of soymilk as a microencapsulant for probiotics, creating probiotic powders that could be used in plant-based and non-dairy foods.
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http://dx.doi.org/10.1016/j.foodres.2020.109831DOI Listing
January 2021

Host Cxcr2-Dependent Regulation of Pancreatic Cancer Growth, Angiogenesis, and Metastasis.

Am J Pathol 2021 04 14;191(4):759-771. Epub 2021 Jan 14.

Department of Pathology and Microbiology, Nebraska Medical Center, Omaha, Nebraska. Electronic address:

Pancreatic ductal adenocarcinoma (PDAC) manifests aggressive tumor growth and early metastasis. Crucial steps in tumor growth and metastasis are survival, angiogenesis, invasion, and immunosuppression. Our prior research showed that chemokine CXC- receptor-2 (CXCR2) is expressed on endothelial cells, innate immune cells, and fibroblasts, and regulates angiogenesis and immune responses. Here, we examined whether tumor angiogenesis, growth, and metastasis of CXCR2 ligands expressing PDAC cells are regulated in vivo by a host CXCR2-dependent mechanism. C57BL6 Cxcr2 mice were generated following crosses between Cxcr2 female and Cxcr2 male. Cxcr2 ligands expressing Kirsten rat sarcoma (KRAS-PDAC) cells were orthotopically implanted in the pancreas of wild-type or Cxcr2 C57BL6 mice. No significant difference in PDAC tumor growth was observed. Host Cxcr2 loss led to an inhibition in microvessel density in PDAC tumors. Interestingly, an enhanced spontaneous and experimental liver metastasis was observed in Cxcr2 mice compared with wild-type mice. Increased metastasis in Cxcr2 mice was associated with an increase in extramedullary hematopoiesis and expansion of neutrophils and immature myeloid precursor cells in the spleen of tumor-bearing mice. These data suggest a dynamic role of host CXCR2 axis in regulating tumor immune suppression, tumor growth, and metastasis.
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http://dx.doi.org/10.1016/j.ajpath.2021.01.002DOI Listing
April 2021

Intracerebral Schwannoma with Perivascular Meningioangiomatosis-like Extension in Surrounding Glia.

Neurol India 2020 Nov-Dec;68(6):1497-1499

Department of Pathology, Dr. Ram Manohar Lohia Institute of Medical Sciences, Gomti Nagar, Lucknow, Uttar Pradesh, India.

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http://dx.doi.org/10.4103/0028-3886.304096DOI Listing
December 2020

Neutrophil Gelatinase-Associated Lipocalin Protects Acinar Cells From Cerulein-Induced Damage During Acute Pancreatitis.

Pancreas 2020 Nov/Dec;49(10):1297-1306

From the Department of Biochemistry and Molecular Biology.

Objectives: Elevated neutrophil gelatinase-associated lipocalin (NGAL) is a promising marker for severe acute pancreatitis (SAP) and multiple organ failure, suggesting systemic and local contributions during pancreatitis. We investigated the role of NGAL locally on acinar cell biology.

Methods: Western blot, reverse transcriptase-polymerase chain reaction, and immunohistochemistry analysis were performed to analyze the levels of NGAL receptors, apoptotic and regeneration markers, and 4-hydroxynonenal (4HNE) levels, 3-[4,5-Dimethylthiazole-2-yl]-2, 5-diphenyltetrazolium bromide assay, and annexin V/propidium iodide staining were used to evaluate cell viability, and effect on endothelial cells was accessed by endothelial permeability assay.

Results: Cerulein treatment at 20 μM for 12 hours significantly reduced acinar cell viability by 40%, which was rescued by NGAL at 800 and 1600 ng/mL concentrations, observed during mild and SAP, respectively. Mechanistically, NGAL significantly reduced the levels of reactive oxygen species and 4HNE adduct formation in a 24p3R-dependent manner and upregulated the expression of acinar cell regeneration markers, like CDK-2, CDK-4, and C-myc. However, SAP levels of NGAL significantly increased endothelial permeability and downregulated the levels of ZO-1, and cerulein treatment in NGAL knockout mice showed increased levels of 4HNE adducts.

Conclusions: Neutrophil gelatinase-associated lipocalin rescues intracellular reactive oxygen species during pancreatitis and promotes survival and regeneration of acinar cells.
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http://dx.doi.org/10.1097/MPA.0000000000001690DOI Listing
October 2020

"Trained immunity" from Mycobacterium spp. exposure or BCG vaccination and COVID-19 outcomes.

PLoS Pathog 2020 10 29;16(10):e1008969. Epub 2020 Oct 29.

Department of Biochemistry, Institute of Science, Banaras Hindu University, Varanasi, India.

Protective variables for Coronavirus Disease 2019 (COVID-19) are unknown. "Trained immunity" of the populace as a result of Bacille Calmette-Guérin (BCG) vaccination policy implementation and coverage had been suggested to be one of the factors responsible for the differential impact of COVID-19 on different countries. Several trials are underway to evaluate the potential protective role of BCG vaccination in COVID-19. However, the lack of clarity on the use of appropriate controls concerning the measures of "trained immunity" or the heterologous cell-mediated immunity conferred by BCG vaccination has been a cause of concern leading to more confusion as exemplified by a recently concluded trial in Israel that failed to find any protective correlation with regard to BCG vaccination. Whereas, when we analyze the COVID-19 epidemiological data of European countries without any regard for BCG vaccination policy but with similar age distribution, comparable confounding variables, and the stage of the pandemic, the prevalence of tuberculin immunoreactivity-a measure of cell-mediated immunity persistence as a result of Mycobacterium spp. (including BCG vaccine) exposure of the populations-is found consistently negatively correlated with COVID-19 infections and mortality. We seek to draw attention toward the inclusion of controls for underlying "trained immunity" and heterologous cell-mediated immunity prevalence that may be preexisting or resulting from the intervention (e.g., BCG vaccine) in such trials to arrive at more dependable conclusions concerning potential benefit from them.
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http://dx.doi.org/10.1371/journal.ppat.1008969DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7595322PMC
October 2020

Myelin Oligodendrocyte Glycoprotein (MOG) Antibody-Associated CNS Demyelination: Clinical Spectrum and Comparison with Aquaporin-4 Antibody Positive Neuromyelitis Optica Spectrum Disorder.

Neurol India 2020 Sep-Oct;68(5):1106-1114

Department of Neurology, Grant Government Medical College, Mumbai, Maharashtra, India.

Background: The clinical phenotypes of myelin oligodendrocyte glycoprotein (MOG) antibody disease, its disease course, and treatment are poorly understood and much work needs to be done towards this.

Objective: To characterize the clinico-radiologic spectrum and treatment outcomes of MOG antibody disease and differentiate it from aquaporin-4 (AQP-4) antibody positive neuromyelitis optica spectrum disorders (NMO-SD).

Methods: A single-center, observational study from Western India during 2017-2019, of 48 patients with either MOG antibody positive (21 patients) or AQP-4 antibody positive (27 patients) central nervous system demyelination.

Results: MOG antibody group had median age 32.2 years, no gender bias, median disease duration 40 months, relapses in 9 patients (43%), and median 2.5 (1-16) episodes per patient. Onset phenotypes included isolated bilateral optic neuritis (ON) (43%), isolated unilateral ON (19%), acute brainstem syndrome (19%), simultaneous ON with myelitis (9%), isolated myelitis (5%), and acute disseminated encephalomyelitis optic neuritis (ADEM-ON) (5%). Characteristic neuroimaging abnormalities were anterior segment longitudinally extensive ON, upper brainstem, and thoracic cord involvement (both short and long segment lesions). Most patients (86%) responded well to steroids, only 3/21 required rescue immunotherapy. In total, 6 out of 46 eyes affected developed permanent visual disability, while one patient had motor disability. The features differentiating MOG from AQP-4 antibody group were: no female predilection, preferential optic nerve involvement, characteristic neuroimaging abnormalities, and favorable therapeutic response and outcome.

Conclusions: MOG disease commonly presents as severe ON, myelitis, acute brainstem syndrome, ADEM or their combinations. Early identification, treatment, and maintenance immunosuppression are necessary. It can easily be differentiated from NMO-SD using clinico-radiological features and therapeutic response.
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http://dx.doi.org/10.4103/0028-3886.294831DOI Listing
October 2020

Breast Cancer Cell-Neutrophil Interactions Enhance Neutrophil Survival and Pro-Tumorigenic Activities.

Cancers (Basel) 2020 Oct 8;12(10). Epub 2020 Oct 8.

Department of Pathology and Microbiology, University of Nebraska Medical Center, 985900 UNMC, Omaha, NE 68198-5900, USA.

Breast cancer remains the most prevalent cancer in women with limited treatment options for patients suffering from therapy-resistance and metastatic disease. Neutrophils play an important role in breast cancer progression and metastasis. We examined the pro-tumorigenic nature of the breast cancer cell-neutrophil interactions and delineated the differences in neutrophil properties between the chemotherapy-resistant and the parent tumor microenvironment. Our data demonstrated that high neutrophil infiltration is associated with disease aggressiveness and therapy resistance. In the human breast cancer dataset, expression of neutrophil-related signature gene expression was higher in tumors from therapy-resistant patients than therapy-sensitive patients. We observed that breast cancer-derived factors significantly enhanced neutrophil survival, polarization, and pro-inflammatory cytokine expression. Breast cancer cell-derived supernatant treated neutrophils significantly expressed high levels of interleukin-1β (IL-1β), CC-chemokine ligand-2-4 (CCL2, CCL3, CCL4), inducible nitric oxide synthase (iNOS), and matrix metallopeptidase-9 (MMP9), and formed extracellular traps (NETs). Moreover, neutrophils showed increased secretion of MMP9 when cultured with the supernatant of chemotherapy-resistant Cl66-Doxorubicin (Cl66-Dox) and Cl66-Paclitaxel (Cl66-Pac) cells in comparison with the supernatant of Cl66-parent cells. Together, these data suggest an important role of breast cancer cell-neutrophil interactions in regulating pro-tumor characteristics in neutrophils and its modulation by therapy resistance.
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http://dx.doi.org/10.3390/cancers12102884DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7599756PMC
October 2020

Cardiac Findings in Pediatric Patients With Multisystem Inflammatory Syndrome in Children Associated With COVID-19.

Clin Pediatr (Phila) 2021 02 25;60(2):119-126. Epub 2020 Sep 25.

Division of Pediatric Cardiology, Hassenfeld Children's Hospital at NYU Langone, New York, NY, USA.

Background: The overall severity of cardiac disease secondary to acute SARS-CoV-2 (severe acute respiratory syndrome coronavirus-2) infection in children appears to be much lower when compared with adults. However, the newly described multisystem inflammatory syndrome in children (MIS-C) associated with coronavirus disease 2019 (COVID-19) has been associated with cardiac complications.

Methods: We reviewed the clinical course and cardiac testing results in pediatric patients hospitalized with MIS-C at 2 large hospital systems in the New York City metropolitan area over a 3-month period.

Results: Of the 33 patients (median age 2.8 years) in the study cohort, 24 (73%) had at least one abnormality in cardiac testing: abnormal electrocardiogram (48%), elevated brain natriuretic peptide (43%), abnormal echocardiogram (30%), and/or elevated troponin (21%). Electrocardiogram and echocardiogram abnormalities all resolved by the 2-week outpatient follow-up cardiology visit.

Conclusion: While 73% of pediatric patients with MIS-C had evidence of abnormal cardiac testing on hospital admission in our study, all cardiac testing was normal by outpatient hospital discharge follow-up. Cardiac screening tests should be performed in all patients diagnosed with MIS-C given the high rate of abnormal cardiac findings in our study cohort.
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http://dx.doi.org/10.1177/0009922820961771DOI Listing
February 2021

Surveillance for cardiac allograft vasculopathy: Practice variations among 50 pediatric heart transplant centers.

J Heart Lung Transplant 2020 Aug 8. Epub 2020 Aug 8.

Department of Pediatrics, Division of Cardiology, Ann & Robert H. Lurie Children's Hospital of Chicago, Chicago, Illinois.

Background: Coronary allograft vasculopathy (CAV) is a leading cause of mortality after heart transplantation (HT) in children. Variation in CAV screening practices may impact detection rates and patient outcomes.

Methods: Among 50 Pediatric Heart Transplant Society (PHTS) sites from 2001 to 2016, coronary evaluations were classified as angiography or non-invasive testing, and angiograms were designated as routine or symptom based. CAV detection rates stratified by routine vs symptom-based angiograms were calculated. Freedom from CAV and mortality after CAV diagnosis, stratified by study indication, were calculated.

Results: A total of 3,442 children had 13,768 coronary evaluations; of these, 97% (n = 13,012) were for routine surveillance, and only 3% (n = 333) were for cause. Over the study period, CAV was detected in 472 patients (14%). Whereas 58% (n = 29) of PHTS sites evaluate by angiography alone, 42% reported supplementing with a non-invasive test, although only 423 non-invasive studies were reported. Angiographic detection of CAV was higher for symptom-based testing than for routine testing (29% vs 4%, p < 0.0001), although routine testing identified a majority of cases (88%; n = 414). The 10-year freedom from CAV was 77% overall. Once CAV is detected, 5-year graft survival was 58%, with lower survival for patients diagnosed after symptoms angiogram than after routine angiogram (30% vs 62%; p < 0.0001).

Conclusions: Development of a robust model for CAV risk should allow low-risk patients to undergo less frequent invasive angiography without adverse impact on CAV detection rates or outcomes.
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http://dx.doi.org/10.1016/j.healun.2020.08.003DOI Listing
August 2020

Aortic Root Thrombosis on ECMO-A Novel Management Strategy.

World J Pediatr Congenit Heart Surg 2020 09;11(5):643-645

Division of Pediatric Cardiology, Department of Pediatrics, 14444Rady Children's Hospital, University of California, San Diego, CA, USA.

A 15-year-old presented in cardiogenic shock secondary to viral myocarditis requiring venoarterial extracorporeal membrane oxygenation (ECMO) support. He developed large thrombi of the left ventricle and aortic root. Anticoagulation was increased, and medications were initiated to decrease the likelihood of aortic valve opening. He underwent balloon atrial septostomy followed by placement of a left atrial vent. A pigtail catheter was placed in the ascending aorta for direct heparin infusion. Serial echocardiograms showed progressive resolution of the thrombi. He was successfully weaned from ECMO and discharged home without neurological deficits.
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http://dx.doi.org/10.1177/2150135120924416DOI Listing
September 2020

Neuroprotection of Rotenone-Induced Parkinsonism by Ursolic Acid in PD Mouse Model.

CNS Neurol Disord Drug Targets 2020 ;19(7):527-540

Department of Biochemistry, Institute of Science, Banaras Hindu University, Varanasi-221005, India.

Background: Parkinson's Disease (PD) is characterized by both motor and non-motor symptoms. The presynaptic neuronal protein, α-Synuclein, plays a pivotal role in PD pathogenesis and is associated with both genetic and sporadic origin of the disease. Ursolic Acid (UA) is a well-known bioactive compound found in various medicinal plants, widely studied for its anti-inflammatory and antioxidant activities.

Objective: In this research article, the neuroprotective potential of UA has been further explored in the Rotenone-induced mouse model of PD.

Methods: To investigate our hypothesis, we have divided mice into 4 different groups, control, drug only control, Rotenone-intoxicated group, and Rotenone-intoxicated mice treated with UA. After the completion of dosing, behavioral parameters were estimated. Then mice from each group were sacrificed and the brains were isolated. Further, the biochemical tests were assayed to check the balance between the oxidative stress and endogenous anti-oxidants; and TH (Tyrosine Hydroxylase), α-Synuclein, Akt (Serine-threonine protein kinase), ERK (Extracellular signal-regulated kinase) and inflammatory parameters like Nuclear Factor-κB (NF-κB) and Tumor Necrosis Factor- α (TNF-α) were assessed using Immunohistochemistry (IHC). Western blotting was also done to check the expressions of TH and α-Synuclein. Moreover, the expression levels of PD related genes like α-Synuclein, β-Synuclein, Interleukin-1β (IL-1β), and Interleukin-10 (IL-10) were assessed by using Real-time PCR.

Results: The results obtained in our study suggested that UA significantly reduced the overexpression of α-Synuclein and regulated the phosphorylation of survival-related kinases (Akt and ERK) apart from alleviating the behavioral abnormalities and protecting the dopaminergic neurons from oxidative stress and neuroinflammation.

Conclusion: Thus, our study shows the neuroprotective potential of UA, which can further be explored for possible clinical intervention.
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http://dx.doi.org/10.2174/1871527319666200812224457DOI Listing
January 2020

Identification of novel Urotensin-II receptor antagonists with potent inhibition of U-II induced pressor response in mice.

Eur J Pharmacol 2020 Nov 1;886:173391. Epub 2020 Aug 1.

Cell Therapy Laboratories, Daiichi Sankyo Co., Limited, Tokyo, Japan.

Urotensin II (U-II) has been found to be one of the most potent vasoconstrictor (Ames et al., 1999; Bohm et al., 2002) reported till date. U-II exerts its response via activation of a G-protein coupled receptor, Urotensin II receptor(UT). Binding of U-II to UT leads to an instant increase in the inositol phosphate turnover and intracellular Ca. Such an instant Ca release and potent vasoconstriction exerted by U-II is expected to have an important role in the progression of cardiac diseases. We have previously shown that UT antagonist DS37001789 prevents U-II induced blood pressure elevation in mice (Nishi et al., 2019) in a dose dependent manner, with potent efficacy at 30 and 100 mg/kg. Further to this, we have also shown that DS37001789 ameliorates mortality in pressure-overload mice with heart failure (Nishi et al., 2020). We therefore conducted an extensive structure-activity relationship studies to identify molecules with superior efficacy. In the present manuscript, we report the identification of two potent, non-peptide small molecule antagonists of Urotensin II receptor (UT), RCI-0879 and RCI-0298 which blocked the action of U-II, both in vitro and in vivo. These molecules were found to be very potent in in vitro Ca and radioligand binding assays using human and mouse UT over-expressing CHO cells. RCI-0879 and RCI-0298 also exhibited superior efficacy in in vivo mouse pressor response model using C57BL/6 mice, compared to our initial molecules (Nishi et al., 2019) and demonstrated ED values of 3.2 mg/kg and 6.8 mg/kg respectively. Our findings reported herewith, further strengthen our concept and belief in UT antagonization as a potential therapeutic approach for the management of chronic heart failure.
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http://dx.doi.org/10.1016/j.ejphar.2020.173391DOI Listing
November 2020

CXCR2 signaling promotes secretory cancer-associated fibroblasts in pancreatic ductal adenocarcinoma.

FASEB J 2020 07 26;34(7):9405-9418. Epub 2020 May 26.

Department of Pathology and Microbiology, University of Nebraska Medical Center, Omaha, NE, USA.

Pancreatic ductal adenocarcinoma (PDAC) remains one of the most challenging malignancies. Desmoplasia and tumor-supporting inflammation are hallmarks of PDAC. The tumor microenvironment contributes significantly to tumor progression and spread. Cancer-associated fibroblasts (CAFs) facilitate therapy resistance and metastasis. Recent reports emphasized the concurrence of multiple subtypes of CAFs with diverse roles, fibrogenic, and secretory. C-X-C motif chemokine receptor 2 (CXCR2) is a chemokine receptor known for its role during inflammation and its adverse role in PDAC. Oncogenic Kras upregulates CXCR2 and its ligands and, thus, contribute to tumor proliferation and immunosuppression. CXCR2 deletion in a PDAC syngeneic mouse model produced increased fibrosis revealing a potential undescribed role of CXCR2 in CAFs. In this study, we demonstrate that the oncogenic Kras-CXCR2 axis regulates the CAFs function in PDAC and contributes to CAFs heterogeneity. We observed that oncogenic Kras and CXCR2 signaling alter CAFs, producing a secretory CAF phenotype with low fibrogenic features; and increased secretion of pro-tumor cytokines and CXCR2 ligands, utilizing the NF-κB activity. Finally, using syngeneic mouse models, we demonstrate that oncogenic Kras is associated with secretory CAFs and that CXCR2 inhibition promotes activation of fibrotic cells (myofibroblasts) and impact tumors in a mutation-dependent manner.
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http://dx.doi.org/10.1096/fj.201902990RDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7501205PMC
July 2020

The biomarker HE4 (WFDC2) promotes a pro-angiogenic and immunosuppressive tumor microenvironment via regulation of STAT3 target genes.

Sci Rep 2020 05 22;10(1):8558. Epub 2020 May 22.

Women and Infants Hospital, Department of Obstetrics and Gynecology, Program in Women's Oncology, Providence, RI, USA.

Epithelial ovarian cancer (EOC) is a highly lethal gynecologic malignancy arising from the fallopian tubes that has a high rate of chemoresistant recurrence and low five-year survival rate. The ovarian cancer biomarker HE4 is known to promote proliferation, metastasis, chemoresistance, and suppression of cytotoxic lymphocytes. In this study, we sought to examine the effects of HE4 on signaling within diverse cell types that compose the tumor microenvironment. HE4 was found to activate STAT3 signaling and promote upregulation of the pro-angiogenic STAT3 target genes IL8 and HIF1A in immune cells, ovarian cancer cells, and endothelial cells. Moreover, HE4 promoted increases in tube formation in an in vitro model of angiogenesis, which was also dependent upon STAT3 signaling. Clinically, HE4 and IL8 levels positively correlated in ovarian cancer patient tissue. Furthermore, HE4 serum levels correlated with microvascular density in EOC tissue and inversely correlated with cytotoxic T cell infiltration, suggesting that HE4 may cause deregulated blood vessel formation and suppress proper T cell trafficking in tumors. Collectively, this study shows for the first time that HE4 has the ability to affect signaling events and gene expression in multiple cell types of the tumor microenvironment, which could contribute to angiogenesis and altered immunogenic responses in ovarian cancer.
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http://dx.doi.org/10.1038/s41598-020-65353-xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7244765PMC
May 2020

Role of trypsin and protease-activated receptor-2 in ovarian cancer.

PLoS One 2020 4;15(5):e0232253. Epub 2020 May 4.

The Wilmot Cancer Institute at the University of Rochester Medical Center, Rochester, NY, United States of America.

Proteases have been implicated in the tumorigenesis and aggressiveness of a variety of cancer types. In fact, proteases have proven to be very clinically useful as tumor biomarkers in the blood of patients. Proteases are typically involved in complex systems of substrates, activators, and inhibitors, thus making our ability to establish their exact function in cancer more difficult. Trypsin, perhaps the most famous of proteases, has been shown to play a role in cancer progression, but its functional role in ovarian cancer has not been much studied. PAR2, a transmembrane receptor that is known to be activated by trypsin, has been reported to be associated with ovarian cancer. Here, we found that stimulation of ovarian cancer cell lines with trypsin or PAR2 activating peptide markedly increased MAPK signaling and cell proliferation. Additionally, HE4, a WAP-family glycoprotein and ovarian cancer biomarker, was found to inhibit trypsin degradation, thereby retaining its activity. Patient data seemed to support this phenomenon, as the serum of ovarian cancer patients with high HE4 expression, revealed significantly elevated trypsin levels. These data support the hypothesis that trypsin plays a tumorigenic role in ovarian cancer, which can be mediated by its receptor PAR2, and potentiated by HE4.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0232253PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7197761PMC
July 2020

Skeletal Muscle Protein Composition Adaptations to 10 Weeks of High-Load Resistance Training in Previously-Trained Males.

Front Physiol 2020 26;11:259. Epub 2020 Mar 26.

School of Kinesiology, Auburn University, Auburn, AL, United States.

While high-load resistance training increases muscle hypertrophy, the intramuscular protein responses to this form of training remains largely unknown. In the current study, recreationally resistance-trained college-aged males ( = 15; mean ± SD: 23 ± 3 years old, 6 ± 5 years training) performed full-body, low-volume, high-load [68-90% of one repetition maximum (1RM)] resistance training over 10 weeks. Back squat strength testing, body composition testing, and a vastus lateralis biopsy were performed before (PRE) and 72 h after the 10-week training program (POST). Fiber type-specific cross-sectional area (fCSA), myofibrillar protein concentrations, sarcoplasmic protein concentrations, myosin heavy chain and actin protein abundances, and muscle tissue percent fluid were analyzed. The abundances of individual sarcoplasmic proteins in 10 of the 15 participants were also assessed using proteomics. Significant increases ( < 0.05) in type II fCSA and back squat strength occurred with training, although whole-body fat-free mass paradoxically decreased ( = 0.026). No changes in sarcoplasmic protein concentrations or muscle tissue percent fluid were observed. Myosin heavy chain protein abundance trended downward (-2.9 ± 5.8%, = 0.069) and actin protein abundance decreased (-3.2 ± 5.3%, = 0.034) with training. Proteomics indicated only 13 sarcoplasmic proteins were altered with training (12 up-regulated, 1 down-regulated, < 0.05). Bioinformatics indicated no signaling pathways were affected, and proteins involved with metabolism (e.g., ATP-PCr, glycolysis, TCA cycle, or beta-oxidation) were not affected. These data comprehensively describe intramuscular protein adaptations that occur following 10 weeks of high-load resistance training. Although previous data from our laboratory suggests high-volume resistance training enhances the ATP-PCr and glycolytic pathways, we observed different changes in metabolism-related proteins in the current study with high-load training.
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http://dx.doi.org/10.3389/fphys.2020.00259DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7135893PMC
March 2020

Healthy and Resilient Cereals and Pseudo-Cereals for Marginal Agriculture: Molecular Advances for Improving Nutrient Bioavailability.

Front Genet 2020 27;11:49. Epub 2020 Feb 27.

Crop Diversification and Genetics Program, International Center for Biosaline Agriculture, Dubai, United Arab Emirates.

With the ever-increasing world population, an extra 1.5 billion mouths need to be fed by 2050 with continuously dwindling arable land. Hence, it is imperative that extra food come from the marginal lands that are expected to be unsuitable for growing major staple crops under the adverse climate change scenario. Crop diversity provides right alternatives for marginal environments to improve food, feed, and nutritional security. Well-adapted and climate-resilient crops will be the best fit for such a scenario to produce seed and biomass. The minor millets are known for their high nutritional profile and better resilience for several abiotic stresses that make them the suitable crops for arid and salt-affected soils and poor-quality waters. Finger millet () and foxtail millet (), also considered as orphan crops, are highly tolerant grass crop species that grow well in marginal and degraded lands of Africa and Asia with better nutritional profile. Another category of grains, called pseudo-cereals, is considered as rich foods because of their protein quality and content, high mineral content, and healthy and balance food quality. Quinoa (), amaranth ( sp.), and buckwheat () fall under this category. Nevertheless, both minor millets and pseudo-cereals are morphologically different, although similar for micronutrient bioavailability, and their grains are gluten-free. The cultivation of these millets can make dry lands productive and ensure future food as well as nutritional security. Although the natural nutrient profile of these crop plant species is remarkably good, little development has occurred in advances in molecular genetics and breeding efforts to improve the bioavailability of nutrients. Recent advances in NGS have enabled the genome and transcriptome sequencing of these millets and pseudo-cereals for the faster development of molecular markers and application in molecular breeding. Genomic information on finger millet (1,196 Mb with 85,243 genes); , a model small millet (well-annotated draft genome of 420 Mb with 38,801 protein-coding genes); amaranth (466 Mb genome and 23,059 protein-coding genes); buckwheat (genome size of 1.12 Gb with 35,816 annotated genes); and quinoa (genome size of 1.5 Gb containing 54,438 protein-coding genes) could pave the way for the genetic improvement of these grains. These genomic resources are an important first step toward genetic improvement of these crops. This review highlights the current advances and available resources on genomics to improve nutrient bioavailability in these five suitable crops for the sustained healthy livelihood.
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http://dx.doi.org/10.3389/fgene.2020.00049DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7056906PMC
February 2020

Development of Potent Forchlorfenuron Analogs and Their Cytotoxic Effect in Cancer Cell Lines.

Sci Rep 2020 02 24;10(1):3241. Epub 2020 Feb 24.

The Wilmot Cancer Institute at the University of Rochester Medical Center, Rochester, NY, United States.

Forchlorfenuron (FCF) is a synthetic plant cytokinin widely used in agriculture to promote fruit size, that paradoxically inhibits proliferation, migration, and invasion in human cancer cell lines. FCF has also been shown to affect HIF-1α and HER2, which are both known to play a crucial role in cancer cell survival. In this study, we have developed potent FCF analogs through structural modification of FCF, coined UR214-1, UR214-7, and UR214-9. Compared to parental FCF, these analogs are more effective in decreasing viability and proliferation in both ovarian and endometrial cancer cell lines. These FCF analogs also suppress HER2 expression at a concentration lower than that of FCF. In addition, we found that treatment with either FCF or its analogs decreases the expression of human epididymis protein 4 (HE4), which is commonly upregulated in ovarian and endometrial cancers. Given the association between cancer behavior and HE4 production in gynecologic cancers, our findings may provide insight useful in the development of new treatment strategies for gynecologic cancers.
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http://dx.doi.org/10.1038/s41598-020-59824-4DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7039965PMC
February 2020

The CD200-CD200R cross-talk helps Leishmania donovani to down regulate macrophage and CD4CD44 T cells effector functions in an NFκB independent manner.

Int J Biol Macromol 2020 May 19;151:394-401. Epub 2020 Feb 19.

Department of Biochemistry, Institute of Science, Banaras Hindu University, Varanasi 221 005, India. Electronic address:

The lacuna in the knowledge of immunobiology, especially in visceral infections that are fatal if left untreated, are a major hurdle in getting a vaccine candidate for leishmaniasis. Till date, only a few drugs are available to combat human leishmaniasis and a vaccine candidate either prophylactic or preventive is still awaited. Therefore, identification of host and parasitic factors involved in the regulation of specific immune mechanisms are essentially needed. In this study, we observed that CD200-CD200R immune inhibitory axis regulates host macrophages effectors properties and helps antigen experienced T cells (CD4CD44 T cells) to acquire anti-inflammatory cytokines (IL-4, IL-10, TGF-β, IL-27) producing abilities in an NFkB independent manner. After CD200 blocking the macrophages effectively inhibited proliferation of Leishmania amastigotes and also induced the production of IL-12, IFN-γ, TNF-α and nitric oxide (NOx). Further, the blocking of CD200 signaling also restored macrophages MHC-II expression and helped CD4CD44 T cells to produce pro-inflammatory cytokines like IL-2, IL-12 and IFN-γ. The finding of this study suggested the importance of immune inhibitory mechanisms in controlling Leishmania growth and survival and therefore, requires more studies to understand its role in vaccine induced immunity.
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http://dx.doi.org/10.1016/j.ijbiomac.2020.02.189DOI Listing
May 2020

Neutrophils in the Tumor Microenvironment.

Adv Exp Med Biol 2020 ;1224:1-20

Department of Pathology and Microbiology, University of Nebraska Medical Center, Omaha, NE, USA.

Neutrophils are the first responders to inflammation, infection, and injury. As one of the most abundant leukocytes in the immune system, neutrophils play an essential role in cancer progression, through multiple mechanisms, including promoting angiogenesis, immunosuppression, and cancer metastasis. Recent studies demonstrating elevated neutrophil to lymphocyte ratios suggest neutrophil as a potential therapeutic target and biomarker for disease status in cancer. This chapter will discuss the phenotypic and functional changes in the neutrophil in the tumor microenvironment, the underlying mechanism(s) of neutrophil facilitated cancer metastasis, and clinical potential of neutrophils as a prognostic/diagnostic marker and therapeutic target.
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http://dx.doi.org/10.1007/978-3-030-35723-8_1DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7325741PMC
February 2020

Role of connexins in female reproductive system and endometriosis.

J Gynecol Obstet Hum Reprod 2020 Jun 1;49(6):101705. Epub 2020 Feb 1.

Department of Bioscience and Biotechnolgy, Banasthali Vidyapith, Banasthali, 304022, Rajasthan, India. Electronic address:

Gap junction form channels between the cells and facilitate the function of cellular cross talk. Connexins, the gap junction proteins play an essential role in female reproductive health and its expression anomalies are correlated with female reproductive disorders like polycystic ovarian syndrome, recurrent miscarriage, pre-term birth and endometriosis. Endometriosis is a chronic gynecologic disorder caused by ectopic endometrial lesions growing outside the uterine cavity. Embryonic implantation is adversely affected in case of endometriosis leading to infertility. Endometriosis also interferes with ovulatory functions, reduces fertilization and impaires blastocyst implantation. There lies a lacunae in understanding of the role of gap junctions protein connexins in endometriosis. Therefore, this study discusses the role of connexins in improving female fertility by taming the processes of oogenesis, germ line development, uterine receptivity, placental growth, implantation, decidualization and concludes by focusing the role of connexins in endometriosis.
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http://dx.doi.org/10.1016/j.jogoh.2020.101705DOI Listing
June 2020

Recent Trends in the Management of Alzheimer's Disease: Current Therapeutic Options and Drug Repurposing Approaches.

Authors:
Rakesh K Singh

Curr Neuropharmacol 2020 ;18(9):868-882

Department of Pharmacology, Amity Institute of Pharmacy, Amity University, Manesar, Gurgaon-122413, Haryana, India

Alzheimer's disease is one of the most progressive forms of dementia, ultimately leading to death in aged populations. The major hallmarks of Alzheimer's disease include deposition of extracellular amyloid senile plaques and intracellular neurofibrillary tangles in brain neuronal cells. Although there are classical therapeutic options available for the treatment of the diseases, however, they provide only a symptomatic relief and do not modify the molecular pathophysiological course of the disease. Recent research advances in Alzheimer's disease have highlighted the potential role of anti-amyloid, anti-tau, and anti-inflammatory therapies. However, these therapies are still in different phases of pre-clinical/clinical development. In addition, drug repositioning/repurposing is another interesting and promising approach to explore rationalized options for the treatment of Alzheimer's disease. This review discusses the different aspects of the pathophysiological mechanism involved in the progression of Alzheimer's disease along with the limitations of current therapies. Furthermore, this review also highlights emerging investigational drugs along with recent drug repurposing approaches for Alzheimer's disease.
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http://dx.doi.org/10.2174/1570159X18666200128121920DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7569317PMC
January 2020

Skeletal Muscle Myofibrillar Protein Abundance Is Higher in Resistance-Trained Men, and Aging in the Absence of Training May Have an Opposite Effect.

Sports (Basel) 2020 Jan 10;8(1). Epub 2020 Jan 10.

School of Kinesiology, Auburn University, Auburn, AL 36849, USA.

Resistance training generally increases skeletal muscle hypertrophy, whereas aging is associated with a loss in muscle mass. Interestingly, select studies suggest that aging, as well as resistance training, may lead to a reduction in the abundance of skeletal muscle myofibrillar (or contractile) protein (per mg tissue). Proteomic interrogations have also demonstrated that aging, as well as weeks to months of resistance training, lead to appreciable alterations in the muscle proteome. Given this evidence, the purpose of this small pilot study was to examine total myofibrillar as well as total sarcoplasmic protein concentrations (per mg wet muscle) from the vastus lateralis muscle of males who were younger and resistance-trained (denoted as YT, n = 6, 25 ± 4 years old, 10 ± 3 self-reported years of training), younger and untrained (denoted as YU, n = 6, 21 ± 1 years old), and older and untrained (denoted as OU, n = 6, 62 ± 8 years old). The relative abundances of actin and myosin heavy chain (per mg tissue) were also examined using SDS-PAGE and Coomassie staining, and shotgun proteomics was used to interrogate the abundances of individual sarcoplasmic and myofibrillar proteins between cohorts. Whole-body fat-free mass (YT > YU = OU), VL thickness (YT > YU = OU), and leg extensor peak torque (YT > YU = OU) differed between groups ( < 0.05). Total myofibrillar protein concentrations were greater in YT versus OU ( = 0.005), but were not different between YT versus YU ( = 0.325). The abundances of actin and myosin heavy chain were greater in YT versus YU ( < 0.05) and OU ( < 0.001). Total sarcoplasmic protein concentrations were not different between groups. While proteomics indicated that marginal differences existed for individual myofibrillar and sarcoplasmic proteins between YT versus other groups, age-related differences were more prominent for myofibrillar proteins (YT = YU > OU, < 0.05: 7 proteins; OU > YT = YU, < 0.05: 11 proteins) and sarcoplasmic proteins (YT = YU > OU, < 0.05: 8 proteins; OU > YT&YU, < 0.05: 29 proteins). In summary, our data suggest that modest (~9%) myofibrillar protein packing (on a per mg muscle basis) was evident in the YT group. This study also provides further evidence to suggest that notable skeletal muscle proteome differences exist between younger and older humans. However, given that our n-sizes are low, these results only provide a preliminary phenotyping of the reported protein and proteomic variables.
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http://dx.doi.org/10.3390/sports8010007DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7022975PMC
January 2020

Stromal Modulation and Treatment of Metastatic Pancreatic Cancer with Local Intraperitoneal Triple miRNA/siRNA Nanotherapy.

ACS Nano 2020 01 13;14(1):255-271. Epub 2020 Jan 13.

Center for Drug Delivery and Nanomedicine, Department of Pharmaceutical Sciences , University of Nebraska Medical Center , Omaha , Nebraska 68198 , United States.

Nanomedicines achieve tumor-targeted delivery mainly through enhanced permeability and retention (EPR) effect following intravenous (IV) administration. Unfortunately, the EPR effect is severely compromised in pancreatic cancer due to hypovascularity and dense desmoplastic stroma. Intraperitoneal (IP) administration may be an effective EPR-independent local delivery approach to target peritoneal tumors. Besides improved delivery, effective combination delivery strategies are needed to improve pancreatic cancer therapy by targeting both cancer cells and cellular interactions within the tumor stroma. Here, we described simple cholesterol-modified polymeric CXCR4 antagonist (PCX) nanoparticles (to block cancer-stroma interactions) for codelivery of anti-miR-210 (to inactivate stroma-producing pancreatic stellate cells (PSCs)) and siKRAS (to kill pancreatic cancer cells). IP administration delivered the nanoparticles to an orthotopic syngeneic pancreatic tumors as a result of preferential localization to the tumors and metastases with disrupted mesothelium and effective tumor penetration. The local IP delivery resulted in nearly 15-fold higher tumor accumulation than delivery by IV injection. Through antagonism of CXCR4 and downregulation of miR-210/KRAS, the triple-action nanoparticles favorably modulated desmoplastic tumor microenvironment via inactivating PSCs and promoting the infiltration of cytotoxic T cells. The combined therapy displayed improved therapeutic effect when compared with individual therapies as documented by the delayed tumor growth, depletion of stroma, reduction of immunosuppression, inhibition of metastasis, and prolonged survival. Overall, we present data that a local IP delivery of a miRNA/siRNA combination holds the potential to improve pancreatic cancer therapy.
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http://dx.doi.org/10.1021/acsnano.9b03978DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7041410PMC
January 2020

IL-17-CXC Chemokine Receptor 2 Axis Facilitates Breast Cancer Progression by Up-Regulating Neutrophil Recruitment.

Am J Pathol 2020 01 22;190(1):222-233. Epub 2019 Oct 22.

Department of Pathology and Microbiology, University of Nebraska Medical Center, Omaha, Nebraska. Electronic address:

Recent evidence suggests that interactions among proinflammatory cytokines, chemokines, and cancer cell-recruited neutrophils result in enhanced metastasis and chemotherapy resistance. Nonetheless, the detailed mechanism remains unclear. Our aim was to discover the role of IL-17, CXC chemokine receptor 2 (CXCR2) ligands, and cancer-associated neutrophils in chemotherapy resistance and metastasis in breast cancer. Mice were injected with Cl66 murine mammary tumor cells, Cl66 cells resistant to doxorubicin (Cl66-Dox), or Cl66 cells resistant to paclitaxel (Cl66-Pac). Higher levels of IL-17 receptor, CXCR2 chemokines, and CXCR2 were observed in tumors generated from Cl66-Dox and Cl66-Pac cells in comparison with tumors generated from Cl66 cells. Tumors generated from Cl66-Dox and Cl66-Pac cells had higher infiltration of neutrophils and T helper 17 cells. In comparison with primary tumor sites, there were increased levels of CXCR2, CXCR2 ligands, and IL-17 receptor within the metastatic lesions. Moreover, IL-17 increased the expression of CXCR2 ligands and cell proliferation of Cl66 cells. The supernatant of Cl66-Dox and Cl66-Pac cells enhanced neutrophil chemotaxis. In addition, IL-17-induced neutrophil chemotaxis was dependent on CXCR2 signaling. Collectively, these data demonstrate that the IL-17-CXCR2 axis facilitates the recruitment of neutrophils to the tumor sites, thus allowing them to play a cancer-promoting role in cancer progression.
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http://dx.doi.org/10.1016/j.ajpath.2019.09.016DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6943375PMC
January 2020

Metabolomic Profiling to Identify Molecular Biomarkers of Cellular Response to Methotrexate In Vitro.

Clin Transl Sci 2020 01 25;13(1):137-146. Epub 2019 Oct 25.

Division of Rheumatology, Department of Pediatrics, Duke Children's Hospital, Durham, North Carolina, USA.

Variation in methotrexate (MTX) efficacy represents a significant barrier to early and effective disease control in the treatment of autoimmune arthritis. We hypothesize that the utilization of metabolomic techniques will allow for an improved understanding of the biochemical basis for the pharmacological activity of MTX, and can promote the identification and evaluation of novel molecular biomarkers of MTX response. In this work, erythroblastoid cells were exposed to MTX at the physiologic concentration of 1,000 nM and analyzed using three metabolomic platforms to give a broad spectrum of cellular metabolites. MTX pharmacological activity, defined as cellular growth inhibition, was associated with an altered cellular metabolomic profile based on the analysis of 724 identified metabolites. By discriminant analysis, MTX treatment was associated with increases in ketoisovaleric acid, fructose, galactose, and 2-deoxycytidine, and corresponding reductions in 2-deoxyuridine, phosphatidylinositol 32:0, orotic acid, and inosine monophosphate. Inclusion of data from analysis of folate metabolism in combination with chemometric and metabolic network analysis demonstrated that MTX treatment is associated with dysregulated folate metabolism and nucleotide biosynthesis, which is in line with its known mechanism of action. However, MTX treatment was also associated with alterations in a diversity of metabolites, including intermediates of amino acid, carbohydrate, and lipid metabolism. Collectively, these findings support a robust metabolic response following exposure to physiologic concentrations of MTX. They also identify various metabolic intermediates that are associated with the pharmacological activity of MTX, and are, therefore, potential molecular biomarker candidates in future preclinical and clinical studies of MTX efficacy in autoimmune arthritis.
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http://dx.doi.org/10.1111/cts.12694DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6951846PMC
January 2020