Role of Base Excision Repair in Innate Immune Cells and Its Relevance for Cancer Therapy.

Innate immunity is critical for immediate recognition and elimination of invading pathogens or defense against cancer cell growth. Dysregulation of innate immune systems is associated with the pathogenesis of different types of inflammatory diseases, including cancer. In addition, the maintenance of innate immune cells' genomic integrity is crucial for the survival of all organisms. Oxidative stress generated from innate immune cells may cause self-inflicted DNA base lesions as well as DNA damage on others neighboring cells, including cancer cells. Oxidative DNA base damage is predominantly repaired by base excision repair (BER). BER process different types of DNA base lesions that are presented in cancer and innate immune cells to maintain genomic integrity. However, mutations in BER genes lead to impaired DNA repair function and cause insufficient genomic integrity. Moreover, several studies have implicated that accumulation of DNA damage leads to chromosomal instability that likely activates the innate immune signaling. Furthermore, dysregulation of BER factors in cancer cells modulate the infiltration of innate immune cells to the tumor microenvironment. In the current review, the role of BER in cancer and innate immune cells and its impact on innate immune signaling within the tumor microenvironment is summarized. This is a special issue that focuses on DNA damage and cancer therapy to demonstrate how BER inhibitor or aberrant repair modulates innate inflammatory response and impact immunotherapy approaches. Overall, the review provides substantial evidence to understand the impact of BER in innate immune response dynamics within the current immune-based therapeutic strategy.

A novel role of snail in regulating tuberin/AMPK pathways to promote renal fibrosis in the new mouse model of type II diabetes.

Epithelial-mesenchymal transition (EMT) plays an important role in tissue fibrosis following chronic exposure to hyperglycemia. This study investigates the role of chronic diabetes in regulating tuberin/snail/AMPK to enhance EMT and increase renal fibrosis. A new mouse model of db/db/TSC2 +/- was generated by backcrossing db/db mice and TSC2 +/- mice. Wild type (WT), db/db, TSC2 +/- and dbdb/TSC2 +/- mice were sacrificed at ages 6 and 8 months old. Tuberin protein level was significantly decreased in kidneys from diabetic compared to WT mice at both ages. In addition, tuberin and E-cadherin protein levels were significantly decreased in dbdb/TSC2 +/- compared to TSC2 +/- and db/db mice. In contrast, p-PS6K, NFkB, snail, vimentin, fibronectin, and α-SMA protein levels were significantly increased in dbdb/TSC2 +/- compared to db/db and TSC2 +/- mice at ages 6 and 8 months. Both downregulation of AMPK by DN-AMPK and downregulation of tuberin by siRNA resulted in increased NFkB, snail, and fibronectin protein expression and decreased E-cadherin protein expression in mouse primary renal proximal tubular cells. Interestingly, downregulation of snail by siRNA increased tuberin expression via feedback through activation of AMPK and reversed the expression of epithelial proteins such as E-cadherin as well as mesenchymal proteins such as fibronectin, NF-KB, vimentin, and α-SMA in mouse primary renal proximal tubular cells isolated from kidneys of four mice genotypes. The data show that chronic diabetes significantly decreases tuberin expression and that provides strong evidence that tuberin is a major key protein involved in regulating EMT. These data also demonstrated a novel role for snail in regulating of AMPK/tuberin to enhance EMT and renal cell fibrosis in diabetes.

mTOR signaling regulates gastric epithelial progenitor homeostasis and gastric tumorigenesis via MEK1-ERKs and BMP-Smad1 pathways.

mTOR, the sensor of nutrients and growth factors, has important roles in tissue homeostasis and tumorigenesis. However, how mTOR controls gastric epithelial cell turnover and gastric cancer development, a leading malignancy, remains poorly understood. Here, we provide genetic evidence that mTOR activation promotes proliferation and inhibits differentiation of Lgr5+ gastric epithelial progenitors (GEPs) in gastric homeostasis and tumorigenesis. mTOR signaling increases MEK1 and Smad1 expression and enhances activation of MEK1-ERKs and BMP-Smad1 pathways, respectively, in GEPs and gastric tumors. Mek1 deletion or inhibition rescues hyperproliferation, whereas Bmpr1a ablation or inhibition rescues differentiation defects of Tsc1-/- GEPs. Tsc1 deficiency in Lgr5+ GEPs accelerates gastric tumor initiation and development, which require MEK1-ERKs for hyperplasia and BMP-Smad1 for differentiation suppression. These findings reveal how mTOR signaling controls Lgr5+ GEP homeostasis and cancerization and suggest that ERKs and Smad1 signaling can be safely targeted to substitute mTOR inhibitors in gastric cancer therapy.

TP63 Is Significantly Upregulated in Diabetic Kidney.

The role of tumor protein 63 (TP63) in regulating insulin receptor substrate 1 (IRS-1) and other downstream signal proteins in diabetes has not been characterized. RNAs extracted from kidneys of diabetic mice (db/db) were sequenced to identify genes that are involved in kidney complications. RNA sequence analysis showed more than 4- to 6-fold increases in TP63 expression in the diabetic mice's kidneys, compared to wild-type mice at age 10 and 12 months old. In addition, the kidneys from diabetic mice showed significant increases in TP63 mRNA and protein expression compared to WT mice. Mouse proximal tubular cells exposed to high glucose (HG) for 48 h showed significant decreases in IRS-1 expression and increases in TP63, compared to cells grown in normal glucose (NG). When TP63 was downregulated by siRNA, significant increases in IRS-1 and activation of AMP-activated protein kinase (AMPK (p-AMPK-Th172)) occurred under NG and HG conditions. Moreover, activation of AMPK by pretreating the cells with AICAR resulted in significant downregulation of TP63 and increased IRS-1 expression. Ad-cDNA-mediated over-expression of tuberin resulted in significantly decreased TP63 levels and upregulation of IRS-1 expression. Furthermore, TP63 knockdown resulted in increased glucose uptake, whereas IRS-1 knockdown resulted in a decrease in the glucose uptake. Altogether, animal and cell culture data showed a potential role of TP63 as a new candidate gene involved in regulating IRS-1 that may be used as a new therapeutic target to prevent kidney complications in diabetes.

mTOR Activation Initiates Renal Cell Carcinoma Development by Coordinating ERK and p38MAPK.

Renal cell carcinoma (RCC) mainly originates from renal proximal tubules. Intriguingly, disruption of genes frequently mutated in human RCC samples thus far has only generated RCC originated from other renal tubule parts in mouse models. This hampers our understanding of the pathogenesis of RCC. Here we show that mTOR signaling, often activated in RCC samples, initiates RCC development from renal proximal tubules. Ablation of Tsc1, encoding an mTOR suppressor, in proximal tubule cells led to multiple precancerous renal cysts. mTOR activation increased MEK1 expression and ERK activation, and Mek1 ablation or inhibition diminished cyst formation in Tsc1-deficient mice. mTOR activation also increased MKK6 expression and p38MAPK activation, and ablation of the p38α-encoding gene further enhanced cyst formation and led to RCC with clear cell RCC features. Mechanistically, Tsc1 deletion induced p53 and p16 expression in a p38MAPK-dependent manner, and deleting Tsc1 and Trp53 or Cdkn2a (encoding p16) enhanced renal cell carcinogenesis. Thus, mTOR activation in combination with inactivation of the p38MAPK-p53/p16 pathway drives RCC development from renal proximal tubules. Moreover, this study uncovers previously unidentified mechanisms by which mTOR controls cell proliferation and suggests the MEK-ERK axis to be a potential target for treatment of RCC. SIGNIFICANCE: Mouse modeling studies show that mTOR activation in combination with inactivation of the p38MAPK-p53/p16 axis initiates renal cell carcinoma that mimics human disease, identifying potential therapeutic targets for RCC treatment.

Oxidatively modified low-density lipoproteins are potential mediators of proteasome inhibitor resistance in multiple myeloma.

Proteasome inhibitor (PI) therapy has improved the survival of multiple myeloma (MM) patients. However, inevitably, primary or acquired resistance to PIs leads to disease progression; resistance mechanisms are unclear. Obesity is a risk factor for MM mortality. Oxidized LDL (OxLDL), a central mediator of atherosclerosis that is elevated in metabolic syndrome (co-occurrence of obesity, insulin resistance, dyslipidemia and hypertension), has been linked to an increased risk of solid cancers and shown to stimulate pro-oncogenic/survival signaling. We hypothesized that OxLDL is a mediator of chemoresistance and evaluated its effects on MM cell killing by PIs. OxLDL potently suppressed the ability of the boronic acid-based PIs bortezomib (BTZ) and ixazomib, but not the epoxyketone-based PI carfilzomib, to kill human MM cell lines and primary cells. OxLDL suppressed BTZ-induced inhibition of proteasome activity and induction of pro-apoptotic signaling. These cytoprotective effects were abrogated when lipid hydroperoxides (LOOHs) associated with OxLDL were enzymatically reduced. We also demonstrated the presence of OxLDL in the MM bone marrow microenvironment as well as numerous granulocytes and monocytes capable of cell-mediated LDL oxidation through myeloperoxidase. Our findings suggest that OxLDL may be a potent mediator of boronic acid-based PI resistance, particularly for MM patients with metabolic syndrome, given their elevated systemic levels of OxLDL. LDL cholesterol-lowering therapy to reduce circulating OxLDL, and pharmacologic targeting of LOOH levels or resistance pathways induced by the modified lipoprotein, could deepen the response to these important agents and offer clinical benefit to MM patients with metabolic syndrome.

CSF-1 in Osteocytes Inhibits Nox4-mediated Oxidative Stress and Promotes Normal Bone Homeostasis.

CSF-1 is a key factor in regulating bone remodeling; osteocytes express CSF-1 and its receptor. Viable osteocytes are essential for bone remodeling through cell-cell contact and secretion of factors that regulate osteoblasts and osteoclasts. Increased oxidative stress contributes to osteocyte death and correlates with bone loss during aging. The NADPH oxidase Nox4 is a major source of ROS in bone. CSF-1 decreases Nox4, suggesting that CSF-1 protects against oxidative stress. Here, we show that osteocyte apoptosis previously reported in our global CSF-1KO mice is associated with increased Nox4, as well as 4-HNE expression in osteocytes. Osteocytes isolated from CSF-1KO mice were less viable and showed increased intracellular ROS, elevated NADPH oxidase activity/Nox4 protein, activation of mTOR/S6K, and downstream apoptosis signals compared with WT osteocytes. Nox4 expression was also increased in CSF-1KO osteocytes and colocalized with MitoTracker Red in mitochondria. Notably, CSF-1 inhibited Nox4 expression and apoptosis cascade signals. In additional studies, shNox4 decreased these signals in CSF-1KO osteocytes, whereas overexpression of Nox4 in WT osteocytes activated the apoptosis pathway. To determine the role of CSF-1 in osteocytes, DMP1Cre-CSF-1cKO (CSF-1cKO) mice that lack CSF-1 in osteocytes/late osteoblasts were developed. Osteocyte defects in CSF-1cKO mice overlapped with those in CSF-1KO mice, including increased apoptosis, Nox4, and 4-HNE-expressing osteocytes. CSF-1cKO mice showed unbalanced cancellous bone remodeling with decreased bone formation and resorption. Continued exposure to high Nox4/ROS levels may further compromise bone formation and predispose to bone loss and skeletal fragility. Taken together, our findings suggest a novel link between CSF-1, Nox4-derived ROS, and osteocyte survival/function that is crucial for osteocyte-mediated bone remodeling. Results reveal new mechanisms by which CSF-1/oxidative stress regulate osteocyte homeostasis, which may lead to therapeutic strategies to improve skeletal health in aging. © 2018 American Society for Bone and Mineral Research.

Gut stem cell aging is driven by mTORC1 via a p38 MAPK-p53 pathway.

Nutrients are absorbed solely by the intestinal villi. Aging of this organ causes malabsorption and associated illnesses, yet its aging mechanisms remain unclear. Here, we show that aging-caused intestinal villus structural and functional decline is regulated by mTORC1, a sensor of nutrients and growth factors, which is highly activated in intestinal stem and progenitor cells in geriatric mice. These aging phenotypes are recapitulated in intestinal stem cell-specific Tsc1 knockout mice. Mechanistically, mTORC1 activation increases protein synthesis of MKK6 and augments activation of the p38 MAPK-p53 pathway, leading to decreases in the number and activity of intestinal stem cells as well as villus size and density. Targeting p38 MAPK or p53 prevents or rescues ISC and villus aging and nutrient absorption defects. These findings reveal that mTORC1 drives aging by augmenting a prominent stress response pathway in gut stem cells and identify p38 MAPK as an anti-aging target downstream of mTORC1.

Tsc1 ablation in Prx1 and Osterix lineages causes renal cystogenesis in mouse.

Tuberous Sclerosis Complex (TSC) is caused by mutations in TSC1 or TSC2, which encode negative regulators of the mTOR signaling pathway. The renal abnormalities associated with TSC include angiomyolipoma, cysts, and renal cell carcinoma. Here we report that specific ablation of Tsc1 using the mesenchymal stem cell-osteoblast lineage markers induced cystogenesis in mice. Using Rosa-tdTomato mice, we found that Prx1- or Dermo1-labeled cells were present in the nephron including glomerulus but they were not stained by markers for podocytes, mesangial cells, endothelial cells, or proximal or loop of Henle tubular cells, while Osx is known to label tubular cells. Tsc1 deficiency in Prx1 lineage cells caused development of mild cysts that were positive only for Tamm-Horsfall protein (THP), a loop of Henle marker, while Tsc1 deficiency in Osx lineage cells caused development of cysts that were positive for Villin, a proximal tubular cell marker. On the other hand, Tsc1 deficiency in the Dermo1 lineage did not produce detectable phenotypical changes in the kidney. Cyst formation in Prx1-Cre; Tsc1f/f and Osx-Cre; Tsc1f/f mice were associated with increase in both proliferative and apoptotic cells in the affected tissue and were largely suppressed by rapamycin. These results suggest that Prx1 and Osx lineages cells may contribute to renal cystogenesis in TSC patients.

Endothelial cell-specific overexpression of endothelial nitric oxide synthase in Ins2Akita mice exacerbates diabetic nephropathy.

Previous studies demonstrated that global deficiency of eNOS in diabetic mice exacerbated renal lesions and that overexpression of eNOS may protect against tissue injury. Our study revealed for the first time overexpression of eNOS leads to disease progression rather than protection. Transgenic mice selectively expressing eNOS in endothelial cells (eNOSTg) were cross bred with Ins2Akita type-1 (AK) diabetic mice to generate eNOS overexpressing eNOSTg/AK mice. Wild type, eNOSTg, AK and eNOSTg/AK mice were assessed for kidney function and blood glucose levels. Remarkably, overexpressing eNOSTg mice showed evidence of unpredicted glomerular injury with segmental mesangiolysis and occasional microaneurysms. Notably, in eNOSTg/AK mice overexpression of eNOS led to increased glomerular/endothelial injury that was associated with increased superoxide levels and renal dysfunction. Results indicate for the first time that overexpressing eNOS in endothelial cells cannot ameliorate diabetic lesions, but paradoxically leads to progression of nephropathy likely due to eNOS uncoupling and superoxide upsurge. This novel finding has a significant impact on current therapeutic strategies to improve endothelial function and prevent progression of diabetic renal disease. Further, the eNOSTg/AK model developed in this study has significant translational potentials for elucidating the underlying mechanism implicated in the deflected function of eNOS in diabetic nephropathy.

Galectin-1 is a new fibrosis protein in type 1 and type 2 diabetes.

Chronic exposure of tubular renal cells to high glucose contributes to tubulointerstitial changes in diabetic nephropathy. In the present study, we identified a new fibrosis gene called galectin-1 (Gal-1), which is highly expressed in tubular cells of kidneys of type 1 and type 2 diabetic mouse models. Gal-1 protein and mRNA expression showed significant increase in kidney cortex of heterozygous Akita+/- and db/db mice compared with wild-type mice. Mouse proximal tubular cells exposed to high glucose showed significant increase in phosphorylation of Akt and Gal-1. We cloned Gal-1 promoter and identified the transcription factor AP4 as binding to the Gal-1 promoter to up-regulate its function. Transfection of cells with plasmid carrying mutations in the binding sites of AP4 to Gal-1 promoter resulted in decreased protein function of Gal-1. In addition, inhibition of Gal-1 by OTX-008 showed significant decrease in p-Akt/AP4 and protein-promoter activity of Gal-1 and fibronectin. Moreover, down-regulation of AP4 by small interfering RNA resulted in a significant decrease in protein expression and promoter activity of Gal-1. We found that kidney of Gal-1-/- mice express very low levels of fibronectin protein. In summary, Gal-1 is highly expressed in kidneys of type 1 and 2 diabetic mice, and AP4 is a major transcription factor that activates Gal-1 under hyperglycemia. Inhibition of Gal-1 by OTX-008 blocks activation of Akt and prevents accumulation of Gal-1, suggesting a novel role of Gal-1 inhibitor as a possible therapeutic target to treat renal fibrosis in diabetes.-Al-Obaidi, N., Mohan, S., Liang, S., Zhao, Z., Nayak, B. K., Li, B., Sriramarao, P., Habib, S. L. Galectin-1 is a new fibrosis protein in type 1 and type 2 diabetes.

Heterozygous Tsc2 (Tsc2+/-) mouse model to study induced renal cancer in response to ionizing radiation at low doses.

Kidneys are one of the main dose-limiting organs in radiotherapeutic procedures of lower abdomen. Likewise, the threat of exposure of radiosensitive organs such as kidneys in warfare or radiation accidents among military personal or due to terrorist activities in general public is of increasing concern. These events warrant the need for appropriate animal models to study the acute and chronic effects of low- and high-dose rate radiation exposures. In this study, for the first time, we validated Tsc2+/- mouse model to study whether radiation accelerates carcinogenesis in kidneys. Tsc2+/- mice at increasing age groups at 8 and 10 months were exposed to repeated doses of gamma radiation (0.4 Gy × 5) and assessed for aggravated kidney tumor formation at 2 months post-irradiation. Animals from irradiated group showed a significant increase in numbers of bilateral, multifocal tumors compared with mock-irradiated animals. Intra-glomerular reactive oxygen species (ROS) levels measured by dihydroethidium florescence showed significant increases in ROS production in irradiated Tsc2+/- mice compared with non-irradiated animals. Similarly, selective hematological parameters and glomerular filtration rate were further reduced significantly in irradiated Tsc2+/- mice. Tsc2 protein, tuberin in irradiated mice, however, remains at the same reduced levels as that of the mock-irradiated heterozygous Tsc2 mice. The results indicate that radiation alters kidney homeostatic function and influences high spontaneous incidence of renal cell carcinoma in this rodent model. Repurposing of Tsc2+/- mice model will, therefore, provide a unique opportunity to study acute and delayed effects of radiation in the development of kidney cancers.

A new drug combination significantly reduces kidney tumor progression in kidney mouse model.

Tuberous sclerosis complex (TSC) disease is associated with tumors in many organs, particularly angiomyolipoma (AML) in the kidneys. Loss or inactivation of TSC1/2 results in high levels of HIF-α activity and VEGF expression. mTOR inhibitor (rapamycin) and the AMPK activator 5-aminoimidazole-4-carboxamide (AICA)-riboside (AICAR) are currently used separately to treat cancer patients. Here, we investigated the effect of a novel combination of rapamycin and AICAR on tumor progression. Our data show that treatment of AML human cells with drug combinations resulted in 5-7-fold increase in cell apoptosis compared to each drug alone. In addition, drug combinations resulted in 4-5-fold decrease in cell proliferation compared to each drug alone. We found that drug combinations abolished Akt and HIF activity in AML cells. The drug combinations resulted in decrease in cell invasion and cell immigration by 70% and 84%, respectively in AML cells. The combined drugs also significantly decreased the VEGF expression compare to each drug alone in AML cells. Drug combinations effectively abolished binding of HIF-2α to the putative Akt site in the nuclear extracts isolated from AML cells. Treatment TSC mice with drug combinations resulted in 75% decrease in tumor number and 88% decrease in tumor volume compared to control TSC mice. This is first evidence that drug combinations are effective in reducing size and number of kidney tumors without any toxic effect on kidney. These data will provide evidence for initiating a new clinical trial for treatment of TSC patients.

Preclinical evidence of the enhanced effectiveness of combined rapamycin and AICAR in reducing kidney cancer.

Loss of Von Hippel-Lindau in renal carcinoma cells results in upregulation of the activity of hypoxia-inducible factor (HIF-α), a major transcription factor involved in kidney cancer. Rapamycin as mammalian target of rapamycin inhibitor and 5-aminoimidazole-4-carboxamide-riboside (AICAR) as AMPK activator are used separately to treat cancer patients. In the current study, the possible additive effect of drug combinations in reducing kidney tumorigenesis was investigated. Treatment with drug combinations significantly decreased cell proliferation, increased cell apoptosis, and abolished Akt phosphorylation and HIF-2α expression in renal cell carcinoma cells, including primary cells isolated from kidney cancer patients. Significant decreases in cell migration and invasion were detected using drug combinations. Drug combinations effectively abolished binding of HIF-2α to the Akt promoter and effected formation of the DNA-protein complex in nuclear extracts from 786-O cells, as demonstrated using electromobility shift assay and examination of Akt promoter activity. Importantly, we tested the effect of each drug and the combined drugs on kidney tumor size in the nude mouse model. Our data show that treatment with rapamycin, AICAR, and rapamycin+AICAR decreased tumor size by 38%, 36%, and 80%, respectively, suggesting that drug combinations have an additive effect in reducing tumor size compared with use of each drug alone. Drug combinations effectively decreased cell proliferation, increased apoptotic cells, and significantly decreased p-Akt, HIF-2α, and vascular endothelial growth factor expression in tumor kidney tissues from mice. These results show for the first time that drug combinations are more effective than single drugs in reducing kidney tumor progression. This study provides important evidence that may lead to the initiation of pre-clinical trials in patients with kidney cancer.

Kidney atrophy vs hypertrophy in diabetes: which cells are involved?

One of the first structural changes in diabetic nephropathy (DN) is the renal enlargement. These changes resulted in renal hypertrophy in both glomerular and tubular cells. Shrink in the kidney size, which described as kidney atrophy resulted from the loss of nephrons or abnormal nephron function and lead to loss of the kidney function. On the other hand, increase in kidney size, which described as hypertrophy resulted from increase in proximal tubular epithelial and glomerular cells size. However overtime, tubular atrophy and tubulointerstitial fibrosis occurs as subsequent changes in tubular cell hypertrophy, which is associated with the infiltration of fibroblast cells into the tubulointerstitial space. The rate of deterioration of kidney function shows a strong correlation with the degree of tubulointerstitial fibrosis. A consequence of long-standing diabetes/hyperglycemia may lead to major changes in renal structure that occur but not specific only to nephropathy. Identifying type of cells that involves in renal atrophy and hypertrophy may help to find a therapeutic target to treat diabetic nephropathy. In summary, the early changes in diabetic kidney are mainly includes the increase in tubular basement membrane thickening which lead to renal hypertrophy. On the other hand, only renal tubule is subjected to apoptosis, which is one of the characteristic morphologic changes in diabetic kidney to form tubular atrophy at the late stage of diabetes.

Nanoparticle as a novel tool in hyperthermic intraperitoneal and pressurized intraperitoneal aerosol chemotheprapy to treat patients with peritoneal carcinomatosis.

The treatment of peritoneal surface malignances has changed considerably over the last thirty years. Unfortunately, the palliative is the only current treatment for peritoneal carcinomatosis (PC). Two primary intraperitoneal chemotherapeutic methods are used. The first is combination of cytoreductive surgery (CRS) and Hyperthermic IntraPEritoneal Chemotherapy (HIPEC), which has become the gold standard for many cases of PC. The second is Pressurized IntraPeritoneal Aerosol Chemotheprapy (PIPAC), which is promising direction to minimally invasive as safedrug delivery. These methods were improved through multicenter studies and clinical trials that yield important insights and solutions. Major method development has been made through nanomedicine, specifically nanoparticles. Here, we are presenting the latest advances of nanoparticles and their application to precision diagnostics and improved treatment strategies for PC. These advances will likely develop both HIPEC and PIPAC methods that used for in vitro and in vivo studies. Several benefits of using nanoparticles will be discussed including: 1) Nanoparticles as drug delivery systems; 2) Nanoparticles and Near Infrred (NIR) Irradiation; 3) use of nanoparticles in perioperative diagnostic and individualized treatment planning; 4) use of nanoparticles as anticancer dressing's, hydrogels and as active beeds for optimal reccurence prevention; and 5) finally the curent in vitro and in vivo studies and clinical trials of nanoparticles. The current review highlighted use of nanoparticles as novel tools in improving drug delivery to be effective for treatment patients with peritoneal carcinomatosis.

Glutamine Addiction in Kidney Cancer Suppresses Oxidative Stress and Can Be Exploited for Real-Time Imaging.

Many cancers appear to activate intrinsic antioxidant systems as a means to counteract oxidative stress. Some cancers, such as clear cell renal cell carcinoma (ccRCC), require exogenous glutamine for growth and exhibit reprogrammed glutamine metabolism, at least in part due to the glutathione pathway, an efficient cellular buffering system that counteracts reactive oxygen species and other oxidants. We show here that ccRCC xenograft tumors under the renal capsule exhibit enhanced oxidative stress compared with adjacent normal tissue and the contralateral kidney. Upon glutaminase inhibition with CB-839 or BPTES, the RCC cell lines SN12PM-6-1 (SN12) and 786-O exhibited decreased survival and pronounced apoptosis associated with a decreased GSH/GSSG ratio, augmented nuclear factor erythroid-related factor 2, and increased 8-oxo-7,8-dihydro-2'-deoxyguanosine, a marker of DNA damage. SN12 tumor xenografts showed decreased growth when treated with CB-839. Furthermore, PET imaging confirmed that ccRCC tumors exhibited increased tumoral uptake of 18F-(2S,4R)4-fluoroglutamine compared with the kidney in the orthotopic mouse model. This technique can be utilized to follow changes in ccRCC metabolism in vivo Further development of these paradigms will lead to new treatment options with glutaminase inhibitors and the utility of PET to identify and manage patients with ccRCC who are likely to respond to glutaminase inhibitors in the clinic. Cancer Res; 77(23); 6746-58. ©2017 AACR.

Bone Size and Quality Regulation: Concerted Actions of mTOR in Mesenchymal Stromal Cells and Osteoclasts.

The bone size and quality, acquired during adolescent growth under the influence of anabolic hormones, growth factors, and nutrients, determine the height and bone stability and forecast osteoporosis risks in late life. Yet bone size and quality control mechanisms remain enigmatic. To study the roles of mammalian target of rapamycin (mTOR) signaling, sensor of growth factors and nutrients, in bone size and quality regulation, we ablated Tsc1, a suppressor of mTOR, in mesenchymal stromal cells (MSCs), monocytes, or their progenies osteoblasts and osteoclasts. mTOR activation in MSCs, but much less in osteoblasts, increased bone width and mass due to MSC hyperproliferation, but decreased bone length and mineral contents due to defective MSC differentiation. mTOR activation promotes bone mineral accretion by inhibiting osteoclast differentiation and activity directly or via coupling with MSCs. Tuberous sclerosis complex patient studies confirmed these findings. Thus, mTOR regulates bone size via MSCs and bone quality by suppressing catabolic activities of osteoclasts.