Expression of cell surface zinc transporter LIV1 in triple negative breast cancer is an indicator of poor prognosis and therapy failure.

Triple negative breast cancers (TNBC) are an aggressive molecular subtype of breast carcinoma (BC) identified by the lack of receptor expression for estrogen, progesterone, & human epidermal growth factor receptor-2. Lack of tangible drug targets warrants further research in TNBC. LIV1, is a zinc (Zn) transporter known to be overexpressed in few cancer types including BCs. Recently, in the United States of America, FDA approved the use of a new drug targeting LIV1, antibody drug conjugate SGN-LIV1A for treatment of TNBC patients. Though LIV1 also has a role in modulating immune cells by its differential transport of Zn, a correlation between the tumor cell expression of LIV1 and immune cell infiltrations were scantily reported. Further adequate baseline data on LIV1 expression in other populations have not been documented. Our objective was to screen a large Indian cohort of TNBC patient samples for LIV1, categorize the immune cell infiltration using CD4/CD8 expression and correlate the findings with therapy outcomes. Further, we also investigated for LIV1 expression in matched samples of primary & secondary tumors; pre & postchemotherapy in TNBC patients. Results showed an elevated expression of LIV1 in TNBC samples as compared to adjacent normal, the mean Q scores being 183.06 ± 6.39 and 120.78 ± 7.37 (p < 0.0001), respectively. Similarly, LIV1 levels were elevated in secondary tumors than primary & in patient samples postchemotherapy as compared to naïve. In the TNBC cohort, using automated method, cell morphology parameters were computed and analysis showed LIV1 levels were elevated in grade 3 TNBC samples presenting with altered cell morphology parameters namely cell size, cell perimeter, & nucleus size. Thus indicating LIV1 expressing TNBC samples portrayed an aggressive phenotype. Finally, TNBC patients with 3+ staining intensity showed poor survival (4.44 year) as compared to patients with 2+ LIV1 expression (5.47 year), emphasizing that LIV1 expression is a poor prognostic factor in TNBC. In conclusion, the study reports elevated expression of LIV1 in a large Indian TNBC cohort; high expression is a poor prognostic factor and correlated with aggressive disease and indicating the need for LIV1 targeted therapies.

Inflammation-induced PELP1 expression promotes tumorigenesis by activating GM-CSF paracrine secretion in the tumor microenvironment.

The inflammatory tumor microenvironment has been implicated as a major player fueling tumor progression and an enabling characteristic of cancer, proline, glutamic acid, and leucine-rich protein 1 (PELP1) is a novel nuclear receptor coregulator that signals across diverse signaling networks, and its expression is altered in several cancers. However, investigations to find the role of PELP1 in inflammation-driven oncogenesis are limited. Molecular studies here, utilizing macrophage cell lines and animal models upon stimulation with lipopolysaccharide (LPS) or necrotic cells, showed that PELP1 is an inflammation-inducible gene. Studies on the PELP1 promoter and its mutant identified potential binding of c-Rel, an NF-κB transcription factor subunit, to PELP1 promoter upon LPS stimulation in macrophages. Recruitment of c-Rel onto the PELP1 promoter was validated by chromatin immunoprecipitation, further confirming LPS mediated PELP1 expression through c-Rel-specific transcriptional regulation. Macrophages that overexpress PELP1 induces granulocyte-macrophage colony-stimulating factor secretion, which mediates cancer progression in a paracrine manner. Results from preclinical studies with normal-inflammatory-tumor progression models demonstrated a progressive increase in the PELP1 expression, supporting this link between inflammation and cancer. In addition, animal studies demonstrated the connection of PELP1 in inflammation-directed cancer progression. Taken together, our findings provide the first report on c-Rel-specific transcriptional regulation of PELP1 in inflammation and possible granulocyte-macrophage colony-stimulating factor-mediated transformation potential of activated macrophages on epithelial cells in the inflammatory tumor microenvironment, reiterating the link between PELP1 and inflammation-induced oncogenesis. Understanding the regulatory mechanisms of PELP1 may help in designing better therapeutics to cure various inflammation-associated malignancies.

Analysis of Morbidity, Mortality and Survival Pattern Following Surgery for Borderline Ovarian and Malignant Ovarian Tumour in Tertiary Care Centre.

BACKGROUNDS: To analyse the morbidity, mortality and survival pattern following surgery for borderline ovarian and malignant ovarian tumours. METHODS: The medical records of 57 consecutive patients with invasive and borderline epithelial ovarian cancer patients registered and operated in our tertiary centre between 2015 to 2017 were reviewed. Patients were followed up for a minimum of 18 months to maximum of 42 months at an interval of 3 months with CA125 values. Various prognostic factors were analysed. The data descriptive statistics of frequency and percentage analysis were used for categorical variables and mean and standard deviation were used for continuous variables. RESULTS: The most common age group was 51 years and above with the majority (56.2%) of women belonging to postmenopausal age group (32/57). In our study, 30 out of 57 women (52.6%) had stage III disease, 17 women had stage I disease (29.8%) and 7 women had stage 2 disease (12.3%). Majority of the women had serous epithelial ovarian tumour (47 out of 57 patients), which contributed to 82.4%. Grade 1 and 2 morbidity was encountered in 8 patients. Six patients had wound infection (grade 1), and 2 patients required blood transfusions (grade 2). One patient had grade 3 morbidity requiring re-laparotomy. Borderline tumours and early-stage epithelial ovarian tumours had good prognosis, less morbidity and good survival. The overall median survival was 25 months. CONCLUSIONS: With meticulous perioperative care, surgery for ovarian cancer in the primary and interval setting can be done with minimal morbidity and no postoperative mortality, especially in patients with co-morbidities. Grade is an important prognostic factor affecting the survival of patients with epithelial ovarian cancers undergoing surgery. Lymph node dissection helps achieve local control but may not improve the survival.

Facile synthesis and nanoscale features of a nanostructured nordihydroguaiaretic acid analog for therapeutic applications.

BACKGROUND: Nordihydroguaiaretic acid (NDGA) is a plant lignan obtained from creosote bush, known to possess anti-oxidant, anti-cancer and anti-viral activities and is being used in traditional medicine. However, toxicity studies indicated liver and kidney damage despite its immense medicinal properties. There has been a recent increase of curiosity in the chemical synthesis of NDGA derivatives for therapeutic applications. NDGA derivatives have been developed as better alternatives to NDGA and for targeted delivery to the site of tissue by chemical derivatives. In this regard, an analog of NDGA, Acetyl NDGA (Ac-NDGA), has been synthesized based on a previous procedure and formulated as a nanostructured complex with Polycaprolactone/Polyethylene glycol polymer matrices, by o/w solvent evaporation method. RESULTS: The drug-incorporated polymeric nanospheres exhibited a drug load of 10.0 ± 0.5 µg drug per mg of nanospheres in acetonitrile solvent with 49.95 ± 10% encapsulation efficiency and 33-41% drug loading capacity with different batches of nanospheres preparation. The in vitro drug release characteristics indicated 82 ± 0.25% drug release at 6 h in methanol. Further, the nanospheres have been characterized extensively to evaluate their suitability for therapeutic delivery. CONCLUSIONS: The present studies indicate a new and efficient formulation of the nanostructured AcNDGA with good therapeutic potential.

KIBRA attains oncogenic activity by repressing RASSF1A.

BACKGROUND: KIBRA-initially identified as a neuronal associated protein is now shown to be functionally associated with other tissue types as well. KIBRA interacts with dyenin light chain 1 and this interaction is essential for oestrogen receptor transactivation in breast cancer cells. KIBRA as a substrate of Cdk1, Aurora kinase and ERK plays an important role in regulating cell cycle, cell proliferation and migration. Despite these evidences, the exact role of KIBRA in cancer progression is not known. METHODS: We studied the expression of KIBRA in breast tissues and breast cancer cell lines by western blotting, immunohistochemisry (IHC) and RT-PCR. Stable over expression and knockdown clones were generated to study the transforming properties of KIBRA by conventional assays. Xenograft studies were performed in nude mice to study the in vivo tumourigenic efficacy of KIBRA. qPCR array was performed to understand the molecular mechanism behind oncogenic activity of KIBRA. RESULTS: Our results showed that KIBRA is upregulated in breast cancer cells and in malignant human breast tumours by both western blotting and IHC. Interestingly, we found that KIBRA expression level goes up with increase in breast cancer progression in well-established MCF10A model system. Further, results from stable overexpression clones of KIBRA in fibroblasts (Rat-1) and epithelial breast cancer cells (ZR75) and lentiviral short hairpin RNA-mediated knockdown (KD) clones of KIBRA in ZR75 showed increase in transforming properties with KIBRA overexpression and vice-versa. Results also showed that fibroblasts stably overexpressing KIBRA showed increased tumourigenic potential in nude mice. By adopting a quantitative PCR array-based approach, we identified RASSF1A, a tumour suppressor, as a transcriptional target of KIBRA. CONCLUSIONS: This is the first study to demonstrate the in vivo tumourigenic property of KIBRA in a nude mouse model and also unravel the underlying molecular mechanism of KIBRA-mediated transformation via repression of RASSF1A.British Journal of Cancer advance online publication, 29 June 2017; doi:10.1038/bjc.2017.192 www.bjcancer.com.

Snail-Modulated MicroRNA 493 Forms a Negative Feedback Loop with the Insulin-Like Growth Factor 1 Receptor Pathway and Blocks Tumorigenesis.

In this study, we have identified one microRNA, microRNA 493 (miR-493), which could simultaneously and directly regulate multiple genes downstream of the insulin-like growth factor 1 receptor (IGF1R) pathway, including IGF1R, by binding with complementary sequences in the 3' untranslated region (UTR) of mRNAs of IGF1R, insulin receptor substrate 1 (IRS1), and mitogen-activated protein kinase 1 (MAPK1), thereby potentiating their inhibitory function at multiple levels in development and progression of cancers. This binding was further confirmed by pulldown of miR with AGO-2 antibody. Further, results from head and neck samples showed that miR-493 levels were significantly downregulated in tumors, with a concomitant increase in the expression of IGF1R and key downstream effectors. Functional studies from miR-493 overexpression cells and nude-mouse models revealed the tumor suppressor functions of miR-493. Regulation studies revealed that Snail binds to the miR-493 promoter and represses it. We found the existence of a dynamic negative feedback loop in the regulation of IGF1R and miR-493 mediated via Snail. Our study showed that nicotine treatment significantly decreases the levels of miR-493-with a concomitant increase in the levels of Snail-an indication of progression of cells toward tumorigenesis, reestablishing the role of tobacco as a major risk factor for head and neck cancers and elucidating the mechanism behind nicotine-mediated tumorigenesis.

Gut Microbiota in Type 2 Diabetes Individuals and Correlation with Monocyte Chemoattractant Protein1 and Interferon Gamma from Patients Attending a Tertiary Care Centre in Chennai, India.

BACKGROUND: Type 2 diabetes mellitus (T2DM) and obesity are associated with changes in gut microbiota and characterized by chronic low-grade inflammation. Monocyte chemoattractant protein-1 (MCP-1) and interferon gamma (IFNγ) are proinflammatory cytokines which play an important role in the development of T2DM. We undertook this study to analyze the gut microbiota of T2DM and nondiabetic subjects and to determine the profile of MCP 1 and IFNγ in the same subjects attending a tertiary care center in Chennai, Tamil Nadu, India. METHODS: The study included 30 subjects with clinical details. Stool and blood samples were collected from all the subjects. DNA was extracted from fecal samples and polymerase chain reaction was done using fusion primers. Metagenomic analysis was performed using ion torrent sequencing. The reads obtained were in FASTA format and reported as operational taxonomic units. Human MCP 1 and IFNγ enzyme linked immunosorbent assay (ELISA) were performed for 23 serum samples. RESULTS: The study consisted of 30 subjects; 17 were T2DM and 13 were nondiabetics. The gut microbiota among T2DM consisted predominantly of Gram negative bacteria; Escherichia and Prevotella, when compared with the nondiabetic group with predominantly Gram positive organisms suchas Faecalibacterium, Eubacterium, and Bifidobacterium. The mean MCP-1 values in the diabetic group were 232.8 pg/ml and in the nondiabetic group 170.84 pg/ml. IFNγ (mean 385.5 pg/ml) was raised in glycated hemoglobin (HbA1c) group of 6.5-7.5% which was statistically significant. Association of Escherichia with T2DM and association of Bifidobacteria in the nondiabetics were also statistically significant. CONCLUSION: Escherichia counts were elevated in T2DM with HbA1c of 6.5-8.5% which was statistically significant suggesting that lipopolysaccharides present in the cell wall of Gram-negative bacteria may be responsible for low-grade inflammation as evidenced by elevated MCP-1 and IFNγ levels in T2DM with the same HbA1c levels.

Molecular Mechanism of Regulation of MTA1 Expression by Granulocyte Colony-stimulating Factor.

Parkinson disease (PD) is a neurodegenerative disorder with loss of dopaminergic neurons of the brain, which results in insufficient synthesis and action of dopamine. Metastasis-associated protein 1 (MTA1) is an upstream modulator of tyrosine hydroxylase (TH), the rate-limiting enzyme in dopamine synthesis, and hence MTA1 plays a significant role in PD pathogenesis. To impart functional and clinical significance to MTA1, we analyzed MTA1 and TH levels in the substantia nigra region of a large cohort of human brain tissue samples by Western blotting, quantitative PCR, and immunohistochemistry. Our results showed that MTA1 and TH levels were significantly down-regulated in PD samples as compared with normal brain tissue. Correspondingly, immunohistochemistry analysis for MTA1 in substantia nigra sections revealed that 74.1% of the samples had a staining intensity of <6 in the PD samples as compared with controls, 25.9%, with an odds ratio of 8.54. Because of the clinical importance of MTA1 established in PD, we looked at agents to modulate MTA1 expression in neuronal cells, and granulocyte colony-stimulating factor (G-CSF) was chosen, due to its clinically proven neurogenic effects. Treatment of the human neuronal cell line KELLY and acute 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine mouse model with G-CSF showed significant induction of MTA1 and TH with rescue of phenotype in the mouse model. Interestingly, the observed induction of TH was compromised on silencing of MTA1. The underlying molecular mechanism of MTA1 induction by G-CSF was proved to be through induction of c-Fos and its recruitment to the MTA1 promoter.