Cell surface CD55 traffics to the nucleus leading to cisplatin resistance and stemness by inducing PRC2 and H3K27 trimethylation on chromatin in ovarian cancer.

BACKGROUND: Platinum resistance is the primary cause of poor survival in ovarian cancer (OC) patients. Targeted therapies and biomarkers of chemoresistance are critical for the treatment of OC patients. Our previous studies identified cell surface CD55, a member of the complement regulatory proteins, drives chemoresistance and maintenance of cancer stem cells (CSCs). CSCs are implicated in tumor recurrence and metastasis in multiple cancers. METHODS: Protein localization assays including immunofluorescence and subcellular fractionation were used to identify CD55 at the cell surface and nucleus of cancer cells. Protein half-life determinations were used to compare cell surface and nuclear CD55 stability. CD55 deletion mutants were generated and introduced into cancer cells to identify the nuclear trafficking code, cisplatin sensitivity, and stem cell frequency that were assayed using in vitro and in vivo models. Detection of CD55 binding proteins was analyzed by immunoprecipitation followed by mass spectrometry. Target pathways activated by CD55 were identified by RNA sequencing. RESULTS: CD55 localizes to the nucleus of a subset of OC specimens, ascites from chemoresistant patients, and enriched in chemoresistant OC cells. We determined that nuclear CD55 is glycosylated and derived from the cell surface pool of CD55. Nuclear localization is driven by a trafficking code containing the serine/threonine (S/T) domain of CD55. Nuclear CD55 is necessary for cisplatin resistance, stemness, and cell proliferation in OC cells. CD55 S/T domain is necessary for nuclear entry and inducing chemoresistance to cisplatin in both in vitro and in vivo models. Deletion of the CD55 S/T domain is sufficient to sensitize chemoresistant OC cells to cisplatin. In the nucleus, CD55 binds and attenuates the epigenetic regulator and tumor suppressor ZMYND8 with a parallel increase in H3K27 trimethylation and members of the Polycomb Repressive Complex 2. CONCLUSIONS: For the first time, we show CD55 localizes to the nucleus in OC and promotes CSC and chemoresistance. Our studies identify a therapeutic mechanism for treating platinum resistant ovarian cancer by blocking CD55 nuclear entry.

Differential expression of IL-6/IL-6R and MAO-A regulates invasion/angiogenesis in breast cancer.

BACKGROUND: Monoamine oxidases (MAO) are mitochondrial enzymes functioning in oxidative metabolism of monoamines. The action of MAO-A has been typically described in neuro-pharmacological domains. Here, we have established a co-relation between IL-6/IL-6R and MAO-A and their regulation in hypoxia induced invasion/angiogenesis. METHODS: We employed various in-vitro and in-vivo techniques and clinical samples. RESULTS: We studied a co-relation among MAO-A and IL-6/IL-6R and tumour angiogenesis/invasion in hypoxic environment in breast cancer model. Activation of IL-6/IL-6R and its downstream was found in hypoxic cancer cells. This elevation of IL-6/IL-6R caused sustained inhibition of MAO-A in hypoxic environment. Inhibition of IL-6R signalling or IL-6R siRNA increased MAO-A activity and inhibited tumour angiogenesis and invasion significantly in different models. Further, elevation of MAO-A with 5-azacytidine (5-Aza) modulated IL-6 mediated angiogenesis and invasive signatures including VEGF, MMPs and EMT in hypoxic breast cancer. High grade invasive ductal carcinoma (IDC) clinical specimen displayed elevated level of IL-6R and depleted MAO-A expression. Expression of VEGF and HIF-1α was unregulated and loss of E-Cadherin was observed in high grade IDC tissue specimen. CONCLUSIONS: Suppression of MAO-A by IL-6/IL-6R activation promotes tumour angiogenesis and invasion in hypoxic breast cancer environment.

Identification of RAB2A and PRDX1 as the potential biomarkers for oral squamous cell carcinoma using mass spectrometry-based comparative proteomic approach.

Despite the recent advances in diagnostic and therapeutic strategies, oral squamous cell carcinoma (OSCC) remains a major health burden. Protein biomarker discovery for early detection will help to improve patient survival rate in OSCC. Mass spectrometry-based proteomics has emerged as an excellent approach for detection of protein biomarkers in various types of cancers. In the current study, we have used 4-Plex isobaric tags for relative and absolute quantitation (iTRAQ)-based shotgun quantitative proteomic approach to identify proteins that are differentially expressed in cancerous tissues compared to normal tissues. The high-resolution mass spectrometric analysis resulted in identifying 2,074 proteins, among which 288 proteins were differentially expressed. Further, it was noticed that 162 proteins were upregulated, while 125 proteins were downregulated in OSCC-derived cancer tissue samples as compared to the adjacent normal tissues. We identified some of the known molecules which were reported earlier in OSCC such as MMP-9 (8.4-fold), ZNF142 (5.6-fold), and S100A7 (3.5-fold). Apart from this, we have also identified some novel signature proteins which have not been reported earlier in OSCC including ras-related protein Rab-2A isoform, RAB2A (4.6-fold), and peroxiredoxin-1, PRDX1 (2.2-fold). The immunohistochemistry-based validation using tissue microarray slides in OSCC revealed overexpression of the RAB2A and PRDX1 gene in 80 and 68 % of the tested clinical cases, respectively. This study will not only serve as a resource of candidate biomarkers but will contribute towards the existing knowledge on the role of the candidate molecules towards disease progression and therapeutic potential.

Erratum to: Mechanistic attributes of S100A7 (psoriasin) in resistance of anoikis resulting tumor progression in squamous cell carcinoma of the oral cavity.

[This corrects the article DOI: 10.1186/s12935-015-0226-9.].

Mechanistic attributes of S100A7 (psoriasin) in resistance of anoikis resulting tumor progression in squamous cell carcinoma of the oral cavity.

BACKGROUND: Squamous cell carcinoma of the oral cavity (SCCOC) is the dominant origin of cancer associated mortality. Previous findings by our study reported that acquisition of anoikis resistance has a significant role in tumor progression of oral cavity. Several genes were over-expressed in anoikis-resistant cells under detached conditions which we confirmed earlier by microarray. Normal oral squamous epithelia grow adherent to a basement membrane, and when detached from the extracellular matrix, undergoes programmed cell death. The acquisition of anoikis-resistance is crucial phenomena in oral tumor advancement. In the current study, we have identified S100A7 expression as contributing factor for anoikis resistance and tumorigenicity in human oral cancer cells. Further, we have explored that elevated S100A7 expression in anoikis-sensitive oral keratinocytes and cancer cells reshape them more resistant to anoikis and apoptosis inducers via activation of cellular intrinsic and extrinsic avenue. METHODS: A subset of human cancer cell lines TU167, JMAR, JMARC39, JMARC42 and MDA-MB-468 were utilized for the generation of resistant stable cell lines. Further, immunohistochemistry, western blot and immunoprecipitation, assays of apoptosis, soft agar assay, orthotopic animal model and signaling elucidation were performed to establish our hypothesis. RESULTS: S100A7 gene is found to be responsible for anoikis resistance and tumorigenicity in human oral cancer cells. We have observed up-regulation of S100A7 in anoikis resistant cell lines, orthotropic model and patients samples with head and neck cancer. It is also noticed that secretion of S100A7 protein in conditioned medium by anoikis resistant head & neck cancer cell and in saliva of head and neck cancer patients. Up-regulation of S100A7 expression has triggered enhanced tumorigenicity and anchorage-independent growth of cancer cells through Akt phosphorylation leading to development of aniokis resistance in head and neck cancer cells. CONCLUSIONS: These data have led us to conclude that S100A7 is the major contributing factor in mediating anoikis-resistance of oral cancer cells and local tumor progression, and S100A7 might be useful as diagnostic marker for early detection of primary and recurrent squamous cell cancer.

Overcoming Akt Induced Therapeutic Resistance in Breast Cancer through siRNA and Thymoquinone Encapsulated Multilamellar Gold Niosomes.

Akt overexpression in cancer causes resistance to traditional chemotherapeutics. Silencing Akt through siRNA provides new therapeutic options; however, poor in vivo siRNA pharmacokinetics impede translation. We demonstrate that acidic milieu-sensitive multilamellar gold niosomes (Nio-Au) permit targeted delivery of both Akt-siRNA and thymoquinone (TQ) in tamoxifen-resistant and Akt-overexpressing MCF7 breast cancer cells. Octadecylamine groups of functionalized gold nanoparticles impart cationic attribute to niosomes, stabilized through polyethylene glycol. TQ's aqueous insolubility renders its encapsulation within hydrophobic core, and negatively charged siRNA binds in hydrophilic region of cationic niosomes. These niosomes were exploited to effectively knockdown Akt, thereby sensitizing cells to TQ. Immunoblot studies revealed enhanced apoptosis by inducing p53 and inhibiting MDM2 expression, which was consistent with in vivo xenograft studies. This innovative strategy, using Nio-Au to simultaneously deliver siRNA (devoid of any chemical modification) and therapeutic drug, provides an efficacious approach for treating therapy-resistant cancers with significant translational potential.

Targeting NANOG and FAK via Cx26-derived Cell-penetrating Peptides in Triple-negative Breast Cancer.

Triple-negative breast cancer (TNBC) represents the most lethal and treatment-resistant breast cancer subtype with limited treatment options. We previously identified a protein complex unique to TNBC composed of the gap junction protein connexin 26 (Cx26), the pluripotency transcription factor NANOG, and focal adhesion kinase (FAK). We sought to determine whether a peptide mimetic of the interaction region of Cx26 attenuated tumor growth in preclinical models. We designed peptides based on Cx26 juxtamembrane domains and performed binding experiments with NANOG and FAK using surface plasmon resonance. Binding studies revealed that the Cx26 C-terminal tail and intracellular loop bound to NANOG and FAK with submicromolar-to-micromolar affinity and that a 5-amino acid sequence in the C-terminal tail of Cx26 (RYCSG) was sufficient for binding. Peptides with high affinity were engineered with a cell-penetrating antennapedia sequence and assessed in functional assays including cell proliferation, tumorsphere formation, and in vivo tumor growth, and downstream signaling changes were measured. The cell-penetrating Cx26 peptide (aCx26-pep) disrupted self-renewal while reducing nuclear FAK and NANOG and inhibiting NANOG target gene expression in TNBC cells but not luminal mammary epithelial cells. In vivo, aCx26-pep reduced tumor growth and proliferation and induced cell death. Here, we provide proof-of-concept that a Cx26 peptide-based strategy inhibits growth and alters NANOG activity specifically in TNBC, indicating the therapeutic potential of this targeting approach.

LCK facilitates DNA damage repair by stabilizing RAD51 and BRCA1 in the nucleus of chemoresistant ovarian cancer.

Poly-ADP Ribose Polymerase (PARP) targeted therapy is clinically approved for the treatment of homologous recombination (HR) repair deficient tumors. The remarkable success of this therapy in the treatment of HR repair deficient cancers has not translated to HR-proficient cancers. Our studies identify the novel role of non-receptor lymphocyte-specific protein tyrosine kinase (LCK) in the regulation of HR repair in endometrioid epithelial ovarian cancer (eEOC) model. We show that DNA damage leads to direct interaction of LCK with the HR repair proteins RAD51 and BRCA1 in a kinase dependent manner RAD51 and BRCA1 stabilization. LCK expression is induced and activated in the nucleus in response to DNA damage insult. Disruption of LCK expression attenuates RAD51, BRCA1, and BRCA2 protein expression by hampering there stability and results in inhibition of HR-mediated DNA repair including suppression of RAD51 foci formation, and augmentation of γH2AX foci formation. In contrast LCK overexpression leads to increased RAD51 and BRCA1 expression with a concomitant increase in HR DNA damage repair. Importantly, attenuation of LCK sensitizes HR-proficient eEOC cells to PARP inhibitor in cells and pre-clinical mouse studies. Collectively, our findings identify a novel therapeutic strategy to expand the utility of PARP targeted therapy in HR proficient ovarian cancer.

PKI 166 induced redox signalling and apoptosis through activation of p53, MAP kinase and caspase pathway in epidermoid carcinoma.

Cellular redox changes have emerged as a pivotal and proximal event in cancer. PKI 166 is used to determine the effects of redox sensitive inhibition of EGFR, metastasis and apoptosis in epidermoid carcinoma. Cytotoxicity study of PKI 166 (IC50 1.0 microM) treated A431 cells were performed by MTT assay for 48 and 72 hrs. Morphological analysis of PKI 166 treated A431 cells for 48 hrs. revealed the cell shrinkage, loss of filopodia and lamellipodia by phase contrast and SEM images in dose dependent manner. It has cytotoxic effects through inhibiting cellular proliferation, leads to the induction of apoptosis, as increased fraction of sub-G1 phase of the cell cycle, chromatin condensation and DNA ladder. It inhibited cyclin-D1 and cyclin-E expression and induced p53, p21 expression in dose dependent manner. Consequently, an imbalance of Bax/Bcl-2 ratio triggered caspase cascade and subsequent cleavage of PARP, thereby shifting the balance in favour of apoptosis. PKI 166 treatment actively stimulated reactive oxygen species (ROS) and mitochondrial membrane depolarization. It inhibited some metastatic properties of A431 cells supressing colony formation by soft agar assay and inhibition of MMP 9 activity by gelatin zymography and western blot analysis. PKI 166 inhibited growth factor induced phosphorylation of EGFR, Akt, MAPK, JNK and colony formation in A431 cells. Thus the inhibition of proliferation was associated with redox regulation of the caspase cascade, EGFR, Akt/PI3K, MAPK/ ERK and JNK pathway. On the other hand, increased antioxidant activity leads to decreased ROS generation inhibit the anti-proliferative and apoptotic properties of PKI 166 in A431 cells. These observations indicated PKI 166 induced redox signalling dependent inhibition of cell proliferation, metastatic properties and induction of apoptotic potential in epidermoid carcinoma.

CD55 in cancer: Complementing functions in a non-canonical manner.

CD55, or decay accelerating factor, is a membrane lipid microdomain-associated, GPI-anchored protein implicated in the shielding of cells from complement-mediated attack via accelerating decay of C3 and C5. Loss of CD55 is associated with a number of pathologies due to hyperactivation of the complement system. CD55 is also implicated in cancer progression thought to be driven via its role in cell shielding mechanisms. We now appreciate that CD55 can signal intracellularly to promote malignant transformation, cancer progression, cell survival, angiogenesis, and inhibition of apoptosis. Outside-in signaling via CD55 is mediated by signaling pathways including JNK, JAK/STAT, MAPK/NF-κB, and LCK. Moreover, CD55 is enriched in the cancer stem cell (CSC) niche of multiple tumors including breast, ovarian, cervical, and can be induced by chemotherapeutics and hypoxic environments. CSCs are implicated in tumor recurrence and chemoresistance. Here, we review the unexpected roles of CD55 in cancer including the roles of canonical and noncanonical pathways that CD55 orchestrates. We will highlight opportunities for therapeutic targeting CD55 and gaps in the field that require more in-depth mechanistic insights.

Disruption of the Gut Microbiota Confers Cisplatin Resistance in Epithelial Ovarian Cancer.

Epithelial ovarian cancer (EOC) is the leading cause of gynecologic cancer death. Despite initial responses to intervention, up to 80% of patient tumors recur and require additional treatment. Retrospective clinical analysis of patients with ovarian cancer indicates antibiotic use during chemotherapy treatment is associated with poor overall survival. Here, we assessed whether antibiotic (ABX) treatment would impact growth of EOC and sensitivity to cisplatin. Immunocompetent or immunocompromised mice were given untreated control or ABX-containing (metronidazole, ampicillin, vancomycin, and neomycin) water prior to intraperitoneal injection with EOC cells, and cisplatin therapy was administered biweekly until endpoint. Tumor-bearing ABX-treated mice exhibited accelerated tumor growth and resistance to cisplatin therapy compared with control treatment. ABX treatment led to reduced apoptosis, increased DNA damage repair, and enhanced angiogenesis in cisplatin-treated tumors, and tumors from ABX-treated mice contained a higher frequency of cisplatin-augmented cancer stem cells than control mice. Stool analysis indicated nonresistant gut microbial species were disrupted by ABX treatment. Cecal transplants of microbiota derived from control-treated mice was sufficient to ameliorate chemoresistance and prolong survival of ABX-treated mice, indicative of a gut-derived tumor suppressor. Metabolomics analyses identified circulating gut-derived metabolites that were altered by ABX treatment and restored by recolonization, providing candidate metabolites that mediate the cross-talk between the gut microbiome and ovarian cancer. Collectively, these findings indicate that an intact microbiome functions as a tumor suppressor in EOC, and perturbation of the gut microbiota with ABX treatment promotes tumor growth and suppresses cisplatin sensitivity. SIGNIFICANCE: Restoration of the gut microbiome, which is disrupted following antibiotic treatment, may help overcome platinum resistance in patients with epithelial ovarian cancer. See related commentary by Hawkins and Nephew, p. 4511.

Coordination-Assisted Self-Assembled Polypeptide Nanogels to Selectively Combat Bacterial Infection.

In the present scenario, the invention of bacteria-selective antimicrobial agent comprising negligible toxicity and hemolytic effect is a great challenge. To surmount this challenge, here, a series of polypeptide nanogels (PNGs) have been fabricated by a coordination-assisted self-assembly of a mannose-conjugated antimicrobial polypeptide, poly(arginine-r-valine)-mannose (poly(Arg-r-Val)-M2), with Zn2+ ions. The fabricated PNGs are spherical in shape with a unique structural appearance similar to that of Taxus baccata fruits. PNGs, with a unique structural arrangement and threshold surface charge density, selectively interact with the bacterial membrane and exhibit potent antimicrobial activity, as reflected in their lower minimum inhibitory concentration values (varies from 2 to 16 µg/mL). PNGs show a remarkably high binding constant, 6.02 × 105 M-1 (from isothermal titration calorimetry, ITC), with the bacterial membrane which manifests its potent bactericidal effect. PNGs are nontoxic against mammalian and red blood cells as reflected from their higher cell viability and insignificant hemolytic effect. PNGs are taken up by the bacterial membrane and selectively undergo structural deformation (scrutinized by ITC) followed by an exposure of free poly(Arg-r-Val)-M2 molecules. The free poly(Arg-r-Val)-M2 molecules are enforced to lyse the bacterial membrane (visualized by cryo-transmission electron microscopy) followed by the diffusion of the cytoplasmic component out of the membrane which culminates in the final death of the bacterium.

Targeting NFE2L2, a transcription factor upstream of MMP-2: A potential therapeutic strategy for temozolomide resistant glioblastoma.

Glioblastoma (GBM) is the most malignant form of brain tumor posing a major threat to cancer amelioration. Temozolomide (TMZ) resistance is one of the major hurdles towards GBM prognosis. Oxidative stress and ECM remodeling are the two important processes involved in gaining chemo-resistance. Here, we established NFE2L2, an important member of oxidative stress regulation elevated in resistant cells, to be playing a transcriptional regulatory role on MMP-2, an ECM remodeling marker. This link led us to further explore targeted molecules to inhibit NFE2L2, thus affecting MMP-2, an important member promoting chemo-resistance. Thus, diosgenin was proposed as a novel NFE2L2 inhibitor acting as an alternative strategy to prevent the high dose administration of TMZ. Combinatorial therapy of diosgenin and TMZ significantly reduced the dosage regimen of TMZ and also showed affectivity in hitherto TMZ resistant GBM cells. GBM cells underwent apoptosis and early cell cycle arrest with significant reduction in MMP-2 levels. Thus preclinical validation of molecular interaction between diosgenin and NFE2L2 down-regulating MMP-2, EMT markers and promoting apoptosis, offers rationale for new therapeutic horizons in the field of glioblastoma management.

A peptide-modified solid lipid nanoparticle formulation of paclitaxel modulates immunity and outperforms dacarbazine in a murine melanoma model.

Melanoma is a highly aggressive skin cancer. A paclitaxel formulation of solid lipid nanoparticles modified with Tyr-3-octreotide (PSM) is employed to treat melanoma that highly expresses somatostatin receptors (SSTRs). PSM exerts more apoptotic and anti-invasive effects in B16F10 mice melanoma cells as compared to dacarbazine (DTIC), an approved chemotherapeutic drug for treating aggressive melanoma. Besides, PSM induces one of the biomarkers of immunogenic cell death in vitro and in vivo as confirmed by calreticulin exposure on the B16F10 cell surface. We observed a significant number of CD8 positive T cells in the tumor bed of the PSM treated group. As a result, PSM effectively reduces tumor volume in vivo as compared to DTIC. PSM also induces a favorable systemic immune response as determined in the spleen and sera of the treated animals. Importantly, PSM can reduce the number of nodule formations in the experimental lung metastasis model. Our experimentations indicate that the metronomic PSM exhibits remarkable anti-melanoma activities without any observable toxicity. This immune modulation behavior of PSM can be exploited for the therapy of melanoma and probably for other malignancies.

Prevention of epithelial to mesenchymal transition in colorectal carcinoma by regulation of the E-cadherin-ß-catenin-vinculin axis.

Epithelial to mesenchymal transition (EMT) is compulsory for metastatic dissemination and is stimulated by TGF-ß. Although targeting EMT has significant therapeutic potential, very few pharmacological agents have been shown to exert anti-metastatic effects. BI-69A11, a competitive Akt inhibitor, displays anti-tumor activity toward melanoma and colon carcinoma. This study provides molecular and biochemical insights into the effects of BI-69A11 on EMT in colon carcinoma cells in vitro and in vivo. BI-69A11 inhibited metastasis-associated cellular migration, invasion and adhesion by inhibiting the Akt-ß-catenin pathway. The underlying mechanism of BI-69A11-mediated inhibition of EMT included suppression of nuclear transport of ß-catenin and diminished phosphorylation of ß-catenin, which was accompanied by enhanced E-cadherin-ß-catenin complex formation at the plasma membrane. Additionally, BI-69A11 caused increased accumulation of vinculin in the plasma membrane, which fortified focal adhesion junctions leading to inhibition of metastasis. BI-69A11 downregulated activation of the TGF-ß-induced non-canonical Akt/NF-κB pathway and blocked TGF-ß-induced enhanced expression of Snail causing restoration of E-cadherin. Overall, this study enhances our understanding of the molecular mechanism of BI-69A11-induced reversal of EMT in colorectal carcinoma cells in vitro, in vivo and in TGF-ß-induced model systems.

Multi-nucleated cells use ROS to induce breast cancer chemo-resistance in vitro and in vivo.

Although there is a strong correlation between multinucleated cells (MNCs) and cancer chemo-resistance in variety of cancers, our understanding of how multinucleated cells modulate the tumor micro-environment is limited. We captured multinucleated cells from triple-negative chemo-resistant breast cancers cells in a time frame, where they do not proliferate but rather significantly regulate their micro-environment. We show that oxidatively stressed MNCs induce chemo-resistance in vitro and in vivo by secreting VEGF and MIF. These factors act through the RAS/MAPK pathway to induce chemo-resistance by upregulating anti-apoptotic proteins. In MNCs, elevated reactive oxygen species (ROS) stabilizes HIF-1α contributing to increase production of VEGF and MIF. Together the data indicate, that the ROS-HIF-1α signaling axis is very crucial in regulation of chemo-resistance by MNCs. Targeting ROS-HIF-1α in future may help to abrogate drug resistance in breast cancer.

Resensitization of Akt Induced Docetaxel Resistance in Breast Cancer by 'Iturin A' a Lipopeptide Molecule from Marine Bacteria Bacillus megaterium.

Development of the resistance is the major problem in cancer therapy. Docetaxel is a taxol alkaloid that is frequently used in metastatic breast cancer. However, resistance often limits the usefulness of this drug in many breast cancer patients. Manipulation of resistant cells to re-sensitize to the therapeutic effect of docetaxel is current strategy to overcome this problem. Here, we have introduced 'Iturin A' as a potent chemosensitizer in docetaxel resistant breast cancer cells. Combination of Iturin A and docetaxel treatment significantly hampered the proliferation of docetaxel resistant MDA-MB-231 and MDA-MB-468 breast cancer cells. Cell cycle analysis also showed massive amount of apoptotic population (Sub G0/G1) in combination therapy. A number of apoptotic and anti-apoptotic proteins were significantly altered in dual drug treated groups. Caspase 3 dependent cell death was observed in dual treatment. Molecular mechanism study showed that over-expression of Akt and its downstream signaling pathway was associated with docetaxel resistance. Iturin A significantly reduced Akt signaling pathway in resistant cells. This mechanistic action might be the reason behind the chemo-sensitization effect of Iturin A in docetaxel resistant breast cancer cells. In conclusion, Iturin A resensitized the resistant breast cancer cells to docetaxel therapy by inhibiting Akt activity.

Therapeutic implication of 'Iturin A' for targeting MD-2/TLR4 complex to overcome angiogenesis and invasion.

Tumor angiogenesis and invasion are deregulated biological processes that drive multistage transformation of tumors from a benign to a life-threatening malignant state activating multiple signaling pathways including MD-2/TLR4/NF-κB. Development of potential inhibitors of this signaling is emerging area for discovery of novel cancer therapeutics. In the current investigation, we identified Iturin A (A lipopeptide molecule from Bacillus megaterium) as a potent inhibitor of angiogenesis and cancer invasion by various in vitro and in vivo methods. Iturin A was found to suppress VEGF, a powerful inducer of angiogenesis and key player in tumor invasion, as confirmed by ELISA, western blot and real time PCR. Iturin A inhibited endothelial tube arrangement, blood capillary formation, endothelial sprouting and vascular growth inside the matrigel. In addition, Iturin A inhibited MMP-2/9 expression in MDA-MB-231 and HUVEC cells. Cancer invasion, migration and colony forming ability were significantly hampered by Iturin A. Expressions of MD-2/TLR4 and its downstream MyD88, IKK-α and NF-κB were also reduced in treated MDA-MB-231 and HUVEC cells. Western blot and immunofluorescence study showed that nuclear accumulation of NF-κB was hampered by Iturin A. MD-2 siRNA or plasmid further confirmed the efficacy of Iturin A by suppressing MD-2/TLR4 signaling pathway. The in silico docking study showed that the Iturin A interacted well with the MD-2 in MD-2/TLR4 receptor complex. Conclusively, inhibition of MD-2/TLR4 complex with Iturin A offered strategic advancement in cancer therapy.

Somatostatin receptor targeted liposomes with Diacerein inhibit IL-6 for breast cancer therapy.

Selective targeting to the tumor niche remains a major challenge in successful cancer therapy. Somatostatin receptor 2 (SSTR2) is overexpressed in breast cancer cells thus making this receptor an attractive target for selective guidance of ligand-conjugated drug liposomes to the tumor site. In this study, a synthetic somatostatin analogue (SST) was used as SSTR2 targeting agent and Diacerein was employed as therapeutic molecule. Diacerein loaded liposomes (DNL) were prepared and they were further decorated with the synthetic and stable analogue of somatostatin (SST-DNL). Fabricated liposomes were nano-size in range and biocompatible. SST-DNL displayed significantly better anti-tumor efficacy as compared to free Diacerein (DN) and DNL in breast cancer models. Enhanced apoptosis in breast cancer cells was detected in SST-DNL treated groups as monitored by cell cycle analysis and changes in expression level of apoptotic/anti-apoptotic proteins Bcl-2, Bax, cleaved Caspase 3 and PARP. SST-DNL more effectively inhibited the oncogenic IL-6/IL-6R/STAT3/MAPK/Akt signalling pathways as compared to DN or DNL in cancer cells. In addition, SST-DNL effectively suppressed angiogenesis and cancer cell invasion. In vivo tumor growth in a MDA-MB-231 mouse xenograft model was significantly suppressed following SST-DNL treatment. In xenograft model, immunohistochemistry of Ki-67 and CD-31 indicated that SST-DNL improved the anti-proliferative and anti-angiogenic impacts of Diacerein. In vivo pharmacokinetic studies in rats showed enhanced circulation time in the DNL or SST-DNL treated groups as compared to free DN. Considering all of these findings, we conclude that SST-DNL provides a novel strategy with better efficacy for breast cancer therapy.