Systemic Delivery of an Adjuvant CXCR4-CXCL12 Signaling Inhibitor Encapsulated in Synthetic Protein Nanoparticles for Glioma Immunotherapy.

Glioblastoma (GBM) is an aggressive primary brain cancer, with a 5 year survival of ∼5%. Challenges that hamper GBM therapeutic efficacy include (i) tumor heterogeneity, (ii) treatment resistance, (iii) immunosuppressive tumor microenvironment (TME), and (iv) the blood-brain barrier (BBB). The C-X-C motif chemokine ligand-12/C-X-C motif chemokine receptor-4 (CXCL12/CXCR4) signaling pathway is activated in GBM and is associated with tumor progression. Although the CXCR4 antagonist (AMD3100) has been proposed as an attractive anti-GBM therapeutic target, it has poor pharmacokinetic properties, and unfavorable bioavailability has hampered its clinical implementation. Thus, we developed synthetic protein nanoparticles (SPNPs) coated with the transcytotic peptide iRGD (AMD3100-SPNPs) to target the CXCL2/CXCR4 pathway in GBM via systemic delivery. We showed that AMD3100-SPNPs block CXCL12/CXCR4 signaling in three mouse and human GBM cell cultures in vitro and in a GBM mouse model in vivo. This results in (i) inhibition of GBM proliferation, (ii) reduced infiltration of CXCR4+ monocytic myeloid-derived suppressor cells (M-MDSCs) into the TME, (iii) restoration of BBB integrity, and (iv) induction of immunogenic cell death (ICD), sensitizing the tumor to radiotherapy and leading to anti-GBM immunity. Additionally, we showed that combining AMD3100-SPNPs with radiation led to long-term survival, with ∼60% of GBM tumor-bearing mice remaining tumor free after rechallenging with a second GBM in the contralateral hemisphere. This was due to a sustained anti-GBM immunological memory response that prevented tumor recurrence without additional treatment. In view of the potent ICD induction and reprogrammed tumor microenvironment, this SPNP-mediated strategy has a significant clinical translation applicability.

G-CSF secreted by mutant IDH1 glioma stem cells abolishes myeloid cell immunosuppression and enhances the efficacy of immunotherapy.

Mutant isocitrate-dehydrogenase 1 (mIDH1) synthesizes the oncometabolite 2-hydroxyglutarate (2HG), which elicits epigenetic reprogramming of the glioma cells' transcriptome by inhibiting DNA and histone demethylases. We show that the efficacy of immune-stimulatory gene therapy (TK/Flt3L) is enhanced in mIDH1 gliomas, due to the reprogramming of the myeloid cells' compartment infiltrating the tumor microenvironment (TME). We uncovered that the immature myeloid cells infiltrating the mIDH1 TME are mainly nonsuppressive neutrophils and preneutrophils. Myeloid cell reprogramming was triggered by granulocyte colony-stimulating factor (G-CSF) secreted by mIDH1 glioma stem/progenitor-like cells. Blocking G-CSF in mIDH1 glioma­bearing mice restores the inhibitory potential of the tumor-infiltrating myeloid cells, accelerating tumor progression. We demonstrate that G-CSF reprograms bone marrow granulopoiesis, resulting in noninhibitory myeloid cells within mIDH1 glioma TME and enhancing the efficacy of immune-stimulatory gene therapy.

Targeting Neuroinflammation in Brain Cancer: Uncovering Mechanisms, Pharmacological Targets, and Neuropharmaceutical Developments.

Gliomas are one of the most lethal types of cancers accounting for ∼80% of all central nervous system (CNS) primary malignancies. Among gliomas, glioblastomas (GBM) are the most aggressive, characterized by a median patient survival of fewer than 15 months. Recent molecular characterization studies uncovered the genetic signatures and methylation status of gliomas and correlate these with clinical prognosis. The most relevant molecular characteristics for the new glioma classification are IDH mutation, chromosome 1p/19q deletion, histone mutations, and other genetic parameters such as ATRX loss, TP53, and TERT mutations, as well as DNA methylation levels. Similar to other solid tumors, glioma progression is impacted by the complex interactions between the tumor cells and immune cells within the tumor microenvironment. The immune system's response to cancer can impact the glioma's survival, proliferation, and invasiveness. Salient characteristics of gliomas include enhanced vascularization, stimulation of a hypoxic tumor microenvironment, increased oxidative stress, and an immune suppressive milieu. These processes promote the neuro-inflammatory tumor microenvironment which can lead to the loss of blood-brain barrier (BBB) integrity. The consequences of a compromised BBB are deleteriously exposing the brain to potentially harmful concentrations of substances from the peripheral circulation, adversely affecting neuronal signaling, and abnormal immune cell infiltration; all of which can lead to disruption of brain homeostasis. In this review, we first describe the unique features of inflammation in CNS tumors. We then discuss the mechanisms of tumor-initiating neuro-inflammatory microenvironment and its impact on tumor invasion and progression. Finally, we also discuss potential pharmacological interventions that can be used to target neuro-inflammation in gliomas.

Human Papillomavirus Induced Cervical and Oropharyngeal Cancers: From Mechanisms to Potential Immuno-therapeutic Strategies.

The human papillomavirus (HPV) associated infections are the hallmark of cervical and neck cancer. Almost all the cases of cervical cancer (CC) and 70% of oropharyngeal cancer (OC) are, more or less, caused by the persistent infection of HPV. CC is the fourth most common cancer globally, and is commenced by the persistent infection with human papillomaviruses (HPVs), predominantly HPV types; 16 and 18. In the light of the above facts, there is an immediate requirement to develop novel preventive and innovative therapeutic strategies that may help in lower occurrences of HPV mediated cancers. Currently, only radiation and chemical-based therapies are the treatment for HPV mediated neck cancer (NC) and CC. Recent advances in the field of immunotherapy are underway, which are expected to unravel the optimal treatment strategies for the growing HPV mediated cancers. In this review, we decipher the mechanism of pathogenesis with current immunotherapeutic advances in regressing the NC and CC, with an emphasis on immune-therapeutic strategies being tested in clinical trials and predominantly focus on defining the efficacy and limitations. Taken together, these immunological advances have enhanced the effectiveness of immunotherapy and promises better treatment results in coming future.

A Study on Serological Reactivity Profile of Different Antigen Preparations with Bancroftian filariasis Human Infection Sera.

BACKGROUND: Lymphatic Filariasis (LF) is one of the incapacitating and mosquito-borne sicknesses that on progression may prompt a few recognizable types of clutters like extreme lymphedema, hydrocele, and elephantiasis. METHODS: Antigenic preparations of B. malayi adult (BmA), S. cervi adult parasites and microfilariae (mf) total parasite extract were used to analyze the serological reactivity profile with human infectious sera collected from endemic areas of Bancroftian filariasis by performing Western blot and ELISA analysis. Sera from healthy human subjects were also included in the study to determine the variation incurred in the reactivity due to the filariasis infection. Gelelectrophoresis analysis of the crude-extract of BmA revealed seven protein bands while more than ten bands were recognized in S. cervi. RESULTS: our results represent a clear variation in protein patterns among the crude-antigens. ELISA results showed highest prevalence of IgG, IgM and IgG4 antibodies against all antigen preparations when recorded among microfilaraemic chronic infected patients. In both the antigenic preparations, the positive reactions were in the order of microfilaraemic>endemic normal>chronic>acute>nonendemic normal subjects. All sera of Mf+ patients were uniformly positive, while sera of both chronic and endemic normal subjects showed less reactivity. CONCLUSION: In the present study, we endeavoured to establish the extent of cross-reactivity of antigens derived from animal filarial parasites such as B. malayi and S. cervi with W. bancrofti filariasis sera of human patients. Besides, we further analyzed antibody-isotype profile of IgG, IgG4 and IgM in various human infection sera of bancroftian filarial subjects reactive to heterologous parasite antigens derived from adult worms of S. cervi from bovine and B. malayi from bovine and jirds.

Bio-mediated synthesis of 5-FU based nanoparticles employing orange fruit juice: a novel drug delivery system to treat skin fibrosarcoma in model animals.

Nano-sized drug delivery systems (NDDS) have been widely exploited to achieve targeted delivery of pharmaco-materials. Traditional pharmaceutical approaches, implied in the synthesis of nano-formulations, are obscure owing to the incompatible physico-chemical properties of the core drug as well as some other factors crucial in development of NDDS. Infact, most of the existing methods used in development of NDDS rely on usage of additives or excipients, a special class of chemicals. Barring few exceptions, the usage of synthetic excipients ought to be curtailed because of several associated undesirable features. Such issues necessitate strategies that lead to development of the synthetic excipient free drug delivery system. Plant based extracts have great potential to induce synthesis of nano-sized particles. Considering this fact, here we propose a prototype employing orange fruit juice (OJ) to facilitate bio-mediated synthesis of nano-sized supra-molecular assemblies of 5-fluorouracil (5-FU), a potent anticancer drug. The as-synthesized 5-FU Nanoparticles (NPs) retained the anti-neoplastic efficacy of the parent compound and induced apoptosis in cancer cells. The novel 5-FU NPs formulation demonstrated enhanced efficacy against DMBA induced experimental fibrosarcoma in the mouse model when compared to the micro-sized crystals of parent 5-FU drug.

Recent Findings on Nanotechnology-based Therapeutic Strategies Against Hepatocellular Carcinoma.

BACKGROUND: Nanotechnology-based therapies are emerging as a promising new anticancer approach. Early clinical studies suggest that nanoparticle-based therapeutics can show enhanced efficacy while reducing side effects minimal, owing to targeted delivery and active intracellular uptake. METHODS: To overcome the problems of gene and drug delivery, nanotechnology based delivery system gained interest in the last two decades. Encouraging results from Nano formulation based drug delivery systems revealed that these emerging restoratives can efficiently lead to more effective, targeted, selective and efficacious delivery of chemotherapeutic agents to the affected target cells. RESULTS: Nanotechnology not only inhibits targeted gene products in patients with cancer, but also taught us valuable lessons regarding appropriate dosages and route of administrations. Besides, nanotechnology based therapeutics holds remarkable potential as an effective drug delivery system. We critically highlight the recent findings on nanotechnology mediated therapeutics strategies to combat hepatocellular carcinoma and discuss how nanotechnology platform can have enhanced anticancer effects compared with the parent therapeutic agents they contain. CONCLUSION: In this review, we discussed the key challenges, recent findings and future perspective in the development of effective nanotechnology-based cancer therapeutics. The emphasis here is focused on nanotechnology-based therapies that are likely to affect clinical investigations and their implications for advancing the treatment of patients with hepatocellular carcinoma.

Zinc oxide-decorated polypyrrole/chitosan bionanocomposites with enhanced photocatalytic, antibacterial and anticancer performance.

A bio-nanocomposite matrix of polypyrrole grafted ZnO/chitosan (Ppy/C/Z) was synthesized via the in situ polymerization of pyrrole with different weight fractions of ZnO. Incorporation of ZnO nanoparticles with polypyrrole enhances the photocatalytic, antibacterial as well as cytotoxic properties of the resultant composite. Characterizations of the synthesized product were performed by X-ray diffraction (XRD), Fourier transform infrared spectroscopy (FTIR) and thermal analysis (TGA and DTA). Surface morphology and particle size were determined by SEM and TEM. The elemental composition of the material was studied by EDX coupled with SEM. Electrochemical surface area was calculated from electrochemical double layer capacitance (EDLC) measurements using cyclic voltammetry. The photocatalytic activity of the composite material was tested by monitoring the degradation of reactive orange-16 (RO-16), Coomassie Brilliant Blue R-250 (CBB-R-250) and Methylene Blue (MB) dyes and the composite was found to be an effective catalyst in the presence of a UV light source. Various scavengers were used to detect the reactive species involved in the photocatalytic process. Furthermore, the stability of the photocatalyst was assessed by recycling experiments. Moreover, the Ppy/C/Z bio-nanocomposite shows potential application with anti-bacterial and anti-cancer activity against Gram-positive and Gram-negative bacterial pathogens and human cancer cell lines (HeLa and MCF-7). The experimental data confirm that the bio-nanocomposite of Ppy/C/Z showed excellent anti-bacterial and anti-cancer activity as compared to a pristine polypyrrole and chitosan formulation (Ppy/C). The apoptosis data with varying concentrations of Ppy/C/Z reveal the remarkable activity against these cancer cell lines.

Status of lymphatic filariasis with progression of age and gender & eradication strategies: A survey among residents of Hardoi district of Uttar Pradesh, an endemic region of North India.

Lymphatic filariasis (LF) is a chronic and debilitating disease that affects people in tropical and sub-tropical areas of Asia, Africa, and Western Pacific. It is one of the leading community health problems in some of the endemic districts in India including Hardoi district of Uttar Pradesh. The disease is caused by the parasites Wuchereria bancrofti (W. bancrofti), Brugia malayi (B. malayi) and Brugia timori (B. timori), transmitted by the vector Culex, Anopheles and other mosquitoes. This cross-sectional survey study was carried out in rural areas, where its inhabitants vary in socio-economic status, from low to middle-income class. 12 villages of Hardoi district, Uttar Pradesh, India were included. The aim was to see the impact of age and gender on various clinical forms of LF and in estimating its economic and social implications. 260 LF affected people in different parts of Hardoi district were surveyed. The results revealed that the Mass Drug Administration (MDA) coverage reached more than 90%. The overall Microfilaria rate had been reduced, however the prevalence of elephantiasis increased with the progression of age and was found to be highest among people of >70 years of age, regardless of their gender.

A Synopsis on the Role of Human Papilloma Virus Infection in Cervical Cancer.

BACKGROUND: Understanding of cervical cancer severity is still an important health issue across the world, especially for developing countries. Cancer or abnormal growth of the cell is one of the major health problems of the world. There are about two hundred types of malignancies reported till date. An updated statistic of all the main types of cancer and pathophysiology of cervical cancer is a significant need for designing the future treatment strategy. OBJECTIVE: In this review, a brief update on cancer, its causes and different types has been discussed along with updated statistics of patient's mortality. A brief overview of cervical cancer and its pathophysiology has been discussed with special emphasis on its causative agent, human papilloma virus (HPV). A brief introduction and update on genetics, molecular pathogenesis and prevalence of HPV and its role in cervical cancer have been added. CONCLUSION: This review delivered an updated status of cervical cancer and provide novel therapeutic approaches for targeting HPV. The detailed molecular and genomic information of the HPV help the researchers to develop more effective and efficacious therapeutic strategies and preventive vaccines that will significantly contribute to the control and anticipation of cervical cancer. Ultimately this may open new vistas to get rid of this deadly disease and may offer significant reduction in the numbers of advanced cervical cancers and deaths from cervical cancer in the affected nations.

Emerging role of Interleukins IL-23/IL-17 axis and biochemical markers in the pathogenesis of Type 2 Diabetes: Association with age and gender in human subjects.

Chronic hyperglycaemia in type 2 diabetes (T2D) is associated with increased oxidative stress and inflammation. Keeping the above fact into consideration we analyse the effect of age and gender on oxidative stress biomarkers and pro-inflammatory cytokines in T2D patients. The study included 148 diabetic and 110 healthy subjects, grouped on the basis of age and gender. Plasma malondialdehyde, protein carbonyl content and nitric oxide levels were elevated significantly in diabetic patients, with significant decrease in Ferric reducing ability of plasma, vitamin C, reduced glutathione, erythrocyte thiol groups and erythrocyte antioxidant enzyme activity and these changes were even more pronounced as age progressed. Serum IL-1ß, IL-6, IL-17A, IL-22 levels and TNF-α mRNA expression was significantly upregulated in all the age groups whereas IL-23 mRNA was upregulated only in the higher age group. Female diabetic patients experienced higher oxidative stress and greater serum IL-6 levels and TNF-α mRNA expression as compared to their male counterparts. This study suggested that diabetes onset is accompanied with increased oxidative stress and elevated levels of inflammatory mediators. The effect was more prominent in aged patients. Female patients experienced greater oxidative stress as compared to males of those age groups with slightly higher levels of inflammatory cytokines.

Undec-10-enoic acid functionalized chitosan based novel nano-conjugate: An enhanced anti-bacterial/biofilm and anti-cancer potential.

Fatty acid functionalized chitosan conjugates are of great interest in cancer therapeutics because of its internalization through receptor mediated endocytosis into the cancer cells. Keeping the above fact into consideration, herein we synthesized the undec-10-enoic acid functionalized chitosan based undecyl-chitosan (U-CS) nano-bioconjugate with the use of DCC as a coupling agent. The U-CS conjugate synthesized was confirmed and characterized by FTIR, 1H NMR, TGA, XRD, SEM and TEM analysis. Generally, it is well established that conjugates of oleic acid with human Alpha-lactalbumin (HAMLET) induce cytotoxicity in the altered cells, but not in healthy cells. To check our presumptions, anti-bacterial and anti-cancer potential of U-CS was evaluated against bacterial pathogens (Gram +ve and Gram -ve) and human cancer cell lines (HeLa, MDA-MB-231 and Hep3B). The results of our study clearly revealed that conjugate showed enhance anti-bacterial, anti-biofilm as well as anti-cancer efficacy as compared to pure and free form of the chitosan.

Molecular and phenotypic characterization of the vancomycin-resistant gene in bacterial isolates acquired from catheter tips.

Gram-positive bacteria, particularly Staphylococcus aureus is a significant pathogen, not only in the hospital setting but the community also. S. aureus is a major cause of serious hospital and community-acquired infections, particularly in the colonized individuals. The emergence of vancomycin-resistant S. aureus (VRSA) strains has led to global concerns about treatments for staphylococcal infections. Until now, few strains of VRSA have been reported worldwide. The conventional disk diffusion method for determination of vancomycin sensitivity often misclassifies intermediately susceptible isolates to fully sensitive. However, non-automated minimum inhibitory concentration (MIC) detection methods are the gold standards. Hence there is a dire need of some advanced methods for rapid detection of VRSA strains. In the present study, Gram-positive clinical isolates were collected from different wards of K.G.M.U.  Hospital, among them, 12 bacterial isolates were identified as Staphylococcus aureus and 18 isolates as Klebsiella spp. Genomic DNA of S. aureus was isolated and used as template in PCR for detection of the presence of van A and van X gene based on a given protocol. Nosocomial infections have an impact on morbidity and probably on mortality as well, and pose a significant economic burden. Rapid molecular identification of antibiotic-resistant strains undoubtedly helps to prevent the hospital-induced infections.

Role of Pro-Inflammatory Cytokines and Biochemical Markers in the Pathogenesis of Type 1 Diabetes: Correlation with Age and Glycemic Condition in Diabetic Human Subjects.

BACKGROUND: Type 1 diabetes mellitus is a chronic inflammatory disease involving insulin producing ß-cells destroyed by the conjoined action of auto reactive T-cells, inflammatory cytokines and monocytic cells. The aim of this study was to elucidate the status of pro-inflammatory cytokines and biochemical markers and possible correlation of these factors towards outcome of the disease. METHODS: The study was carried out on 29 T1D subjects and 20 healthy subjects. Plasma levels of oxidative stress markers, enzymatic and non-enzymatic antioxidants were estimated employing biochemical assays. The levels of pro-inflammatory cytokines such as by IL-1ß & IL-17 in the serum were determined by ELISA, while the expression of TNF-α, IL-23 & IFN-γ was ascertained by qRT-PCR. RESULTS: The onset of T1D disease was accompanied with elevation in levels of Plasma malondialdehyde, protein carbonyl content and nitric oxide while plasma vitamin C, reduced glutathione and erythrocyte sulfhydryl groups were found to be significantly decreased in T1D patients as compared to healthy control subjects. Activity of antioxidant enzymes, superoxide dismutase, catalase, glutathione reductase and glutathione-s-transferase showed a significant suppression in the erythrocytes of T1D patients as compared to healthy subjects. Nevertheless, the levels of pro-inflammatory cytokines IL-1ß and IL-17A were significantly augmented (***p≤.001) on one hand, while expression of T cell based cytokines IFN-γ, TNF-α and IL-23 was also up-regulated (*p≤.05) as compared to healthy human subjects. CONCLUSION: The level of pro-inflammatory cytokines and specific biochemical markers in the serum of the patient can be exploited as potential markers for type 1 diabetes pathogenesis. The study suggests that level of inflammatory markers is up-regulated in T1D patients in an age dependent manner.

TLR Agonist Augments Prophylactic Potential of Acid Inducible Antigen Rv3203 against Mycobacterium tuberculosis H37Rv in Experimental Animals.

In general, the members of Lip gene family of Mycobacterium tuberculosis evoke strong immune response in the host. Keeping this fact into consideration, we investigated role of Rv3203, a cell wall associated protein with lipolytic activity, in imparting protection against experimental murine tuberculosis. The data of the present study suggested that archaeosome encapsulated Rv3203 induce strong lymphocyte proliferation, up-regulated Th-1 biased cytokines profile, increased expression of co-stimulatory markers on both antigen presenting cells and T lymphocytes. The immuno-prophylactic response was further modulated by exposure of the animals to zymosan, a TLR2/6 agonist, prior to immunization with archaeosome encapsulated Rv3203. Interestingly, pre-treatment of experimental animals with zymosan boosted strong immunological memory as compared to archaeosome encapsulated Rv3203 as well as BCG vaccine. We conclude that priming of immunized animal with TLR agonist followed by immunization with archaeosomes encapsulated Rv3203 offer substantial protection against tuberculosis infection and could be a potential subunit vaccine based prophylactic strategy.

Amyloid form of ovalbumin evokes native antigen-specific immune response in the host: prospective immuno-prophylactic potential.

Amyloids are highly organized protein aggregates that arise from inappropriately folded versions of proteins or polypeptides under both physiological as well as simulated ambiences. Once thought to be irreversible assemblies, amyloids have begun to expose their more dynamic and reversible attributes depending upon the intrinsic properties of the precursor protein/peptide and experimental conditions such as temperature, pressure, structural modifications in proteins, or presence of chemicals in the reaction mixture. It has been repeatedly proposed that amyloids undergo transformation to the bioactive peptide/protein forms under specific conditions. In the present study, amyloids assembled from the model protein ovalbumin (OVA) were found to release the precursor protein in a slow and steady manner over an extended time period. Interestingly, the released OVA from amyloid depot was found to exhibit biophysical characteristics of native protein and reacted with native-OVA specific monoclonal as well as polyclonal antibodies. Moreover, antibodies generated upon immunization of OVA amyloidal aggregates or fibrils were found to recognize the native form of OVA. The study suggests that amyloids may act as depots for the native form of the protein and therefore can be exploited as vaccine candidates, where slow antigen release over extended time periods is a pre-requisite for the development of desired immune response.