Modulation of carcinogenesis with selected GRAS nutraceuticals via Keap1-Nrf2 signaling pathway.

Keap1-Nrf2 is a fundamental signaling cascade known to promote or prevent carcinogenesis. Extensive studies identify the key target of modulatory aspects of Keap1-Nrf2 signaling against cancer. Nutraceuticals are those dietary agents with many health benefits that have immense potential for cancer chemoprevention. The nutritional supplements known as nutraceuticals are found to be one of the most promising chemoprevention agents. Upon investigating the dual nature of Nrf2, it became clear that, in addition to shielding normal cells from numerous stresses, Nrf2 may also promote the growth of tumors. In the present review, we performed a systematic analysis of the role of 12 different nutraceuticals like curcumin, sulforaphane, resveratrol, polyunsaturated fatty acids (PUFA) from fish oil, lycopene, soybean, kaempferol, allicin, thymoquinone, quercetin, gingerol, and piperine in modulating the Nrf2/Keap1 signaling mechanism. Among these, 12 Generally Recognized As Safe (GRAS) certified nutraceuticals, sulforaphane is the most extensively studied compound in modulating Keap1-Nrf signaling. Even though there is much evidence at preclinical levels, further high-quality research is still required to validate the potential role of these nutraceuticals in Keap1-Nrf2 modulation.

Corrigendum: Targeting thymidylate synthase enhances the chemosensitivity of triple-negative breast cancer towards 5-FU-based combinatorial therapy.

[This corrects the article DOI: 10.3389/fonc.2021.656804.].

Harnessing the immune system against cancer: current immunotherapy approaches and therapeutic targets.

Cancer immunotherapy is a rapidly evolving concept that has been given the tag "fifth pillar" of cancer therapy while radiation therapy, chemotherapy, surgery and targeted therapy remain the other four pillars. This involves the stimulation of the immune system to control tumor growth and it specifically targets the neoplastic cells rather than the normal cells. Conventional chemotherapy has many limitations which include drug resistance, recurrence of cancer and severe adverse effects. Immunology has made major treatment breakthroughs for several cancers such as colorectal cancer, prostate cancer, breast cancer, lung cancer, liver cancer, kidney cancer, stomach cancer, acute lymphoblastic leukaemia etc. Currently, therapeutic strategies harnessing the immune system involve Checkpoint inhibitors, Chimeric antigen receptor T cells (CAR T cells), Monoclonal antibodies, Cancer vaccines, Cytokines, Radio-immunotherapy and Oncolytic virus therapy. The molecular characterization of several tumor antigens (TA) indicates that these TA can be utilized as promising candidates in cancer immunotherapy strategies. Here in this review, we highlight and summarize the different categories of emerging cancer immunotherapies along with the immunologically recognized tumor antigens involved in the tumor microenvironment.

Targeting Thymidylate Synthase Enhances the Chemosensitivity of Triple-Negative Breast Cancer Towards 5-FU-Based Combinatorial Therapy.

BACKGROUND: The ongoing treatment modalities for breast cancer (BC) primarily rely on the expression status of ER, PR and HER-2 receptors in BC tissues. Our strategy of chemosensitization provides new insights to counter chemoresistance, a major obstacle that limits the benefits of chemotherapy of mammary cancers. METHODS: By utilizing a murine breast cancer model employing NSG mice bearing orthotopic triple-negative breast cancer (TNBC) xenografts, we have evaluated the ability of phytochemical curcumin in chemosensitizing BC to 5-Fluorouracil (5-FU) chemotherapy and the differential modulations of cellular events in response to this strategy, independent of their receptor status. RESULTS: A significant synergistic antitumor potential was observed in the murine model with a sub-optimal dose treatment of 5-FU plus curcumin, as evaluated by a reduction in the tumor-related parameters. We authenticated the pivotal role of thymidylate synthase (TS) in regulating the 5-FU-curcumin synergism using the TNBC pre-clinical model. Our study also confirmed the pharmacological safety of this chemotherapeutic plus phytoactive combination using acute and chronic toxicity studies in Swiss albino mice. Subsequently, the molecular docking analysis of curcumin binding to TS demonstrated the affinity of curcumin towards the cofactor-binding site of TS, rather than the substrate-binding site, where 5-FU binds. Our concomitant in vivo and in silico evidence substantiates the superior therapeutic index of this combination. CONCLUSION: This is the first-ever pre-clinical study portraying TS as the critical target of combinatorial therapy for mammary carcinomas and therefore we recommend its clinical validation, especially in TNBC patients, who currently have limited therapeutic options.

Chitosan Encapsulation Enhances the Bioavailability and Tissue Retention of Curcumin and Improves its Efficacy in Preventing B[a]P-induced Lung Carcinogenesis.

The rate of lung cancer incidence is alarmingly mounting, despite the decline of smoking and tobacco consumption. Recent reports indicate a very high correlation between the growing fast food culture and lung cancer incidence. Benzo[a]pyrene (B[a]P) is a potent carcinogen abundantly present in grilled and deep-fried food and in tobacco smoke. Our previous studies have proved the efficacy of curcumin in curbing B[a]P-induced lung carcinogenesis. However, the poor pharmacokinetic profile of the compound considerably hampers its potential as an effective chemopreventive. This study was intended to evaluate whether encapsulation of curcumin in chitosan nanoparticles can improve the cellular uptake and prolong the tissue retention of curcumin yielding better chemoprevention. The curcumin-loaded chitosan nanoparticles (chitosan nanocurcumin) exhibited a size of 170-200 nm in transmission electron microscopy. In vitro drug release studies showed sustained release of curcumin over a period of approximately 180 hours and excellent intracellular uptake and cytotoxicity in lung cancer cells. Bioavailability studies using healthy Swiss albino mice demonstrated drastic enhancement in lung localization of chitosan nanocurcumin compared with free curcumin. Toxicologic evaluation using chronic toxicity model in Swiss albino mice confirmed the pharmacologic safety of the formulation. Moreover, the formulation, even at a dose equivalent to one fourth that of free curcumin, exhibits better efficacy in reducing tumor incidence and multiplicity than free curcumin, thereby hampering development of B[a]P-induced lung adenocarcinomas in Swiss albino mice. Hence, our study underscores the supremacy of the formulation over free curcumin and establishes it as a potential chemopreventive and oral supplement against environmental carcinogenesis.

Novel flourescent spiroborate esters: potential therapeutic agents in in vitro cancer models.

The current treatment system in cancer therapy, which includes chemotherapy/radiotherapy is expensive and often deleterious to surrounding healthy tissue. Presently, several medicinal plants and their constituents are in use to manage the development and progression of these diseases.They have been found effective, safe, and less expensive. In the present study, we are proposing the utility of a new class of curcumin derivative, Rubrocurcumin, the spiroborate ester of curcumin with boric acid and oxalic acid (1:1:1), which have enhanced biostability for therapeutic applications. In vitro cytocompatibility of this drug complex was analysed using MTT assay, neutral red assay, lactate dehydrogenase assay in 3T3L1 adipocytes. Anti tumour activity of this drug complex on MCF7 and A431 human cancer cell line was studied by morphological analysis using phase contrast microscopy, Hoechst staining and cell cycle analysis by FACS. To explore the chemotherapeutic effect, the cytotoxic effect of this compound was also carried out. Rubrocurcumin is more biostable than natural curcumin in physiological medium. Our results prove that this curcumin derivative drug complex possess more efficacy and anti-cancer activity compared with curcumin. The findings out of this study suggests this novel compound as potential candidate for site targeted drug delivery.

JUN is a key transcriptional regulator of the unfolded protein response in acute myeloid leukemia.

The transcription factor JUN is frequently overexpressed in multiple genetic subtypes of acute myeloid leukemia (AML); however, the functional role of JUN in AML is not well defined. Here we report that short hairpin RNA (shRNA)-mediated inhibition of JUN decreases AML cell survival and propagation in vivo. By performing RNA sequencing analysis, we discovered that JUN inhibition reduces the transcriptional output of the unfolded protein response (UPR), an intracellular signaling transduction network activated by endoplasmic reticulum (ER) stress. Specifically, we found that JUN is activated by MEK signaling in response to ER stress, and that JUN binds to the promoters of several key UPR effectors, such as XBP1 and ATF4, to activate their transcription and allow AML cells to properly negotiate ER stress. In addition, we observed that shRNA-mediated inhibition of XBP1 or ATF4 induces AML cell apoptosis and significantly extends disease latency in vivo tying the reduced survival mediated by JUN inhibition to the loss of pro-survival UPR signaling. These data uncover a previously unrecognized role of JUN as a regulator of the UPR as well as provide key new insights into the how ER stress responses contribute to AML and identify JUN and the UPR as promising therapeutic targets in this disease.

Reed-Sternberg cells in Hodgkin's lymphoma present features of cellular senescence.

Hodgkin's Lymphoma (HL) is one of the most prevailing malignancies in young adults. Reed-Sternberg (RS) cells in HL have distinctive large cell morphology, are characteristic of the disease and their presence is essential for diagnosis. Enlarged cells are one of the hallmarks of senescence, but whether RS cells are senescent has not been previously investigated. Here we show that RS cells have characteristics of senescent cells; RS cells in HL biopsies specifically express the senescence markers and cell cycle inhibitors p21Cip1 and p16INK4a and are negative for the proliferation marker Ki-67, suggesting that these cells have ceased to proliferate. Moreover, the RS-like cells in HL lines, stained specifically for senescence-associated ß-galactosidase (SA-ß-gal). Oxidative stress promoted senescence in these cells as demonstrated by their staining for p21Cip1, p16INK4a, p53 and γH2AX. Senescent cells produce copious amounts of inflammatory cytokines termed 'senescence-associated secretory phenotype' (SASP), primarily regulated by Nuclear Factor κB (NF-κB). Indeed, we show that NF-κB activity and NF-κB-dependent cytokines production (e.g., IL-6, TNF-α, GM-CSF) were elevated in RS-like cells. Furthermore, NF-κB inhibitors, JSH-23 and curcumin reduced IL-6 secretion from RS-like cells. Thus, defining RS cells as senescent offers new insights on the origin of the proinflammatory microenvironment in HL.

The complete genome sequence of a south Indian isolate of Rice tungro spherical virus reveals evidence of genetic recombination between distinct isolates.

In this study, complete genome of a south Indian isolate of Rice tungro spherical virus (RTSV) from Andhra Pradesh (AP) was sequenced, and the predicted amino acid sequence was analysed. The RTSV RNA genome consists of 12,171 nt without the poly(A) tail, encoding a putative typical polyprotein of 3,470 amino acids. Furthermore, cleavage sites and sequence motifs of the polyprotein were predicted. Multiple alignment with other RTSV isolates showed a nucleotide sequence identity of 95% to east Indian isolates and 90% to Philippines isolates. A phylogenetic tree based on complete genome sequence showed that Indian isolates clustered together, while Vt6 and PhilA isolates of Philippines formed two separate clusters. Twelve recombination events were detected in RNA genome of RTSV using the Recombination Detection Program version 3. Recombination analysis suggested significant role of 5' end and central region of genome in virus evolution. Further, AP and Odisha isolates appeared as important RTSV isolates involved in diversification of this virus in India through recombination phenomenon. The new addition of complete genome of first south Indian isolate provided an opportunity to establish the molecular evolution of RTSV through recombination analysis and phylogenetic relationship.

Uncorrectable coagulopathy due to intestinal obstruction: A clinical dilemma of competing priorities in a child.

Disseminated intravascular coagulation (DIC) is a contraindication for major surgical operations. However, if surgery is required to correct the underlying cause of DIC, a clinical conflict is created. In such complex scenario, standard resources such as textbooks and journals offer very little guidance. In this communication, we report a 22-month-old boy who developed sepsis-induced DIC following intestinal obstruction. Pre-operative attempts to normalize coagulation parameters failed. Damage control laparotomy was undertaken as it was considered essential to control the underlying cause of DIC. His abnormal coagulation status reverted quickly after surgical relief of intestinal obstruction. Paradoxically intraoperative blood loss was less than anticipated amount. There are a few case reports of adult patients who have successfully undergone major surgery despite the presence of abnormal coagulation. However, this appears to be the first paediatric report of successful surgery in DIC status. Lessons learnt from this case and hypothetical speculations of clinical paradoxes are discussed.

Phytochemicals as chemosensitizers: from molecular mechanism to clinical significance.

This review provides an overview of the clinical relevance of chemosensitization, giving special reference to the phenolic phytochemicals, curcumin, genistein, epigallocatechin gallate, quercetin, emodin, and resveratrol, which are potential candidates due to their ability to regulate multiple survival pathways without inducing toxicity. We also give a brief summary of all the clinical trials related to the important phytochemicals that emerge as chemosensitizers. The mode of action of these phytochemicals in regulating the key players of the death receptor pathway and multidrug resistance proteins is also abridged. Rigorous efforts in identifying novel chemosensitizers and unraveling their molecular mechanism have resulted in some of the promising candidates such as curcumin, genistein, and polyphenon E, which have gone into clinical trials. Even though considerable research has been conducted in identifying the salient molecular players either contributing to drug efflux or inhibiting DNA repair and apoptosis, both of which ultimately lead to the development of chemoresistance, the interdependence of the molecular pathways leading to chemoresistance is still the impeding factor in the success of chemotherapy. Even though clinical trials are going on to evaluate the chemosensitizing efficacy of phytochemicals such as curcumin, genistein, and polyphenon E, recent results indicate that more intense study is required to confirm their clinical efficacy. Current reports also warrant intense investigation about the use of more phytochemicals such as quercetin, emodin, and resveratrol as chemosensitizers, as all of them have been shown to modulate one or more of the key regulators of chemoresistance.

Nicotine-induced survival signaling in lung cancer cells is dependent on their p53 status while its down-regulation by curcumin is independent.

BACKGROUND: Lung cancer is the most lethal cancer and almost 90% of lung cancer is due to cigarette smoking. Even though nicotine, one of the major ingredients of cigarette smoke and the causative agent for addiction, is not a carcinogen by itself, several investigators have shown that nicotine can induce cell proliferation and angiogenesis. We observed that the proliferative index of nicotine is different in the lung cancer cell lines H1299 (p53-/-) and A549 (p53+/+) which indicates that the mode of up-regulation of survival signals by nicotine might be different in cells with and without p53. RESULTS: While low concentrations of nicotine induced activation of NF-κB, Akt, Bcl2, MAPKs, AP1 and IAPs in H1299, it failed to induce NF-κB in A549, and compared to H1299, almost 100 times higher concentration of nicotine was required to induce all other survival signals in A549. Transfection of WT-p53 and DN-p53 in H1299 and A549 respectively, reversed the mode of activation of survival signals. Curcumin down-regulated all the survival signals induced by nicotine in both the cells, irrespective of their p53 status. The hypothesis was confirmed when lower concentrations of nicotine induced NF-κB in two more lung cancer cells, Hop-92 and NCI-H522 with mutant p53 status. Silencing of p53 in A549 using siRNA made the cells susceptible to nicotine-induced NF-κB nuclear translocation as in A549 DN-p53 cells. CONCLUSIONS: The present study reveals a detrimental role of nicotine especially in lung cancer patients with impaired p53 status and identifies curcumin as a potential chemopreventive.

Temporal variability of benzene concentration in the ambient air of Delhi: a comparative assessment of pre- and post-CNG periods.

CNG (compressed natural gas) was fully implemented in public transport system in Delhi in December 2002. The study assesses the benzene concentration trends at two busy traffic intersections and a background site in Delhi, India. Monitoring was done for two different time periods viz; in the year 2001-2002 (pre-CNG) and two winter months (January and February) of the year 2007 (post-CNG) to assess the impact of various policy measures adopted by the government of Delhi to improve the air quality in the city. Annual average benzene concentration for the pre-CNG period was found to be 86.47+/-53.24 microg m(-3). Average benzene concentrations for the winter months (January-February) of pre- and post-CNG periods were 116.32+/-51.65 microg m(-3) and 187.49+/-22.50 microg m(-3), respectively. Enhanced values could be solely attributed to the increase in the vehicular population from 3.5 million in the year 2001-2002 to approximately 5.1 millions in the year 2007.

Histamine release during adult cardiopulmonary bypass.

Histamine, an inflammatory mediator in its own right, may also be a marker for a more widespread systemic inflammatory process. In this study we have examined variations in plasma histamine concentrations produced during the course of cardiac surgery involving cardiopulmonary bypass, the relationship between these variations and intra-operative events. By assays of serum tryptase and CD-63 expression we have also attempted to identify the source of histamine. Histamine concentrations that were significantly raised from baseline level were demonstrated. These were elevated from the time of aortic cross-clamping and continued to be raised for 24 h postoperatively (p < 0.00625). This was associated with an increase in CD-63 expression (p < 0.025) (but not an increase in tryptase concentration) following aortic cross-clamping and protamine administration, suggesting that basophils are the source of histamine. 41% of patients had arrhythmias in the post bypass period. The rise in histamine levels was not related to the incidence of cardiac arrhythmias.

Scoliosis in cystic fibrosis: is it idiopathic?

STUDY DESIGN: This is a retrospective study of all the patients registered with the Yorkshire Regional Cystic Fibrosis unit from 1982 to 1997. Of the 316 patients, there were 184 adults (age 17 years and above) and 132 children (age 0-6 years). OBJECTIVES: This study was aimed at determining the prevalence of scoliosis in people with cystic fibrosis and describes the characteristics and progression of scoliosis in these patients and highlights predictive factors, which account for high prevalence of scoliosis in this condition. SUMMARY OF THE BACKGROUND DATA: Two previous North American studies (1978 and 1982) have indicated a high prevalence of scoliosis in patients with cystic fibrosis. METHODS: The patients were divided into 3 groups based on their chronologic age as on January 1998. Chest, abdomen, and whole spine radiographs were studied for the presence of any spinal deformity, and measurements were made using the Oxford Cobbmeter (Oxford Metrics). The extent, apex, and the side of the curves were described. The disease-specific scores, ie, Chrispin-Norman score (score for radiologic severity of lung disease) and Shwachman score (score for general condition), were noted from the patient follow-up database maintained by the Regional Cystic Fibrosis Unit. Multiple linear regression analysis was used to study the correlation between Cobb angle and the previously mentioned scores. RESULTS: In the 4- to 16-year age group, the prevalence of scoliosis was 15.6%, which is 20 times the prevalence in 15,793 school children with a similar age and sex distribution from the same geographic area. The majority of curves were single-thoracic, apical around T6-T8 with no side predilection. In the adult population (above 16 years), the prevalence was 9.8%, which is higher than that of the general population. These curves were thoracic, apical around T7-T8, and approximately two thirds of them were right-sided. Infantile curves are described for the first time in our study; these tend to be nonprogressive, right-sided, upper thoracic curves. CONCLUSION: Our study shows a high prevalence of scoliosis in people with cystic fibrosis. These are benign short midthoracic curves, apical between T6-T8 with no side predilection.

Oncolytic activity of vesicular stomatitis virus is effective against tumors exhibiting aberrant p53, Ras, or myc function and involves the induction of apoptosis.

We have recently shown that vesicular stomatitis virus (VSV) exhibits potent oncolytic activity both in vitro and in vivo (S. Balachandran and G. N. Barber, IUBMB Life 50:135-138, 2000). In this study, we further demonstrated, in vivo, the efficacy of VSV antitumor action by showing that tumors that are defective in p53 function or transformed with myc or activated ras are also susceptible to viral cytolysis. The mechanism of viral oncolytic activity involved the induction of multiple caspase-dependent apoptotic pathways was effective in the absence of any significant cytotoxic T-lymphocyte response, and occurred despite normal PKR activity and eIF2alpha phosphorylation. In addition, VSV caused significant inhibition of tumor growth when administered intravenously in immunocompetent hosts. Our data indicate that VSV shows significant promise as an effective oncolytic agent against a wide variety of malignant diseases that harbor a diversity of genetic defects.

Analyzing the mechanisms of interferon-induced apoptosis using CrmA and hepatitis C virus NS5A.

The dsRNA-dependent protein kinase, PKR, is a key component of interferon (IFN)-mediated anti-viral action and is frequently inhibited by many viruses following infection of the cell. Recently, we have demonstrated that IFN and PKR can sensitize cells to apoptosis predominantly through the FADD/caspase-8 pathway (S. Balachandran, P. C. Roberts, T. Kipperman, K. N. Bhalla, R. W. Compans, D. R. Archer, and G. N. Barber. (2000b) J. Virol. 74, 1513-1523). Given these findings, it is thus plausible that rather than specifically target IFN-inducible genes such as PKR, viruses could also subvert the mechanisms of IFN action, in part, at locations that could block the apoptotic cascade. To explore this possibility, we analyzed whether the poxvirus caspase-8 inhibitor, CrmA, was able to inhibit IFN or PKR/dsRNA-mediated apoptosis. Our findings indicated that CrmA could indeed inhibit apoptosis induced by both viral infection and dsRNA without blocking PKR activity or inhibiting IFN signaling. In contrast HCV-encoded NS5A, a putative inhibitor of PKR, did not appear to inhibit cell death mediated by a number of apoptotic stimuli, including IFN, TRAIL, and etoposide. Our data imply that viral-encoded inhibitors of apoptosis, such as CrmA, can block the innate arms of the immune response, including IFN-mediated apoptosis, and therefore potentially constitute an alternative family of inhibitors of IFN action in the cell.

Vesicular stomatitis virus (VSV) therapy of tumors.

Vesicular stomatitis virus (VSV) is an essentially nonpathogenic negative-stranded RNA virus, the replication of which is extremely sensitive to the antiviral effects of interferon (IFN). We demonstrate here that VSV selectively induces the cytolysis of numerous transformed human cell lines in vitro, with all the morphological characteristics of apoptotic cell death. Importantly, VSV can also potently inhibit the growth of p53-null C6 glioblastoma tumors in vivo without infecting and replicating in normal tissue. With our previous findings demonstrating that primary cells containing the double-stranded RNA-activated protein kinase PKR and a functional IFN system are not permissive to VSV replication, these results suggest that signaling by IFN may be defective in many malignancies. Thus VSV might be useful in novel therapeutic strategies for targeting neoplastic disease.

Activation of the dsRNA-dependent protein kinase, PKR, induces apoptosis through FADD-mediated death signaling.

The dsRNA-dependent protein kinase (PKR) is considered to play a key role in interferon-mediated host defense against viral infection and conceivably malignant transformation. To investigate further the mechanisms of PKR-induced growth inhibition, we have developed tetracycline-inducible murine cell lines that express wild-type PKR or a catalytically inactive PKR variant, PKRdelta6. Following induction, the growth of the wild-type PKR-expressing cells was similar to that of cells transfected with vector alone, while cells expressing PKRdelta6 became malignantly transformed. Significantly, treatment with dsRNA caused the wild-type PKR-overexpressing cells to undergo programed cell death while, conversely, cells expressing PKRdelta6 were completely resistant. Our studies demonstrated that activation of PKR induces the expression of members of the tumor necrosis factor receptor (TNFR) family, including Fas (CD95/Apo-1) and pro-apopotic Bax. In contrast, transcripts representing Fas, TNFR-1, FADD (Fas-associated death domain), FLICE, Bad and Bax were ablated in cells expressing PKRdelta6. The involvement of the death receptors in PKR-induced apoptosis was underscored by demonstrating that murine fibroblasts lacking FADD were almost completely resistant to dsRNA-mediated cell death. Thus, PKR, a key cellular target for viral repression, is a receptor/inducer for the induction of pro-apoptotic genes by dsRNA and probably functions in interferon-mediated host defense to trigger cell death in response to virus infection and perhaps tumorigenesis.