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Three universal types of cancer are identified, based on the malfunction of a certain set of proteins and regulatory RNAs, irrespective of the organ in which they are located: Cancer Type 1, where cancer cells lack either a functional (a) P14ARF gene, or (b) a P53 gene; Cancer Type 2, where cancer cells lack a functional DINO lncRNA; and Cancer Type 3, where cancer cells have abnormally high MDM2 protein activity. New therapeutic targets were discovered for each type of cancer that pave the way for treating cancer irrespective of the organ it lies in. Until now, current cancer treatments have been organ-specific, and no common pan-organ denominator has been identified. Furthermore, cancer biochemistry has been studied in isolation, one pathway at a time, without considering the complex interactions between proteins and regulatory RNAs, which are characteristic of a living human organism. This work develops a new unified therapeutic theory that identifies novel master regulators of apoptosis as targets for treating cancer regardless of which organ the cancer lies in, through a novel biochemical flowsheet of a universal apoptosis network comprising approximately 100 pathways (80% activation and 20% inhibition), based on a critical analysis of 172 scientific publications that considered all the complex interactions between proteins and regulatory RNAs.

Mouse mammary tumour virus-like (MMTV-like) is suspected to be involved in human breast cancer and it has been hypothesized that companion animals might have a role in viral transmission. The aim of our study was to investigate the presence of MMTV-like nucleotide sequences and viral protein in a larger number of feline (FMCs) and canine mammary carcinomas (CMCs) by nested PCR and immunohistochemistry. Results showed that the presence of MMTV-like env sequence in FMCs was 7% (6/86), while all the CMCs and canine dysplastic lesions scored negative. All PCR-positive FMCs scored positive for the MMTV p14 signal peptide of the envelope precursor protein of the virus. In contrast, all PCR-negative FMCs and canine mammary lesions were also negative for immunohistochemistry analysis. Canine and feline normal mammary gland tissues scored negative for both PCR and MMTV-p14 protein. Multiple nucleotide alignment of MMTV-like env gene sequences isolated from cat showed 97% and 99% similarity with HMTV and MMTV, respectively, while the others two presented some polimorphisms. Particularly the sequences of one of these two tumors showed a polymorphism (c.7575 A> G), that causes a previously unreported amino acid substitution (Thr > Ala). In conclusion, the results of our study showed the presence of MMTV-like sequences and viral protein in some FMCs. Further studies are needed to understand whether this virus does play a role in the development of FMCs, if MMTV-like is an exogenous virus as these data suggest and, in such a case, how and from whom this virus was acquired.

The physical entry of virus particles into cells triggers an innate immune response that is dependent on both calcium and nucleic acid sensors, with particles containing RNA or DNA genomes detected by RNA or DNA sensors, respectively. While membrane fusion in the absence of viral nucleic acid causes an innate immune response that is dependent on calcium, the involvement of nucleic acid sensors is poorly understood. Here, we used lipoplexes containing purified reovirus p14 fusion protein as a model of exogenous or fusion from without and a cell line expressing inducible p14 protein as a model of endogenous or fusion from within to examine cellular membrane fusion sensing events. We show that the cellular response to membrane fusion in both models is dependent on calcium, IRF3 and IFN. The method of sensing fusion, however, differs between fusion from without and fusion from within. Exogenous p14 lipoplexes are detected by RIG-I-like RNA sensors, whereas fusion by endogenous p14 requires both RIG-I and STING to trigger an IFN response. The source of nucleic acid that is sensed appears to be cellular in origin. Future studies will investigate the source of endogenous nucleic acids recognized following membrane fusion events.

Background: Chimeric antigen receptor (CAR)-based immune cell therapies attack neighboring cancer cells after receptor recognition but are unable to directly affect distant tumor cells. This limitation may contribute to their inefficiency in treating solid tumors, given the restricted intratumoral infiltration and immunosuppressive tumor microenvironment. Therefore, cell-cell fusion as a cell-killing mechanism might develop a novel cytotherapy aimed at improving the efficacy against solid tumors. Methods: We constructed a fusogenic protein, fusion-associated small transmembrane (FAST) p14 of reptilian reovirus, into cancer cells and mesenchymal stem cells (MSCs), which cocultured with various colon cancer cells and melenoma cells to validate its ability to induce cell fusion and syncytia formation. RNA sequencing, quantitative reverse transcription polymerase chain reaction, and Western blot were performed to elucidate the mechanism of syncytia death. Cell viability assay was employed to assess the killing effects of MSCs carrying the p14 protein (MSCs-p14), which was also identified in the subcutaneous tumor models. Subsequently, the Tet-On system was introduced to enhance the controllability and safety of therapy. Results: Cancer cells incorporated with fusogenic protein p14 FAST from reovirus fused together to form syncytia and subsequently died through apoptosis and pyroptosis. MSCs-p14 cocultured with different cancer cells and effienctly induced cancer cell fusion and caused widespread cancer cell death in vitro. In mouse tumor models, mMSCs-p14 treatment markedly suppressed tumor growth and also enhanced the activity of natural killer cells and macrophages. Controllability and safety of MSCs-p14 therapy were further improved by introducing the tetracycline-controlled transcriptional system. Conclusion: MSC-based cytotherapy carrying viral fusogenic protein in this study kills cancer cells by inducing cell-cell fusion. It has demonstrated definite efficacy in treating solid tumors and is worth considering for clinical development.

Trafficking of integral membrane proteins between the ER and Golgi complex, and protein sorting and trafficking between the TGN and endosomal/lysosomal compartments or plasma membranes, are dependent on cis-acting, linear amino acid sorting signals. Numerous sorting signals of this type have been identified in the cytoplasmic domains of membrane proteins, several of which rely on basic residues. A novel Golgi export signal that relies on a membrane-proximal polybasic motif (PBM) was recently identified in the reptilian reovirus p14 protein, a representative of an unusual group of bitopic fusion-associated small transmembrane (FAST) proteins encoded by fusogenic orthoreoviruses and responsible for cell-cell fusion and syncytium formation. Using immunofluorescence microscopy, cell surface immunofluorescence, and endoglycosidase H assays, we now show the p14 PBM can mediate several distinct trafficking functions depending on its proximity to the transmembrane domain (TMD). When present within 4-residues of the TMD it serves as a Golgi export signal, but when located at the C-terminus of the 68-residue p14 cytoplasmic endodomain it functions as an ER retention signal. The PBM has no effect on protein trafficking when located at an internal position in the cytoplasmic domain. When present in both membrane-proximal and -distal locations, the PBMs promote export to, and efficient retrieval from, the Golgi complex. Interestingly, the conflicting trafficking signals provided by two PBMs induces extensive ER tubulation and segregation of ER components. These studies highlight how a single trafficking signal in a simple transmembrane protein can have remarkably diverse, position-dependent effects on protein trafficking and ER morphogenesis.

Genes of the polycomb group function by silencing homeotic selector genes that regulate embryogenesis. In mice, downregulation of one of the polycomb genes, bmi-1, leads to neurological alterations and severe proliferative defects in lymphoid cells, whilst bmi-1 overexpression, together with upregulation of myc-1, induces lymphoma. An oncogenic function has been further supported in primary fibroblast studies where bmi-1 overexpression induces immortalization due to repression of p16/p19ARF, and where together with H-ras, it readily transforms MEFs. It was the aim of this study to assess the expression of bmi-1 in resectable non-small cell lung cancer (NSCLC) in association with p16 and p14ARF (=human p19ARF). Tumours (48 resectable NSCLC (32 squamous, 9 adeno-, 2 large cell, 4 undifferentiated carcinomas and 1 carcinoid); stage I, 29, II, 7, III, 12; T1, 18, T2, 30; differentiation: G1 12, G2 19, G3 17) were studied by immunohistochemistry for protein expression and by comparative multiplex PCR for gene amplification analysis. In tumour-free, normal lung tissue from patients, weak – moderate bmi-1 staining was seen in some epithelial cells, lymphocytes, glandular cells and in fibroblasts, whereas blood, endothelial, chondrocytes, muscle cells and adipocytes did not exhibit any bmi-1 expression. In tumours, malignant cells were negative/weakly, moderately and strongly positive in 20, 22 and 6 cases, respectively. As assessed by multiplex PCR, bmi-1 gene amplification was not the reason for high-level bmi-1 expression. Tumours with moderate or strong bmi-1 expression were more likely to have low levels of p16 and p14ARF ( P = 0.02). Similarly, tumours negative for both, p16 and p14ARF, exhibit moderate–strong bmi-1 staining. 58% of resectable NSCLC exhibit moderate–high levels of bmi-1 protein. The inverse correlation of bmi-1 and the INK4 locus proteins expression (p16/p14ARF) supports a possible role for bmi-1 misregulation in lung carcinogenesis. http://www.bjcancer.com © 2001 Cancer Research Campaign

We conducted a meta-analysis to explore the relationships between p14ARF gene methylation and clinicopathological features of colorectal cancer (CRC). Databases, including Pubmed, Embase and Cochrane Library, were searched and, finally, a total of 18 eligible researches encompassing 1988 CRC patients were selected. Combined odds ratios (ORs) with 95% confidence intervals (95% CIs) were evaluated under a fixed effects model for absence of heterogeneity. Significant associations were observed between p14ARF gene methylation and tumor location (OR = 2.35, 95% CI: 1.55-3.55, P = 0.001), microsatellite instability (MSI) status (OR = 3.28, 95% CI: 2.12-5.07, P<0.0001). However, there were no significant associations between p14ARF gene methylation and tumor stage, tumor differentiation. We concluded that p14ARF gene methylation may be significantly associated with tumor location, and MSI status of CRC.

Atopic dermatitis (AD) is a chronic inflammatory skin disease, with a complex etiology encompassing immunologic responses. AD is frequently associated with elevated serum immunoglobulin (Ig) E levels and is exacerbated by a variety of environmental factors, which contribute to its pathogenesis. However, the etiology of AD remains unknown. Recently, reports have documented the role of lactic acid bacteria (LAB) in the treatment and prevention of AD in humans and mice. The LAB, Lactobacillus casei (LC), is frequently used in the treatment of AD. To identify the active component of LC, we screened fractions obtained from the ion exchange chromatography of LC extracts. Using this approach, we identified the candidate protein, P14. We examined whether the P14 protein has anti-atopic properties, using both in vitro and in vivo models. Our results showed that the P14 protein selectively downregulated serum IgE and interleukin-4 cytokine levels, as well as the AD index and scratching score in AD-like NC/Nga mice. In addition, histological examination was also effective in mice. These results suggest that the P14 protein has potential therapeutic effects and that it may also serve as an effective immunomodulatory agent for treating patients with AD.

Undifferentiated nasopharyngeal carcinoma (NPC) is an epithelial malignancy that is consistently associated with Epstein-Barr virus (EBV) but which very rarely has p53 gene mutations in primary tumours. Since the tumour suppressor p53 is mutated in most human cancers or the wild type protein is inactivated in a significant number of the remainder, here we have investigated cellular factors that could compromise p53 function in primary NPC. Twenty-five primary tumours were judged to carry only wild type p53 by SSCP analysis of all exons and sequence determination of exons 4–9. Only one tumour was found to express significant levels of hMdm2 and in 24/25 there were no detectable mutations or deletions in exons 1β and 2 of the p14ARF gene. However, immunohistochemistry consistently revealed that all the tumour cells express substantial amounts of the p53-related protein p63. Semi-quantitative RT–PCR analysis of mRNA from tumour biopsies showed that the dominant species expressed was invariably the truncated ΔN-isotype. Since this can block p53-mediated transactivation, it is potentially a dominant-negative isoform. In normal nasopharyngeal epithelium the distribution of p63 was restricted to the proliferating basal and suprabasal layers. We suggest that ΔN-p63 is a good candidate as a suppressor of wild type p53 function in these tumours and also that it may prove to be a valuable diagnostic marker for undifferentiated NPC.

The INK4a/ARF locus encodes the cyclin dependent kinase inhibitor, p16INK4a and the p53 activator, p14ARF. These two proteins have an independent first exon (exon 1α and exon 1β, respectively) but share exons 2 and 3 and are translated in different reading frames. Germline mutations in this locus are associated with melanoma susceptibility in 20–40% of multiple case melanoma families. Although most of these mutations specifically inactivate p16INK4a, more than 40% of the INK4a/ARF alterations located in exon 2, affect both p16INK4a and p14ARF. We now report a 16 base pair exon 1β germline insertion specifically altering p14ARF, but not p16INK4a, in an individual with multiple primary melanomas. This mutant p14ARF, 60ins16, was restricted to the cytoplasm, did not stabilize p53 and was unable to arrest the growth of a p53 expressing melanoma cell line. This is the first example of an exon 1β mutation that inactivates p14ARF, and thus implicates a role for this tumour suppressor in melanoma predisposition.