Thomas Schmitt

Staphylococcus aureus (S. aureus) is an important class of food-borne pathogens that cause food contamination and food poisoning, and is also a symbiont of human skin and mucous membranes. Studies have found that about 20%~30 % of healthy individuals carry Staphylococcus aureus. Animal foods are susceptible to Staphylococcus aureus infection, such as milk, butter, cured ham, etc. Food poisoning incidents caused by Staphylococcus aureus have been frequently reported, and more than 95% of them are caused by enterotoxin. According to reports, the United States and Canada accounted for 33% and 45% of food poisoning incidents caused by staphylococcal enterotoxins (SEs), respectively. Epidemiological studies have shown that Staphylococcus aureus enterotoxin A can be produced under more suitable environmental conditions (temperature 7~47, pH 4℃~10) and growth phase (logarithmic growth phase or transition to stable phase) Staphylococcal enterotoxin A(SEA), Staphylococcal enterotoxin B (SEB), Staphylococcal enterotoxin C1 (SEC1), Staphylococcal enterotoxin D (SED) and other about 23 serotypes, they are similar in structure and function with molecular weight of 27.5~30 kDa. Among the above-mentioned SEs, SEB is one of the most common biotoxins reported so far, and it is also a typical biotoxin that causes food poisoning. In order to detect SEB, researchers have developed different detection methods, among which the main detection methods are biological methods, immunological methods, gene probe methods, instrumental analysis methods and biosensing methods. Figure 1. New insights into Staphylococcus aureus immune evasion. Structural Properties and Hazards of SEB SEB is mostly found in spoiled milk and meat products in food. According to relevant research reports, the content of SEB in the culture supernatant of Staphylococcus aureus (strain S-6) is as high as 0.2 mg/mL, and the semi-lethal dose LD50 of mice is 0.02 µg/kg. The molecular weight of SEB is 28.4 KDa, which contains 239 amino acid fragments. The structure of SEB is mainly composed of 2 regions, of which 129 amino acid residues in region 1 contain 2 β-sheets and 3 α-helices (α1, α2 and α3). The 2 β-sheets form a cylindrical structure, the inner wall is lined with hydrophilic residues, and they form “crosses” with each other. Region 2 is divided into 2 different parts, one part contains 2 α-helices (α4 and α5) and 2 very short β-strands, and the other part has α-helices located between the curved sheet and β cylinder. One side of α4 and α5 is exposed to the solvent, and the two domains are connected by a ring structure. Compared with other enterotoxins, SEB has stronger heat resistance, its structure and function do not change significantly after being heated at 100 ℃ for 30 min, and it can resist the hydrolysis of intestinal proteases. Therefore, enterotoxin-contaminated food can still cause potential toxic effects even after being cooked and digested by the human body. As a superantigen substance, enterotoxin B simultaneously forms a bridge by cross-linking with major histocompatibility complex (MHC) class II molecules on antigen-presenting cells (APCs), Stimulates T cells in a manner that does not involve antigen recognition by generating receptor complexes with MHC II sites on APCs and T cells. It can effectively bypass the treatment of conventional APCs, induce T cells to produce cytokines, and cause the activation and proliferation of T cell subsets. Related studies have further confirmed that SEB increases the expression of components of the arachidonic acid pathway, leading to inflammation, edema and shock. Current studies have found that ingesting a small amount of SEB in the human body can cause immune disorders, organ damage and other symptoms, and the clinical manifestations are fever, respiratory diseases (cough, dyspnea and chest pain) and digestive tract diseases. Therefore, it is of great significance to study the highly sensitive and rapid detection method of SEB for the clinical diagnosis, prevention and control of food poisoning. Method for the Detection of Staphylococcus aureus Enterotoxin B Biological Detection Method The detection method of Staphylococcus aureus enterotoxin B was initially based on related detection methods such as zoological experiments. Through intraperitoneal injection and feeding of samples added with enterotoxin B to the tested animals, the abnormal physiological reactions and activities of the animals were observed and recorded. The existence and toxicity of enterotoxin B are qualitatively analyzed. This type of experiment is intuitive in the interpretation of the results, but the disadvantages such as poor specificity limit the application of this method. Immunoassay Immunological detection is mainly based on the principle of specific combination of antigen and antibody, including immunoagglutination test, agarose diffusion method and enzyme-linked immunosorbent assay. However, the detection sensitivity of these methods is only 0.1 ng/mL, and the sample preparation and detection take a long time. In addition, this method needs to ensure the homogeneity of monoclonal antibody structure and avoid cross-reaction between each other. Immunoagglutination Test The immunoagglutination test is the agglutination produced by the combination of antigen and antibody in the presence of electrolytes. These include the reverse indirect hemagglutination test and the reverse passive latex agglutination test. The former uses the antiserum of enterotoxin B to adsorb on the surface of red blood cells of animals. When the test specimen contains SEB, hemagglutination occurs. This method can interpret the results in about 2 hours, and the detection sensitivity is greatly improved compared with the previous method; the latter adsorbs the specific anti-SEB antibody on the latex particles, and when the electrolyte and SEB exist at the same time, the latex particles agglutinate phenomenon, the detection time is about 18 h. 2. Agarose Diffusion Method Immuno-agar diffusion method refers to the diffusion of soluble antigens and antibodies in semi-solid agar. It is not suitable for the detection of low concentration SEB (below ng level). If the antigen and antibody react according to a certain concentration relationship, a white precipitate line appears on the surface of the plate, which is a positive result, and the standard curve is made by the logarithm of the distance of the SEB diffusion agarose layer and the antigen concentration, which is used for the indirect quantification of SEB . The method is simple to operate and does not require any equipment, but the disadvantages of long detection time and low sensitivity limit the application of this method. 3. Immunolabeling Technique Immunolabeling techniques can be divided into detection techniques such as enzyme-linked immunosorbent immunoassay, radioimmunoassay, and immunofluorescence. It is based on the traditional antigen-antibody reaction, by modifying fluorescent groups on antibodies such as hydroxyfluorescein (financial analyst meeting, FAM), Fluoresceine isothiocyanate (FITC), water-soluble indocyanine-type fluorescein (cyanine-5, Cy5), etc., radioactive isotopes, horseradish peroxidase (horseradish peroxidase, HRP) or alkaline phosphatase (alkaline phosphatase, AP) and other trace substances, using relevant equipment to output signal intensity to achieve the purpose of detection. Enzyme-linked Immunoassay Technology Enzyme linked immunosorbent assay (ELISA) is based on the antigen-antibody reaction of a solid-phase carrier (such as polystyrene, etc.), and the color development of the substrate is catalyzed by HRP or AP, thereby amplifying the detection signal. Double-antibody sandwich and indirect competition methods are usually used to detect SEB, and the former can obtain lower detection sensitivity and wider linear range. At present, enzyme-linked immunoassay technology has been widely used in actual detection, and a targeted detection kit has been developed. Radioimmunoassay Technology Radioimmunoassay is based on the competitive reaction of labeled antigen (labeled with radioactive isotope) and non-labeled antigen to antibody, and is used to detect the antigen concentration of the sample to be tested. Immunofluorescence Detection Technique Immunofluorescence detection technology uses fluorescent substances (FITC, Cy5, quantum dots, upconversion particles, etc.) as signal molecules labeled on antibodies, and the concentration of SEB is indirectly measured by the fluorescence intensity of the labeled antibody bound to SEB, usually using quantitative determination by photometer. Colloidal Gold Test Strip Detection Technology Colloidal gold-labeled immune test strip mainly utilizes the excellent physical and chemical properties of colloidal gold. Since the surface of colloidal gold has a negative charge, under alkaline conditions, the antibody can be immobilized on the surface by physical adsorption without affecting the biological activity of the antibody itself. This method can quickly interpret the results under naked eye conditions, and is generally used for preliminary screening on the spot. Gene Probe Method Enterotoxin B is transcribed and translated from the toxin expression gene sequence seb of Staphylococcus aureus. By detecting the enterotoxin B gene in Staphylococcus aureus in food, it can be used for rapid and high-throughput detection of positive strains of Staphylococcus aureus. Specifically, the method mainly uses nucleic acid amplification techniques, including polymerase chain amplification and nucleic acid isothermal amplification techniques. Instrumental Analysis Method With the continuous improvement of the current detection instruments and equipment, many scholars are no longer limited to the traditional immunology and other related methods, but instead use modern large-scale precision instruments for qualitative and quantitative detection of enterotoxin B. Some instrumental analysis methods can directly reveal the molecular structure of toxins. Biosensing Detection Method Biosensor (biosensor) connects biochemical reactions with transducers through physical and chemical instruments, and expresses them in the form of electricity. It mainly recognizes biological components of specific substances (such as enzymes, antibodies, nucleic acids, lectins, Cells or tissues) and biological reaction signal amplification are organically combined, which has the advantages of less sample usage and fast analysis speed, and can be used for simultaneous detection of multiple substances. At present, the biosensors used for detection mainly include electrochemical sensors, surface plasmon resonance and piezoelectric crystal sensors.

The incidence and prevalence of diabetes are increasing year by year globally, and patients with diabetes are at increased risk of cardiovascular disease and heart failure. Diabetic cardiomyopathy (DCM) refers to the presence of abnormal myocardial structure and performance in the absence of other heart-related risk factors (such as coronary artery-related disease, hypertension, and severe valvular disease). DCM can lead to cardiac systolic or diastolic dysfunction, further causing heart failure with reduced ejection fraction and heart failure with normal ejection fraction, seriously affecting the patient’s quality of life. The pathogenesis of DCM is particularly complex. Current research has discovered multiple mechanisms of action, including metabolic disorders, insulin resistance, formation of advanced glycation end products, mitochondrial damage, oxidative stress, endoplasmic reticulum stress (ERS), Inflammatory response and cell apoptosis, etc. Glucagon-like peptide-1 (GLP-1) receptor agonist is an incretin secreted by intestinal L cells. It reduces the blood sugarof diabetes in patients with diabetes by increasing glucose-dependent insulin secretion and inhibiting glucagon secretion, and also regulates a variety of cardiovascular risk factors, thereby inhibiting the process of atherosclerosis and reducing the occurrence of coronary events and cardiovascular death. GLP-1 receptor agonists have a longer half-life, are more effective than many oral hypoglycemic drugs in clinical trials, and have a low risk of hypoglycemia. This article reviews the effects of GLP-1 receptor agonists on DCM to provide reference value for further research on GLP-1 receptor agonists. Figure 1. Major cardiometabolic actions of GLP1. GLP-1 Receptor Agonists Affect DCM by Regulating Metabolic Disorders Hyperglycemia and insulin resistance are the most significant features of DCM. In addition, they are also accompanied by changes in myocardial structure and function caused by calcium imbalance, abnormal lipid metabolism, lipid deposition, and the combined effects of multiple hormone changes. The metabolic disorder of myocardial cells is mainly caused by the increase in blood sugar levels caused by insulin resistance. When diabetes occurs, there are conversion obstacles in both the anaerobic glycolysis and aerobic oxidation of glucose in the body. The body’s utilization of glucose is reduced, lipolysis is increased, and excessive free Fatty acids are absorbed by cardiomyocytes, reducing the heart’s ability to use glucose. At this time, the heart mainly relies on fatty acid oxidation to provide energy, but high oxygen consumption is accompanied by low energy metabolism efficiency. The accumulation of free fatty acids in the form of triglycerides promotes fatty degeneration of the myocardium. Therefore, long-term hyperglycemic environment, abnormal lipid metabolism, and lipid deposition lead to the occurrence and development of DCM in diabetic patients. Multiple studies have shown that GLP-1 receptor agonists can directly or indirectly improve the function of cardiomyocytes by controlling the above multiple metabolic factors and related signaling pathways. ​GLP-1 Receptor Agonists Affect DCM by Improving ERS In diabetic patients, the endoplasmic reticulum dysfunction caused by cardiac oxidative stress, abnormal lipid metabolism and inflammatory response is called ERS when the body is in a high glucose state. Pathophysiological factors such as ischemia, hypoxia, oxidative stress, or calcium homeostasis disorders can disrupt endoplasmic reticulum homeostasis, cause endoplasmic reticulum dysfunction, and lead to the accumulation of unfolded or misfolded proteins, thus inducing ERS and triggering unfolded protein response, thereby activating the corresponding endoplasmic reticulum and calcium regulatory mechanisms and other signal transduction pathways to restore endoplasmic reticulum homeostasis. When the ERS intensity is too high or lasts for a long time, the endoplasmic reticulum cannot restore homeostasis. At this time, multiple pathways induce cell apoptosis, which together with multiple factors lead to cardiomyopathy and heart failure. Increasing evidence shows that ERS is associated with diabetes, endothelial dysfunction, and atherosclerosis. GLP-1 receptor agonists can inhibit ERS and reduce intracellular calcium concentration through indirect effects, thereby reducing myocardial damage. Their effects may be related to the inactivation of the ERS signaling pathway. GLP-1 Receptor Agonists Affect DCM by Interfering with Inflammatory Response Oxidative stress is an important factor driving the inflammatory response. High glucose status and insulin resistance in diabetic patients promote cell metabolism disorders and release a large amount of reactive oxygen species, which in turn activates nuclear factor κB, induces the expression of inflammatory factors, damages vascular endothelial cells, and causes abnormal proliferation of smooth muscle cells. It will affect the structure and function of the heart in the long term, leading to the occurrence and development of DCM. GLP-1 receptor agonists can inhibit the inflammatory pathway through direct or indirect effects, activate the AMP-activated protein kinase (AMPK) pathway, reduce the activity of nuclear factor κB, reduce the production and release of inflammatory mediators, inhibit the differentiation of inflammatory monocytes and macrophages, and Reduce heart and blood vessel inflammation, etc. GLP-1 Receptor Agonists affect DCM by protecting Mitochondria and Improving Oxidative Stress The tissue damage of DCM mainly has the following five mechanisms: (1) Increased metabolism of glucose and other sugars through the polyol pathway; (2) Formation of intracellular advanced glycation end products; (3) Advanced glycation end products Increased expression of receptors and ligands; (4) activation of protein kinase C subtype; (5) overexpression of hexosamine pathway activity. Mitochondria are one of the main sources of reactive oxygen species in cardiomyocytes, and reactive oxygen species are the upstream products of the above five pathways. In the body’s high-glucose state, due to changes in metabolic substrates, the oxidation of free fatty acids increases the transfer of reducing equivalents, which can damage the ability of the electron transport chain, increase the generation of reactive oxygen species, excessive reactive oxygen species deposition, or weaken the antioxidant mechanism. The redox imbalance leads to oxidative stress; this process and the high glucose state can simultaneously promote oxidative stress and the body’s inflammatory response, damage myocardial cells, and then lead to myocardial fibrosis, ventricular hypertrophy, coronary microvascular damage, cardiac systolic and diastolic function obstacles, eventually developing into DCM. In the entire pathogenesis of DCM, the effects of GLP-1 receptor agonists on metabolic disorders, accumulation of reactive oxygen species, ERS, and inflammatory responses all require the oxidative stress link. The oxidative stress link can be called the key to the pathogenesis of DCM. Therefore, intervening in the oxidative stress process is an important step in improving the prognosis of DCM. Studies have shown that exenatide reduces oxidative stress in cardiomyocytes and improves mitochondrial function by activating autophagy; it can also remove dysfunctional mitochondria to inhibit cardiomyocyte apoptosis, achieve anti-apoptosis and antioxidant effects, and protect cardiac function. Exenatide can also inhibit the activation of caspase-1, indirectly attenuating mitochondrial damage and reducing reactive oxygen species deposition, thereby attenuating oxidative stress.

Alzheimer’s disease (AD) is a progressive neurodegenerative disease with an insidious onset. Clinically, it is characterized by comprehensive dementia manifestations such as memory impairment, aphasia, apraxia, agnosia, impairment of visuospatial skills, executive dysfunction, and personality and behavioral changes. Alzheimer’s disease is progressive and irreversible and can only be delayed with medication. Once it develops into moderate or severe disease, it will cause significant harm to the lives of patients and their families. Figure 1. Schematic overview of invasive biomarkers. Pathological Mechanism of AD The pathological mechanism of AD is currently unclear, and is currently summarized as neurodegenerative changes and inflammatory response. Neurodegenerative changes involve neuronal degeneration, nerve cell apoptosis, and the generation of new neurons. Inflammatory response refers to the inflammatory response in the brain of AD patients, including the release of activated cytokines and inflammatory cytokines. In addition, many studies have shown that the pathological mechanism of AD is also affected by protein oxidative stress. Oxidative stress refers to the production of free radicals and the increase in reactive oxygen species under conditions of intracellular hypoxia or anoxia, leading to the destruction of cell structure and function. Studies have found that oxidative stress can affect the apoptosis and degeneration of nerve cells in the brains of AD patients, thus increasing the severity of the disease. There are reports that AD is also affected by neurotransmitters. The level of neurotransmitters in the brains of AD patients is lower than that of normal people. This may be one of the reasons for the cognitive impairment of AD patients. In addition, AD patients have obvious amyloidosis in their brains. Amyloidosis refers to a variety of pathological changes that occur in the brains of AD patients, including amyloid crystals, neuronal degeneration, and nerve cell apoptosis. Such lesions may be a key cause of AD and serve as diagnostic markers for AD. AD Diagnostic Markers Apolipoprotein Eε4 Apolipoprotein Eε4 (APOEε4) is a very clear risk gene for late-onset AD, which may advance the age of onset and accelerate the development of cognitive decline. It has been recognized as a genetic marker of AD. ApoE is a key component of plasma lipoproteins, which can produce β-amyloid peptide and maintain neurological function. β⁃amyloid Under the action of hydrolyzing amyloid precursor protein (APP), β⁃ and γ⁃secretase can produce β⁃amyloid protein (Aβ). Such polypeptides are distributed in many cells and can also be found in blood, cerebrospinal fluid (CSF) and brain interstitial fluid. The most widespread subtypes of Aβ are Aβ40 and 42. The latter are neurotoxic and are related to the formation of AD. Most of them are generated from microglia and astrocytes, have hydrophobic characteristics, and play a major role in plaque formation. Aβ42 in CSF is generally considered to be a biomarker of AD and is helpful in diagnosing preclinical AD. Lower levels of Aβ42 in the cerebrospinal fluid are associated with higher Aβ plaques in the brain, and the concentration levels of Aβ42 in the cerebrospinal fluid of AD patients are significantly reduced compared with normal controls. Currently, the detection of Aβ42 in cerebrospinal fluid can be carried out using immunological technology or mass spectrometry. However, the specificity of Aβ42 in detecting AD needs to be improved. Some studies have shown that the ratio of Aβ42/Aβ40 can be used to judge amyloid deposition in the brain in the early stages of the disease. The specificity of this method is significantly better than that of measuring Aβ42 alone. However, the detection of cerebrospinal fluid is invasive and is not conducive to early screening. Currently, more and more scholars are beginning to explore blood biomarkers. Aβ protein can be measured in plasma, but the Aβ concentration in plasma will be affected by factors such as platelets, and its correlation with AD is not as good as the Aβ concentration in cerebrospinal fluid. However, gratifyingly, studies have shown that using mass spectrometry to detect plasma Aβ concentration can show a significant correlation with cerebral amyloidosis. Therefore, exploring more effective measurement technologies is the core of breakthroughs in blood screening. Tau The microtubule system is a component of the neuronal cytoskeleton, and its composition contains tubulin and microtubule-related proteins, with the latter having a higher content of Tau protein. The main function of Tau protein is to bind tubulin and promote the production of microtubules. If Tau is over-phosphorylated, it can accumulate in cells, damage the function of microtubules, and damage the cytoskeleton or axonal transport of nerve cells, thus triggering neurodegeneration. It is generally believed that total⁃t tau (T⁃t tau) and phosphorylated⁃Tau (P⁃tau) concentrations are related to AD. The increase in T-tau reflects the degree of degeneration and damage of neurons and axons. High levels of P-tau suggest the formation of neurofibrillary tangles. CSF P-tau is regarded as the most specific marker of AD, and the concentration of CSF P-tau in AD patients will be significantly increased. Studies have shown that P-tau protein can be used to judge the conversion of MCI to AD, and therefore can be used as a key marker for early clinical diagnosis. Nowadays, P-tau levels in CSF can be measured through immunological methods such as ELISA. Neurofilament lightweight peptide (NFL) NFL is a peptide encoded by the NEFL gene. It is an intrinsic cytoskeletal protein that can be used as a biomarker of axonal injury and can be obtained in both cerebrospinal fluid and plasma. Neurons in the brain interstitium and cerebrospinal fluid release NFL after death, and then enter the blood. The serum or plasma level of NFL is closely related to CSF ​​NFL, and is increased in both familial and sporadic AD. In familial AD, patients show increased concentrations of NFL 10 years before the onset of significant clinical symptoms. . Therefore, plasma or serum NFL may be a reliable blood biomarker for neurodegenerative diseases in AD and other neurodegenerative diseases. Plasma glial fibrillary acidic protein (GFAP) GFAP is a key indicator used to monitor astrocyte viability. Previous studies have shown that GFAP is significantly increased in the brain and CSF of AD patients, suggesting that GFAP may become a potential biomarker of AD. Studies have shown that serum GFAP levels in AD patients are significantly higher than those in normal controls. In addition, reports also suggest that the plasma concentration of early-onset AD patients is significantly higher than that of normal controls. While the increase in GFAP concentration in the plasma of late-onset AD patients is not as significant as that of early-onset AD, it is also significantly higher than that of normal controls group. Therefore, GFAP is expected to become a member of the blood AD markers and provide new clues for the clinical diagnosis of AD. Neurogranin (Ng) Ng is also a potential biomarker for early diagnosis of AD and is usually expressed in granular structures in pyramidal cells in the hippocampus and cortex. Ng has been shown to be related to synaptic plasticity, synaptic regeneration and other functions mediated by the calcium/calmodulin signaling pathway. Some scholars have compared the levels of neurogranin in the cerebrospinal fluid of patients with AD, frontotemporal dementia, Lewy body dementia, Parkinson’s disease, etc. The results show that the concentration of neurogranin in the cerebrospinal fluid of AD patients is significantly higher than that of other diseases, further illustrating that Higher Ng protein concentration in cerebrospinal fluid has a specific screening effect on AD.

Herpes simplex virus (HSV) belongs to the herpesvirus alpha subfamily and has a unique 4-layer structure. The core of HSV is a linear double-stranded DNA of about 152 000 bp, surrounded by an icosahedral capsid. The outer capsid is a membrane, which contains more than 20 important proteins that can regulate the virus replication cycle. It is a characteristic structure of herpes viruses, can connect the capsid and envelope to form a complete virus particle. The outermost layer of HSV is the lipid bilayer envelope, which contains at least 12 viral membrane proteins (gB, gC, gD, gE, gG, gH, gI, gJ, gK, gL, gM, gN). Entry into host cells and viral immune evasion are both critical. Figure 1. Schematic Representation of Acute and Latent Herpes Simplex Virus-1 (HSV-1) Infection in Humans. HSV Type There are 2 serotypes of HSV: herpes simplex virus ⁃1 (HSV⁃1) and HSV 2, which share 83% homology in their amino acid sequences. HSV⁃1 infection can cause inflammation of the lips in the human body. When the infection is severe, it can induce conjunctivitis and lead to blindness. It may also invade nerve cells and damage the nervous system, leading to encephalitis. HSV ⁃2 is the main causative agent of genital herpes. Once infected, patients will carry the virus for life and develop periodic genital herpetic lesions. Recent epidemiological studies have shown that genital damage caused by HSV⁃2 infection increases the risk of HSV⁃2 carriers contracting and transmitting human immunodeficiency virus (HIV). Even if viral shedding occurs without causing genital damage, will significantly increase the probability of HIV transmission. Different herpesviruses have similar viral structures and can enter host cells through similar membrane fusion mechanisms. Membrane fusion requires the interaction of four basic glycoproteins (gB, gD, gH and gL) on the virus surface. The complex composed of these four glycoproteins is called the core fusion mechanism. They are usually located on the surface of the viral envelope and can pass through the cascade The action transmits the activation signal from gD to gB, causing conformational changes in gB, forming a fusion pore on the host cell membrane to achieve membrane fusion, allowing the virus to successfully release the nucleocapsid and DNA into the host cell for a new round of replication. The Mechanism of HSV Entry into Host Cells HSV can infect a variety of cells including neurons, epithelial cells, and fibroblasts with the help of fusion proteins. In the current research model, specific binding of HSV to receptors on host cells can cause conformational changes in viral surface glycoproteins, thereby mediating HSV entry into cells. The mechanism of action is achieved through two similar pathways: 1. Entering cells through the direct fusion of the viral envelope and the host cell plasma membrane, and the virus uses this pathway to enter fibroblasts and lymphocytes; 2. The virus triggers the invagination of the host cell plasma membrane to form endosomes, Endocytosis into the cytoplasm is the pathway used by the virus to enter neuronal cells. However, the specific entry route may depend on the type of host cell surface receptors, such as the paired immunoglobulin-like receptor α (PILRα) of gB, and the integrin receptors αvβ3, αvβ6, and αvβ8 of gH⁃gL, which may be related to It is related to the role of virus non-essential envelope proteins (gC, gE, gG, gI, gJ, gK, gM, gN), and the number of receptors on the host cell membrane may also affect the entry method of the virus.

As a tumor suppressor, p53 is one of the most commonly mutated genes in human cancers. Due to its important role in tumor suppression, p53 has attracted great interest from researchers in drug development. As a transcription factor, p53 is directly or indirectly involved in regulating many genes, including cell cycle arrest, apoptosis, senescence, and DNA repair. In recent years, p53 has become one of the most important and attractive drug targets in cancer treatment. This article will review research on p53-based gene therapy, targeted therapy and immunotherapy. Figure 1. The p53 pathway. Overview of the Structure and Function of p53 Gene The human p53 gene is located on chromosome 17p13.1, contains 11 exons, encodes 393 amino acids, and has a relative molecular mass of 43,700. p53 is a sequence-specific DNA-binding protein with two transactivation domains at its N-terminus, followed by a conserved proline-rich domain, a central DNA-binding domain, and finally a The C terminus encodes its nuclear localization signal and an oligomerization domain required for transcriptional activity. p53 mutations or deletions occur in about half of cancer patients, and it is one of the most commonly mutated genes in cancer. p53 mutations mainly occur in the DNA-binding domain, and several amino acid residues have a much higher mutation frequency than other amino acids. These residues are often called hotspot TP53 mutations, among which R175, G245, R248, R249, R273, and R282 are the major hot spot. As a tumor suppressor, p53 can regulate cell division, prevent cells with mutated or damaged DNA from dividing, and transmit apoptosis signals to these cells, thereby preventing tumor formation. p53 can respond to cellular stress or DNA damage, activate a variety of transcription targets, and coordinate a variety of responses, including cell cycle arrest, DNA repair, antioxidant effects, anti-angiogenesis effects, metabolism, autophagy, senescence, and apoptosis. Under non-stress conditions, p53 is ubiquitinated by E3 proteins such as mouse two-microbody protein 2 (MDM2), constitutive photomorphogen 1 (COP1), RING domain family member 2 (PIRH2), and triple motif protein 24 (TRIM24), and subsequent degradation by the proteasome maintains p53 protein at a low level. Another important negative regulator of p53 is mouse double microbody protein X(MDM4) can inhibit cell cycle progression in a variety of ways, one of which is to upregulate p21 protein expression. Subsequently, p21 protein can bind to cell cycle proteins, causing cell cycle G1 phase arrest. In addition, p53 can also bind to other target genes, such as 14-3-3σ and cell division cycle protein 25 (CDC25), to arrest the cell cycle in the G2/M phase. In addition, p53 also plays an active role in many different types of DNA repair, including nucleotide excision repair, base excision repair, mismatch repair and non-homologous end joining. Beyond this, p53 mutations provide a selective advantage to tumor cells, allowing them to circumvent cell cycle checkpoints, avoid apoptosis and senescence, and proliferate under conditions where normal cells cannot proliferate. Targeting MDM2 and MDMX MDM2 inhibitors MDM2 is a major negative regulator of p53. In cancer cells carrying WT p53, MDM2 is often overexpressed through gene amplification or transcriptional upregulation. Therefore, MDM2 is a good target for cancer treatment. Common inhibitors of MDM2 are Nutlin3a and Nutlin-3a-aa, which are cis-imidazole compounds that can induce p53 activation in WT p53 cancer cells but have no effect in mutant p53 (mutp53) cells. Nutlin- 3a-aa is more active than Nutlin-3a against purified wild-type MDM2 and is more effective in increasing p53 levels and releasing the transcription of p53 target genes from MDM2-induced repression. MDMX inhibitors Like MDM2, as a negative regulator of p53, MDMX has also attracted great attention from inhibitor developers, but few inhibitors have been discovered so far, and most of them are short peptides that inhibit MDMX activity. To inhibit the interaction between MDMX and p53, the researchers designed a highly specific short peptide (SAH-p53-8) that can activate p53 in MDMX-dependent cancer cells and induce apoptosis in vivo. However, later studies showed that SAH-p53-8 binds very tightly to serum, which limits its entry into tumor cells and further clinical development. Targeting p53 mutants Most tumor cells contain p53 gene alterations, including mutations, deletions, and translocations. However, the incidence of p53 gene mutations is higher than deletions and translocations. p53 mutations not only lead to changes in p53 function, but also confer new functions to these mutants, called gain of function (GOFs). To target cancer cells with p53 mutations, several therapies have been developed. Activate WT p53 Activity As a tumor suppressor gene, reactivating the activity of WT p53 is a better treatment method. The p53-Y220C mutant is an excellent example of developing mutant p53 therapeutics through protein stabilization. Some studies have found that small molecule compounds PK083 and PK7080, which target the unique surface cracks produced by the Y220C mutation, can bind to the Y220C mutant, restore the wild conformation, and induce Y220C-dependent cell cycle arrest and apoptosis. Studies have shown that the correct folding of p53 requires the presence of zinc, and the lack of zinc in the DNA-binding domain leads to misfolding of the protein. By providing zinc to certain p53 mutants such as R175H, these mutants can restore the conformation and function of WT p53, once again functioning as a tumor suppressor gene. GOFs that Inhibit Mutant p53 Most p53-based drug development targets WT p53 activity in cancer cells, but there are also attempts to eliminate mutp53 GOFs activity by targeting mutp53 for rapid degradation. Since the introduction of mutp53 GOFs, many mechanisms of mutp53 GOFs have been proposed, such as: BAG5 interacts with mutp53 protein and promotes the accumulation of mutp53 protein, resulting in GOFs promoting cell proliferation, tumor growth, cell migration and chemical resistance. p53 mutants down-regulate solute carrier family member SLC7A11 (solute carrier family 7 member 11) through the epidermal growth factor receptor (EGFR)/extracellular regulated protein kinase (ERK) cycle or through transcriptional regulatory protein 1 (BACH1)-mediated down-regulation, enhances the invasion and metastasis of cancer cells. A general strategy for GOFs targeting mutant p53 is to utilize existing therapeutics that target pathways or genes affected by mutant p53, such as EGFR inhibitors, statins of the metovalerate pathway, and inhibitors of mixed lineage leukemias (MLLs). Furthermore, there is emerging evidence that the interaction between mutp53 and the tumor microenvironment may shape the GOFs of mutant p53. The GOFs of p53 play a key role in the occurrence and development of cancer. A comprehensive understanding of the GOFs of p53 will help provide a better theoretical basis for personalized drug therapy in cancer treatment. Targeting Mutant p53 Stability One strategy to target p53 mutants is to reduce their stability. Some p53 mutants are oncogenic, and reduced levels can lead to cancer cell death. Studies have found that the stability of mutant p53 can be enhanced by the heat shock protein (HSPs) family, and the binding of the HSPs complex prevents the degradation of mutant p53 by E3 ubiquitin ligases such as MDM2 and HSP70-interacting protein (CHIP).