Malignancy currently ranks first among the causes of immature human mortality (death before the average life expectancy of the population). The development and progression of the vast majority of cancers depends on chronic inflammation. Chronic infection, environmental exposure, and immune imbalances can cause chronic, uncontrollable inflammation in the host, which provides a microenvironment for cancer to occur and affects its entire development. The process of cancer occurrence and development reflects the evolutionary process of "mutation-selection-adaptation" and the existence of a "degeneration" process opposite to development. Through the in-depth study of the intrinsic mechanism of hepatocellular carcinoma (HCC) caused by hepatitis B virus (HBV), combined with the results of other tumor second-generation sequencing studies, the team proposed the theoretical framework of "Cancer Evolutionary Development (Cancer Evo-Dev)" for the first time in the world. The basic content of the theoretical framework is as follows: in the early stage of cancer, some cells in precancerous diseased tissues induce nucleic acid-editing enzyme expression through inflammation, resulting in somatic mutations, changing the original cell signaling pattern; through the evolutionary selection of the inflammatory microenvironment, a small number of mutated cells survive, the de-epigenetic modification (such as demethylation) of the dedifferentiation process occurs, and then from epithelial cells to interstitial cells, that is, the EMT process, thus giving these cells the ability to overcome "aging", plunder nutrients, and obtain unlimited expansion Some of these cells have evolved further and have been given the potential to invade. In the evolutionary process of "mutation-selection-adaptation", these cancer-related common or characteristic somatic cell variations and signaling networks play a decisive role in cancer evolution and development. In the evolution and development of cancer, from precancerous lesions to cancer occurrence and early invasion, there are a series of molecular events such as epigenetic modification, somatic mutations, and dynamic signal network regulation. In this paper, colorectal cancer (CRC) and HBV-associated hepatocellular carcinoma (HBV-HCC), two representative malignant tumors with a heavy malignant tumor burden and inflammation-cancer transformation characteristics in China, are selected, and the early key molecular events in the process of occurrence and invasion are described, in order to determine which precancerous disease changes are more likely to worsen and which cancers are more likely to occur early invasion, so as to achieve the goal of "gate forward shift" for CRC and HBV-HCC-specific prevention and treatment.
1 The relationship between genetic susceptibility to inflammatory signals such as HLA-II antigen and NF-κB and chronic inflammation and its role in cancer evolution and development
1.1 Relationship between genetic susceptibility to inflammatory signals and chronic inflammation
In HBV-HCC, previous studies have shown that key pathway molecules for immune and inflammatory signaling, such as human leukocyte antigen class II, HLA-II.-DP, -DQ and -DR, as well as STAT3 and NF-κB innate single nucleotide polymorphisms-genetic predisposition are associated with chronicity of HBV infection. Rare genotypes (or alleles) of congenital genetic polymorphisms of the HLA gene are significantly and negatively correlated with chronic HBV infection, particularly in people with HBV genotype C infection. Genetic polymorphisms that increase NF-κB activity have been found to contribute to B-type HBV clearance, while the rs2233406 variant genotype may significantly increase the risk of HCC by promoting immune selection for HBV mutations. STAT3-related studies suggest that the interaction of SNPs with HBV mutations makes hosts with HBV mutations more susceptible to liver cancer.
In colorectal cancer, during the evolution of ulcerative colitis to CRC, the regulatory T cell (Treg) subpopulation IL-8 + Foxp3 + CD4+ T cells and KLRG + T cells increased, and both had immunosuppressive and pro-inflammatory bifunctional activities. At the same time, the epimodulatory molecule PRC2 complex and EZH block T cells enter the transformation microenvironment, which can strengthen the formation of immunosuppressive networks.
In summary, under the genetic predisposition dominated by certain genetic factors, continuous stimulation of the external environment is the basis for the production and maintenance of chronic inflammation. Community-based epidemiological research is irreplaceable for studying the role of genetic and environmental factors in the etiology of cancer. Therefore, etiological research should strengthen the links between epidemiology, genetics, immunology, molecular biology and clinical medicine.
1.2 Inflammatory signaling molecules influence the selective effect of HBV variants by influencing immune HBV function
The innate genetic polymorphisms of the HLA-II antigen gene and the inflammatory signaling pathways STAT3 and NF-κB, which play a decisive role in the carcinogenic process of viral hepatitis B, determine the inflammatory response in which the immune system participates, and have a definite selective effect on the characteristic variants of HBV. The main genetic polymorphisms of these HLA-II immune genes and key molecules of inflammatory signaling networks such as STAT3 and NF-κB not only affect the chronicization of HBV infection, but also promote the selection of HCC-related HBV variants, reflecting the process of "optimization" of environmental carcinogenic factors by genetic factors through genetic function. During the development of "HBV-chronic HBV-cirrhosis-HCC", HCC-associated high-risk mutations of HBV gradually accumulate but the diversity of T cell receptors gradually decreases. Genetic polymorphisms that affect the expression of immune class II leukocyte antigen genes HLA-DR can significantly reduce the frequency of high-risk HBV mutations A1762T/G1764A and reduce the risk of HCC.
1.3 Key molecular events of chronic inflammation in the early commonalities and properties of cancer evolution
Our team compared the HBV sequences in peripheral blood, paracancerous liver tissue and HCC tissue, and found that the complexity of HBV mutations gradually decreased in peripheral blood-paracancerous liver tissue-HCC tissue, but the proportion of high-risk mutations increased, showing typical environmental screening characteristics. In the process of intestinal inflammation or adenoma evolution to CRC, the fungal flora appears in the characteristic screening of diversity and composition; Candida albicans has an abnormal activation of the intestinal epithelium Wnt/β-catenin signaling pathway and inhibits the expression of chemokine CXCL10 and other "bidirectional" regulatory activities, which is a key molecular event for chronic inflammation to promote cancer occurrence.
Recent studies have used a variety of pathogenic animal models of liver fibrosis to confirm that LECT2-Tie1 and Slit2-Robo1 signals can regulate the early liver fibrosis process of cirrhosis. Fetal-hepatic-based related studies have shown that the MBNL3-lncRNA-PXN regulatory network active in the embryonic stage is an early key event in the occurrence of HBV-HCC, further confirming that the early molecular events of HCC are closely related to the embryonic development events of liver tissue. MiR-155 expression levels in PERIPHERAL BLOOD CD4+ and CD8+ T cells in patients with chronic viral hepatitis B are significantly elevated and may be positively associated with immune activation. The study believes that some of the newly discovered key molecules are expected to provide new biomolecular markers for the early diagnosis of cancer. It can be seen that most of the early molecular events of cancer are related to inflammation-related molecules and early embryonic expression molecules.
2 A key molecular event in which chronic inflammation promotes the transformation of precancerous lesions into cancer
2.1 The APOBECs family is the bridge between chronic inflammation and cancer
The occurrence and development of cancer is the evolutionary development process in the environment of chronic uncontrollable inflammation, and the evolution of species in the natural environment follows the evolutionary law of "mutation-selection-adaptation". The apolioprotein B mRNA-editing enzyme catalytic polypeptide (APOBEC) family is a highly potent class of cytidine deaminases whose transcription can be activated by pro-inflammatory cytokines and chemokines, and play an important role in acquired immune mechanisms such as innate immunity and antibody production. While inhibiting viral replication through mutationalization, the APOBECs family can also drive the formation of viral mutants with pro-cancer effects, and their gene coding function can also induce the generation of drive mutations with pro-cancer effects. As a signature enzyme that plays a bridging role in inflammation-cancer transformation, the APOBECs family is essential for the evolutionary development of cancer and is a breakthrough point in the process of through-inflammatory-cancer transformation.
2.2 Uracil DNA transglucosase
Maintaining a "pro-mutation-repair" balance with APOBECs is uracil DNA transglucosase (UNG). UNG plays a key role in maintaining genome integrity by reducing mutagenic events caused by mutations in the transition from G:C to A:T. UNG initiates a variety of downstream repair pathways that eliminate mutations C>U while also producing a small number of other mutations. If UNG removes uracil to form an AP site, the base can be inserted without a template during complementary chain synthesis, and if guanine is inserted again, the nucleic acid chain can be restored to a normal state; if adenine is inserted, a C/G to T/A mutation is formed. In fact, the latter often occurs, which is one of the characteristic labels of APOBECs. At the same time, intranuclear subtypes (UNG2) are mainly concentrated in the G1 stage of cell division, which means that the metabolism of "mutation-excision-repair/degradation" is more frequent, and it accelerates the accumulation of mutations on cells with vigorous division such as cancer initiating cells.
2.3 How APOBECs and UNG Maintain Balance
The equilibrium relationship between APOBECs and UNG is mainly reflected in the following three aspects: (1) UNG can recognize the C>U mutation caused by the APOBECs family, and excise uracil to form AP sites; (1) UNG includes mitochondrial subtypes (UNG1) and UNG2, of which UNG2 activity is stronger, which corresponds to the mutagenic function of APOBECs clearing HBV cccDNA into the nucleus and the mutagenic function of the human genome ;(3) The deamination activity of the APOBECs family on single-stranded DNA is 200-300 times that of double-stranded DNA, and the uracil excision activity of UNG2 pair of single-stranded DNA is 50 times higher than that of double-stranded DNA. However, the effect of the balance between UNG and APOBECs is not simply mutual restriction and mutual cancellation, but both containment and cooperation.
2.4 Role of promoters and enhancers SNPs of APOBECs and UNG in inflammation-cancer transformation
"Yin" and "yang" inflammatory factors can induce imbalances in APOBECs and UNG, which in turn affect the inflammatory carcinogenic process. SNP genotype rs2267401: APOBEC3B promoter (-338 bp:T→G) and rs3890995:UNG enhancer (-1 948 bp:T→C) in the transcriptional regulatory region enhance gene damage-repair imbalances triggered by the "negative" cytokine IL-6, increasing the probability of viral mutations, thereby increasing the risk of cancer. The expression level of APOBEC3B in adenomatous polyposis tissue was significantly higher than that of normal tissue, reaching the level of CRC tissue, and significantly enriched APOBEC-labeled mutations. In the pre-evolutionary stage of chronic carcinogenicity of HBV (chronic hepatitis, cirrhosis), APOBEC with upregulation of inflammatory factors promotes the production of APOBEC-related mutations in the HBV genome, which significantly increase the risk of HCC. It was found that activating inflammatory pathways significantly upregulated the expression of the cytidine deaminase APOBECs family that promote gene mutations in both HCC and CRC.
3 Key molecular events in the evolution of carcinoma in situ towards early invasion
The theory of "cancer evolutionary development" elaborates that the key molecular events that drive cancer evolution give tumor cells survival advantages by regulating the "dry" signaling pathway, inducing the transformation of epithelial cells into interstitial cells, changing energy metabolism patterns, influencing the microenvironment, etc., so that they adapt to inflammatory selection pressures, and constantly evolve to lead to invasion.
3.1 Regulation of "dry" signaling pathways promotes early invasion
It was found that Cortactin and Prohibitin overexpression can regulate the formation of aggressive pseudopods in CRC cells; KRAS mutation can inhibit IRF2 expression, thereby inhibiting the CXCL3-CXCR2 axis, so that the myelogenous inhibition of cell proliferation, thereby inhibiting the immune response and T cell infiltration, and promoting CRC evolution. A number of key molecular events that significantly promote the evolution of HCC by modulating various dry signaling pathways have also been identified in HCC, including HBV integration mutations significantly enriched in the mTOR signaling pathway, PRMT1 activating the SATA3 signaling pathway, hypoxia-induced 14-3-3ζ, ICR and ICAM abnormal upregulation, lncRNA-FTX abnormal downregulation, and Bif-1-mediated CDC42 upregulation. These key signaling molecules all play an important role in the "development" of cancer.
3.2 EMT promotes early invasion
EMT refers to the biological process by which epithelial cells are transformed into cells with interstitial phenotypes through specific procedures. Studies have shown that FOXF1, TUSC3-induced WNT/β-catenin and MAPK signaling pathway activation, miR-20a/miR-106a-mediated WTX deletion can promote EMT, which in turn promotes CRC invasion. In HCC, macrophages with low expression of miR-26a promote EMT in HCC cells by secreting TGF-β, while CD31 induces EMT through the ITGB1-FAK-Akt signaling pathway, both of which promote HCC metastasis and invasion. Other studies have shown that tumor exosome lncRNA promotes HCC and CRC invasion. Based on the results of domestic and foreign studies, it is expected to find the molecular markers of mutation-induced carcinogenesis that lead to "reverse differentiation" and the key molecules of targeted intervention.
3.3 Inflammatory microenvironment and microbes promote early invasion
It was found that miR-25-3p can promote tumor vascular formation and increase vascular permeability through exosomes to vascular endothelial cells, and then promote the formation of microenvironment before metastasis, while the detection of miR-25-3p expression levels of peripheral blood circulation exosomes can predict distal CRC metastasis. The intestinal flora stimulates the production of cathepsin K by modulating the polarization of TLR4-dependent M2-type macrophages and promoting CRC invasion and metastasis. Among them, specific strains can be used as biomarkers to predict the effect of immunotherapy, and intestinal microbiota intervention methods such as related microbiota preparations and microbiota transplantation are expected to become auxiliary means of tumor immunotherapy.
4 Summary
In the study of inflammation-cancer transformation, from the perspective of chronic inflammation and cancer genetic susceptibility, such as human class II leukocyte antigen (HLA-DP, -DQ, -DR) and the role of genetic susceptibility of inflammation-related gene expression and function on the maintenance of chronic uncontrollable inflammation, etc., using epidemiological large-scale population cohort studies, the etiological time series linkage was established, and the various types that appeared in the complete development process of "chronic inflammation - precancerous lesions - carcinoma in situ - invasion cancer" were studied in depth" Immunity-selection-adaptation" phenomenon and key molecular events. It was found that the molecules associated with the occurrence and invasion of CRC and HCC were related to the balance between APOBECs that promote gene mutations and UNG in mutation repair, so it is proposed that the balance between APOBECs and UNG is an important step in pro-cancer mutations and a bridge for inflammatory transformation. It was also found that the inflammatory microenvironment can also promote the dedifferentiation of differentiated cells, which is manifested in the gradual removal of epigenetic modifications, which promotes the occurrence of EMT, and the further "degeneration" of differentiated cells to early non-differentiated stem cells, showing a "reverse development" process. These findings also further enrich the evidence at all levels of cancer evolutionary development and improve the theoretical system of "cancer evolutionary development". The theoretical system of "cancer evolution and development" still needs to be continuously improved in the future, and the current evidence supports the authenticity of the "cancer evolutionary development" theory, and it is expected to use the principle and method of "cancer evolution and development" to guide the specific practice of cancer prevention and treatment, such as determining what kind of precancerous lesion population is more prone to cancer, finding out cancer-specific preventive measures; determining what kind of early cancer will have early invasion, so as to carry out targeted treatment to reduce cancer mortality rate and avoid excessive medical treatment.