Game of clones: Battles in the field of carcinogenesis.

The first hints of alterations occurring in normal cells came from early studies revealing unexpected pathological findings in what appeared to be phenotypically normal tissues. In 1953, pathologic atypia was characterized in “normal appearing” tissues surrounding oropharyngeal carcinoma ( O’Shaughnessy et al., 2002 ; Slaughter et al., 1953 ). This phenomenon, which was coined “ field of cancerization ”, comprises cellular states that harbor some but not all phenotypic traits entailed for malignancy ( Braakhuis et al., 2003 ; Curtius, Wright, & Graham, 2018 ). Since then, field effects have been described in various epithelia, whereby alterations were shown to adopt a multifocal spatial pattern, followed by expansion of smaller “patches”, each comprising a group of cells that share a common cancer-associated genetic alteration, such as TP53 mutations ( Garcia, Park, Novelli, & Wright, 1999 ). A clonal unit or patch may acquire additional tissue-relevant genetic alterations that provide a growth advantage, thus gradually displacing the normal mucosa and enabling the conversion of that clonal patch into an expanding field ( Fig. 1 ). Random somatic mutations attributed to the infidelity of DNA replication machinery or induced by mutagenic insults induce cancerized fields comprised of genetically distinct clones in competition, until the fittest clone dominates the field ( Curtius et al., 2018 ). A concept related to field of cancerization is the “field of injury”, which encompasses molecular changes in tissues incurred by mutagen exposure such as smoking or UV radiation ( Dakubo, Jakupciak, Birch-Machin, & Parr, 2007 ; Steiling, Ryan, Brody, & Spira, 2008 ). This field of injury reflects host mechanisms of defense and the damage caused by exposure to a carcinogen, and which may not necessarily lead to overtly malignant fates ( Dakubo et al., 2007 ; Gower, Steiling, Brothers 2nd, Lenburg, & Spira, 2011 ). Field of injury and cancerization have been documented and described across multiple epithelial tissue types. Exposure of cervical epithelia to high-risk strains of human papillomavirus affected the function of p53, thus initiating a field of pre-invasive cervical intraepithelial neoplasia ( Burd, 2003 ). H. pylori is a known trigger for a field characteristic of intestinal metaplasia in the stomach, yet the mutational spectrum was shown to be distinct from that seen in gastric tumors ( Curtius et al., 2018 ; Wang et al., 2014 ; Yakirevich & Resnick, 2013 ). A field of cancerization can also be ignited by gastroesophageal reflux disease (GERD), a known risk factor for esophageal carcinoma, which is typified by a mutational signature characteristic of acid exposure, implying that esophageal cancerized fields may be implicated in tumor formation ( Dulak et al., 2013 ; Rubenstein & Taylor, 2010 ). In the context of exposure to tobacco smoke, molecular changes throughout the entire respiratory tree have been documented ( Steiling et al., 2008 ), and mutations in TP53 in the lung and airways were frequently observed in cancerized fields ( de Bruin et al., 2014 ; Steiling et al., 2008 ). In the next section, we will provide an overview of how cancerized fields develop as well as their implications on cell fate and possibly carcinogenesis. More specifically, we will focus on studies done on cancerized fields in the lung epithelium and the possible link to development of premalignant lesions and different subtypes of lung cancers, particularly non-small cell lung cancers (NSCLC). As previously described, clonal patches have been evident in lung epithelium of cancer-free individuals at increased risk for lung cancer and with a long period of tobacco exposure ( Mao et al., 1997 ; Park et al., 1999 ; Wistuba et al., 1997 ) ( Fig. 1 ). Indeed, it was postulated that repeated insults, such as exposure to tobacco smoke, promote chronic wound healing, which in turn enables cells with genetic alterations needed for survival and preferential expansion ( Wynn & Vannella, 2016 ). A mosaic of clonal (and subclonal) outgrowths may develop in the tobacco-exposed airway, and damage-surviving clonal clusters continue to accumulate mutations as they evolve into overt lesions. Histological and molecular changes have been also studied in cancerized fields of airway of patients with lung cancer and in smokers ( Kadara & Wistuba, 2012 ). A detailed analysis of histologically normal, premalignant and malignant epithelia from smokers and patients with lung squamous carcinoma (LUSC) indicated that several, consecutive allele-specific chromosomal and genomic events begin in dispersed and clonally-independent foci at early stages of pathogenesis, including in normal-appearing bronchial epithelium ( Wistuba, Behrens, Milchgrub, Bryant, & Hung, 1999 ) ( Fig. 1 ). In addition, it has been demonstrated that KRAS ( Nelson, Wymer, & Clements Jr., 1996 ) and EGFR ( Tang et al., 2005 ) mutations, common driver mutations in patients with lung adenocarcinoma (LUAD), are found in peripherally located histologically normal lung tissue adjacent to the LUADs themselves. Higher frequency of EGFR mutation was found in normal bronchial cells with increasing proximity to the tumor, possibly signifying a localized field effect in LUAD pathogenesis ( Tang et al., 2005 ). Increased Ki67 expression, proliferation, as well as metabolic pathway activity have been shown in airways of patients with dysplastic lesions, in lesions adjacent to LUSCs, as well as in smokers at high risk for lung cancer ( Beane et al., 2019 ; Gómez-López, Whiteman, & Janes, 2021 ). Spatial gradients (i.e., gradients along macro-space regions with differing relative distances from the primary tumor) of allelic imbalance events were also reported in lung field of injury whereby defined chromosomal aberrations were more frequent in tumor-adjacent normal appearing (to the eye), relative to tumor-distant airways ( Jakubek et al., 2016 ; Humam Kadara et al., 2019 ). Single-cell analysis of the geospatially epithelial and immune ecosystem of early-stage LUADs revealed cellular lineages, states, and transcriptional features that gradually evolved across normal regions and with increasing proximity to LUADs ( Sinjab et al., 2021 ). The normal-appearing regions also harbored cells with features of malignant-enriched subsets and heterogeneous copy number variation profiles, possibly pointing towards molecular cancerized field effects or signifying mutagenic cellular clones that did not progress to cancer ( Sinjab et al., 2021 ). Field carcinogenesis could also partially explain the irreversible and continued risk to develop lung cancer among former smokers even after smoking cessation; mutagenized lung fields have a “mixture” of somatic mutations increasing the probability of transformation to a cancerized field across the lung. Further evidence on the relevance of cancerized field is relapse following curative surgeries where progressive transformation of tissues surrounding the excised tumor (rather than expansion of pre-existing cancer cells) can lead to relapse. These and other field cancerization observations may signify important features in the progression of phenotypically normal pre-neoplastic cells to cancer. A deeper understanding of the evolution of cellular clones in injury/cancerized fields could offer promising means to assess cancer risk and influence clinical directions, particularly in the space of cancer prevention. To understand how normal cells acquire these alterations as well as their implications, we provide a brief overview of the main concepts that have elucidated mutation acquisition, interrogating tissues with diverse mutational rates and patterns, giving rise to cellular clones that compete for “normal” tissue space, and we discuss how these concepts may impact our understanding of carcinogenesis. While most of our knowledge on somatic mutations and clonal expansion comes from our interrogation of pre-cancerous and tumor tissues, we further summarize a handful of landmark studies that were able to chart rare alterations in normal tissues, largely due to the advent of sensitive and deep sequencing technologies as well as cutting-edge mathematical modeling. We also show how these observations highlight the remarkable implications that field carcinogenesis studies can have on our understanding of preneoplasia, evolution, pathology and therapy outcomes.

Classically, cancer was perceived as a result of accumulating mutations in key genes over the years ( Fearon & Vogelstein, 1990 ). Recent advances in bulk sequencing approaches as well as genomic decoding at the single-cell level have revealed a surprisingly high mutational burden in normal somatic tissues which, for the most part, seems to be pathologically insignificant, prompting us to believe this could be the “new normal.” Making sense of those mutations to understand the transition from normal to malignant tissues is crucial for prevention, early detection, as well as the development of targeted pharmacotherapeutics to halt the progression of normal tissues to full-blown tumors. Despite being in its infancy, this branch of science reveals a new perspective for the pathobiology of cancer development where the accumulation of mutations alone is not sufficient to drive pathogenic transformation. Understanding the relevance of mutations in somatic tissues is evolving with technological advances. Scientists first believed that accumulating mutations are the root cause of cancers. Identification of mutations in normal tissues surrounding tumors and the ensuing field of cancerization led to new insights on how cells interact, how they evolve to survive, and how they impact the ability of early lesions to progress. Today, we are also learning that mutations are present in normal tissues irrespective of cancer development; normal tissues can harbor cancer-driving mutations and not have a higher rate of tumors. A deeper understanding of the mutational landscape of normal tissues, the clonal dynamics and the cellular interactions that ensue could lead to new ways to prevent the progression of expanded clones to cancer. The missing key is the ability to accurately determine the potential of cancer development from clones with specific mutations, since putatively mutated fields are abundant, but few are the ones that are actually tumorigenic. More recently, we are learning about clonal competition which might protect from early tumorigenesis rather than cause it. This challenges our current understanding of early tumorigenesis and may explain why the rate of cellular transformation to cancerous states is rare, despite the high frequency at which normal tissues accumulate harmful genetic mutations. Those ground-breaking findings are an invitation to reconsider our understanding of mutations in normal tissues and present an opportunity to revisit the definition of field of cancerization which was initially suggested to characterize mutated normal tissues. Those mutated fields may be in fact shields protecting us against early tumorigenesis. Clearly, there is a pressing need to identify biomarkers that can distinguish mutated fields that will remain indolent from those that will transform into cancers. Furthermore, the molecular mechanisms underlying clonal competition are yet to be elucidated. What makes a cell more competitive and able to eliminate neighboring cells? In the context of mutagen- or aging- induced somatic mutations, which alterations confer survival benefits? Are cells capable of evolving to select for those mutations that protect against tumorigenesis? Answering those questions would pave the way for better understanding of cancer evolution. Indeed, variation in cancer incidence across different tissues is well documented. While some of these differences are attributed to well-known risk factors such as smoking, ultraviolet exposure, alcohol use, or even familial predisposition, these hereditary and environmental dynamics cannot fully explain the differences in organ-specific cancer risk. As the endogenous mutation rate of all human cell types is nearly identical ( Tomasetti et al., 2013 ), it is plausible to predict that there should be a solid correlation between the total number of divisions of the normal self-renewing (stem) cells maintaining that tissue’s homeostasis and the lifetime risk of cancer development in that organ ( Tomasetti & Vogelstein, 2015 ). However, these intrinsic factors impacting cancer are further complicated by exogenous key players such as smoking (and the degree of it). In similar vein, the emergence of clones within normal tissues, their expansion, and their competitiveness (which depends on their accruing mutations and the microenvironment) are likely to be strongly associated with the degree of cell division in that particular tissue. In fact, half or more of the somatic mutations in cancers of self-renewing tissues originate prior to tumor initiation ( Tomasetti et al., 2013 ). Furthermore, a key aspect to be taken into consideration while examining the anti-tumorigenic potential of a clone depends on the regenerative and reparative capacity of the tissue. One such example is the facultative progenitor cell populations residing in the lungs that can be induced to proliferate and differentiate in response to various insults ( Kotton & Morrisey, 2014 ). This reparative response differs from that of other organs which exhibit a baseline high level of turnover via a population of undifferentiated stem cell populations such as the intestine, or that of organs with no regenerative capacity such as the brain and heart. Those various repair mechanisms and responses to injury are further complicated by an altered cellular response to injury associated with aging ( Poulose & Raju, 2014 ). All in all, the competitive advantage of a clone and its anti-tumorigenic potentials depend on an intertwined complex variety of factors that are yet to be deciphered. While we decode the dynamics of clonal competition to understand the factors that can favor an anti-tumorigenic clone, an open eye should be kept on the enemy. It is crucial to understand the factors that drive the pathogenic transformation of a clone into a malignant one. One supposition is related to the order of acquisition of mutations in a particular tissue. It is plausible that stochastic events leading to mutation accumulation can lead to different outcomes based on the order of these occurrences. It is also possible, that a certain mutation can drive a phenotype only if accompanied by a co-occurring culprit. The degree of exposure to an insult (continuous vs. intermittent exposure to a carcinogen) can also contribute to malignant transformation possibly by exhausting repair mechanisms and by means of topographical factors where strength is in numbers (many mutated cells are more potent than a solitary one). Furthermore, dampening of tissue repair mechanisms as a result of aging can also contribute to the tumorigenic potentials of a clone. Another provocative supposition is immune evasion which can lead to a positive selection of malignant clones. Moreover, the challenge in charting the evolution of a mutated field is further complicated by the presence of various exogenous and endogenous key players. For instance, a benign clone’s transition to acquire tumorigenic potential can be influenced by the microenvironment, whereby stromal reprogramming was found to be a key regulator of the surrounding epithelium. Hence, changes in the stroma (e.g., immune cell states and composition, fibroblast phenotypes, inflammatory molecules such as cytokines) can directly impact surrounding tissues ( Fig. 2 ). Studies on patients with inflammatory bowel disease and Barrett’s esophagus showed how inflammation can predict cancer risk ( Kaz, Grady, Stachler, & Bass, 2015 ; Nordenvall, Ekbom, Bottai, Smedby, & Nilsson, 2014 ; Prasad et al., 2007 ). Loss of Notch signaling in mesenchymal cells of transgenic mouse models was also shown to induce epithelial tumorigenesis (B. Hu et al., 2012 ). More recently, spatial characterization of co-existent clones in ductal in situ carcinoma tissues revealed characteristic differences between clones that share many genetic and all host features yet manifest divergent fates and clinical outcomes, with some bearing histology of ancestral stages despite carrying genetic signals of progression ( Lomakin et al., 2021 ). The stromal environment’s long-standing active role in driving tumor progression was not only visually confirmed in situ, but also further shown to be clone specific, a finding which could help explain why some, but not all lesions progress to a potentially lethal invasive cancer ( Lomakin et al., 2021 ). While the mechanisms underlying these dynamics are not fully understood, there is evidence that microenvironmental changes can enable field cancerization by altering the fitness of mutations in epithelial cells, thus promoting the expansion of cancerized clones ( Curtius et al., 2018 ; Davis et al., 2015 ; Gatenby & Gillies, 2008 ). The microenvironment provides selective pressure for epithelial cells which will have progenies with a genotype reflecting the phenotype of cells that successfully overcame the environmental challenge ( Curtius et al., 2018 ). Indeed, development of a malignant phenotype was shown to be mediated by overcoming one or more of six microenvironmental barriers: ischemia, acidosis, hypoxia, senescence, inadequate growth promotion, and apoptosis with loss of basement membrane contact ( Gatenby & Gillies, 2008 ). Whether there is clonal competition beyond the epithelial compartment is yet to be elucidated. There is a pressing need to better understand the interplay between the epithelium and the microenvironment. For example, is clonal expansion a direct result of the crosstalk between two biologically and functionally distinct cellular compartments? How does clonal hematopoiesis – i.e., the formation of genetically distinct populations of blood cells as we age - affect the function of the immune system and inflammation, and hence, impact clonal competition and expansion? One of the key advantages of understanding the relevance of mutations in normal somatic tissues is our ability to develop preventive therapies that can halt the transition from normal to diseased state. An elegant example of that is the use of nonsteroidal anti-inflammatory drugs (NSAIDs) to reduce the risk of cancer development in patients with Barrett’s esophagus ( Kostadinov et al., 2013 ). It could be hypothesized that by modifying microenvironmental cues, NSAIDs impact the fate of certain clones and drive clonal competition to adopt alternative outcomes ( Galipeau et al., 2007 ). Through a similar mechanism of action, 5-aminosalicylate anti-inflammatory drugs (e.g., aspirin) which reduce the risk of colorectal cancer development in inflammatory bowel disease patients, could modulate the evolution of mutated fields by establishing a microenvironment that favors non-malignant clones to preside ( Rothwell et al., 2011 ; Zhao et al., 2014 ). Furthermore, Bacillus Calmette–Guérin therapy prescribed for patients with superficial bladder cancer may function by modulating the interactions between the mutated clones and the microenvironment ( Gee, Sabichi, & Grossman, 2002 ; Herr et al., 1995 ; Lamm et al., 2000 ; Redelman-Sidi, Glickman, & Bochner, 2014 ). Along those lines, it is crucial to understand the mechanisms by which some clones are selected for, and thus, identify mutations that could increase fitness in the face of other clones with malignant potential. For instance, pro-inflammatory signals within the microenvironment can abrogate cellular competition and clearance of mutant cells, whereas anti-inflammatory treatments can sway the competitive advantage towards normal cells ( D’Ambrogio et al., 2022 ). The overarching goal would be to identify markers for early detection and targets for prevention. In conclusion, normal somatic tissues experience silent battlefields between mutating clones. While competitive interactions are governed by the differences in fitness between neighboring cells, external factors can impact the outcomes of the game. Decoding the dynamics of clonal competition and the factors contributing to diverging fitness levels presents an understudied avenue that can be leveraged to tilt the odds towards certain clones, and thus, protect the species from tumorigenesis.

The accumulation of mutations in normal somatic cells has been demonstrated across various tissues including bladder, appendix, pancreas, prostate, stomach, bile duct, endometrium, adipose tissue, kidney, muscle, esophagus, blood, skin, lung, small and large intestines, and liver ( Brunner et al., 2019 ; Franco et al., 2018 ; Franco et al., 2019 ; Grossmann et al., 2021 ; Lawson et al., 2020 ; Lee-Six et al., 2018 ; Lee-Six et al., 2019 ; Mitchell et al., 2022 ; Moore et al., 2020 ; Moore et al., 2021 ; Osorio et al., 2018 ; Park et al., 2021 ; Yokoyama et al., 2019 ; Yoshida et al., 2020 ). It is important to remember that the mutations arise in phenotypically normal-appearing tissues, meaning in cells that maintain their normal functions. Therefore, from an “outsider’s” perspective, it would seem that order is in place and homeostasis is maintained while instead, a looming “silent” battle of clones is clearly underway. With this in mind, it is logical to assume that the significance of genetic alterations ought to be always studied in a context dependent manner; the functions of genetic mutations in normal tissues are likely to be different from the roles of these same alterations when seen and described in frank malignancy. Simply put, cancer mutations could be classified as either drivers that confer survival growth to tumor cells within a permissive microenvironment, or passenger mutations which passively accumulate in somatic cells ( Calabrese, Tavaré, & Shibata, 2004 ; Greaves, 2015 ; Stratton, Campbell, & Futreal, 2009 ). In normal cells, one way of characterizing mutations could be based on their ability to confer phenotypic changes that underpin the formation of a cancerized field. Despite the “chaotic” nature of the battleground events, the final outcome will be determined based on the fitness of the clone that presides, which is itself primarily dictated by its acquired genomic alteration(s) that discriminate it from the rest of the clones. One can expect that mutations occurring in the normal-appearing field may have one (or more) of the following consequences: 1) - negatively selected mutations (majority), 2) selectively advantageous mutations (minority) that are retained as long as the mutations subsequently acquired by cellular clones exhibit additional advantages that can trigger substantial clonal expansion ( Abbosh, Venkatesan, Janes, Fitzgerald, & Swanton, 2017 ), and 3) neutral or even mildly disadvantageous mutations that can be spared or even undergo some expansion due to genetic drift ( Marusyk & Polyak, 2010 ) ( Fig. 2 ). If “proto-cancer” mutations in the field accumulate additional events capable of contributing to the carcinogenic process, field cancerization can be equated with initiation of tumorigenesis. This has been strongly supported by the identification of premalignant fields in diseases previously known to predispose tissue to cancer, such as Crohn’s disease, ulcerative colitis and Barrett’s esophagus ( Baker, Graham, & Wright, 2013 ; Graham, McDonald, & Wright, 2011 ). One expected consequence of this phenomenon is that, while a fully malignant tumor is enriched with the advantageous mutation(s), other regions within the tumor or in the adjacent normal-appearing and damage-exposed field may contain alternate mutational patterns which can be used to backtrack the evolution of the tumor from the premalignant field. In this way, field carcinogenesis consorts with the multiclonal evolution of tumors. For instance, by sequencing multiple regions across lung tumors, the accumulation of mutations was found to adopt a branched evolutionary model whereby early “founder” mutational events present in every tumor region constitute trunk mutations and are comprised of the majority of canonical cancer mutations, whereas subsequent spatially confined mutations are represented as “branch” events acquired later during lung carcinogenesis ( de Bruin et al., 2014 ; Zhang et al., 2014 ). Importantly, the expanded clone (with advantageous mutations) will also carry passenger mutations, which are expected to be overrepresented in the mosaic of clones in terms of their frequency despite their “passive” nature, i.e., they are not driving the main event(s) that caused this clone to outgrow. Therefore, the bumped-up frequency of such passive mutations ought to be interpreted with caution because it is a mere consequence of serendipitously existing in positively selected clone and being “dragged up along” in its cells as they expand. Furthermore, tissue damage may incur changes that alter the tumor microenvironment, including extra-cellular matrix and immune contexture, further exacerbating the effect of genetic instability ( Hittelman, 1999 ). Also, evidence of molecular heterogeneity, whether across clonal outgrowths (e.g. higher frequencies of genetic modulations seen in dysplastic lesions relative to normal-appearing and metaplastic lesions ( Gutierrez-Gonzalez et al., 2011 )), as well as between the clonal outgrowths and the primary tumor (e.g. different loss of heterozygosity patterns ( Park et al., 1999 )) signifies that clonal patches may adopt distinct multistep tumorigenesis pathways such as in the lung cancerization field. One possibility is that lesions become progressively more heterogeneous as they advance into late-stage disease ( Jamal-Hanjani et al., 2017 ; Reuben et al., 2017 ; Zhang et al., 2014 ). Other observations showing higher intra-tumor heterogeneity (subclonal and branch mutations) among early pre-neoplastic lesions compared to advanced-stage lesions support a clonal sweep model, whereby fit subclones selectively outgrow other subclones during initiation and early progression of lung preneoplasia ( Hu et al., 2019 ). Overall, these scenarios show that despite being phenotypically normal, cells can accrue mutations that could lead to clonal outgrowths. In the ensuing cancerized field, the normal-appearing tissue may thus comprise a battleground for “silently” competing clones. In the next section, we provide an overview of the large spectrum of possible mechanisms of the competition and its outcomes, mainly based on studies interrogating cellular competition in non-malignant settings (e.g., development, experimental genetic mosaicism). Evolutionarily speaking, somatic cell competition describes how cells with higher fitness outcompete neighboring cells with lower fitness. Cell competition was first described in Drosophila melanogaster harboring mutations in ribosomal genes ( Lambertsson, 1998 ; Marygold et al., 2007 ). The homozygous mutations are lethal, while heterozygotes can survive in a genetically mosaic environment, albeit with a slower developmental rate when compared to the wild-type counterparts. In fact, heterozygote cells were found to be eliminated by caspase-dependent apoptosis induced by the wild-type cells in drosophila. As a result, a single wild-type cell is capable of colonizing an entire developmental compartment of heterozygotes ( Marygold et al., 2007 ). Cell competition was then adopted to describe the phenomenon of context-dependent active elimination of intrinsically viable cells via short range interaction with distinct neighboring cells, as opposed to passive clonal contraction or expansion ( Kale, Li, Lee, & Baker, 2015 ; Lambertsson, 1998 ; Li & Baker, 2007 ; Morata & Ripoll, 1975 ; Moreno, Basler, & Morata, 2002 ; Simpson & Morata, 1981 ). The existence of an active cell competition process was further validated by the discovery of “super-competitors”, such as Drosophila melanogaster expressing elevated levels of Myc (the mammalian homologue of which is an oncogene), which are themselves able to eliminate nearby wild-type cells. Subsequent studies in Drosophila melanogaster showed similar competitive cell elimination induced by other factors, such as loss of apicobasal cell polarity ( Brumby & Richardson, 2003 ; Igaki, Pastor-Pareja, Aonuma, Miura, & Xu, 2009 ), or differences in the activity signaling pathways such as Wnt/Wg ( Vincent, Kolahgar, Gagliardi, & Piddini, 2011 ) and JAK-STAT ( Ballesteros-Arias, Saavedra, & Morata, 2014 ; Enomoto & Igaki, 2013 ; Menut et al., 2007 ; Moberg, Schelble, Burdick, & Hariharan, 2005 ; Tamori et al., 2010 ; Thompson et al., 2005 ; Vaccari & Bilder, 2005 ; Vidal, Larson, & Cagan, 2006 ). Cell competition was further demonstrated in other model systems such as mammalian cells cultured in vitro, zebrafish and mice ( Chiba et al., 2016 ; Clavería, Giovinazzo, Sierra, & Torres, 2013 ; Clavería & Torres, 2016 ; Di Gregorio et al., 2016 ; Ellis et al., 2019 ; Hogan et al., 2009 ; Kajita et al., 2010 ; Kale et al., 2015 ; Li & Baker, 2007 ; Maruyama & Fujita, 2017 ; Moreno et al., 2002 ; Norman et al., 2012 ; Tamori et al., 2010 ). Liver repopulation following donor liver cells transplant is among the first descriptions of cell competition in mammals. Following the introduction of donor fetal stem and progenitor cells into a rat liver, apoptosis of surrounding hepatocytes occurs including human α1-anti-trypsin-mutants and replacement with donor tissues take place. Intriguingly, this active process did not occur in the absence of the transplanted cells ( Ding et al., 2011 ; Oertel, Menthena, Dabeva, & Shafritz, 2006 ). In addition to apoptosis-mediated elimination introduced above, several other mechanisms have been involved in cell-cell competition. For instance, autophagy eliminates cardiomyocytes with lower Myc expression ( Villa Del Campo, Clavería, Sierra, & Torres, 2014 ). Furthermore, oriented cell division directing stratified epithelial cells with less Myc ( Ellis et al., 2019 ) or more p53 ( Fernandez-Antoran et al., 2019 ) leading to terminal differentiation and sloughing. Also, cell-cell competition can be fueled by differences between cells which themselves are identifiable due to specific molecular recognition signals that enable competing cells to label other clones as foreign ( Bilder, Li, & Perrimon, 2000 ; Brumby & Richardson, 2003 ; Ohsawa et al., 2011 ; Sanaki, Nagata, Kizawa, Léopold, & Igaki, 2020 ; Vaughen & Igaki, 2016 ; Yamamoto, Ohsawa, Kunimasa, & Igaki, 2017 ). Alternatively, if the normal balance between cell death and survival is disrupted, one cell population will be facing a more hostile environment and will be less likely to preside, in which case cell competition would have occurred without the need for specific molecular recognition events ( Alpar, Bergantiños, & Johnston, 2018 ; Germani, Hain, Sternlicht, Moreno, & Basler, 2018 ; Meyer et al., 2014 ). Moreover, differential growth rates across diverse cell populations can generate mechanical stresses which might affect cell survival, proliferation and mobility, and lead to cell competition ( Aegerter-Wilmsen et al., 2010 ; Eisenhoffer et al., 2012 ; Gudipaty et al., 2017 ; Levayer, Dupont, & Moreno, 2016 ; Levayer, Hauert, & Moreno, 2015 ; Li, Kale, & Baker, 2009 ; Matamoro-Vidal & Levayer, 2019 ; Moreno, Valon, Levillayer, & Levayer, 2019 ; Puliafito et al., 2012 ; Shraiman, 2005 ; Simpson, 1979 ; Wagstaff et al., 2016 ). The immune system can be the ace in the hole in the game of clones, and this has been demonstrated in the success of immunotherapies as well as the prognostic value of immune infiltrates ( Cercek et al., 2022 ; Galon et al., 2014 ; Sharma & Allison, 2015 ). When cancers accumulate mutations, they acquire neoantigens – novel peptides that elicit an immune response- and consequently, they undergo evolutionary selection ( Fig. 2 ). In a process called “immune escape”, tumor cells can evolve to block the immune system’s ability to recognize or react to their neoantigens, leading to positive selection ( DuPage, Mazumdar, Schmidt, Cheung, & Jacks, 2012 ). The clonality of neoantigens in a growing cancer can also be shaped by negative selection. Neoantigens that are recognized by the immune system will cause the clones bearing them to expand less and disproportionately produce fewer offspring, thereby undergoing negative selection ( Lakatos et al., 2020 ). While these processes are mostly studied in the context of overt cancer, the neoantigen landscape in normal-appearing phases, i.e., in normal tissues, and whether the immune system is implicated in the “silent” battle of clones, are concepts that remain largely unexplored. It is worth noting that while the majority of the afore-mentioned examples of cellular competition were described in the context of experimental genetic mosaicism, normal somatic cells are also capable of accumulating mutations with age and are thus a potential site for endogenous genetic mosaicism that can result in clonal selection and competition-like population swings ( Martincorena & Campbell, 2015 ). For instance, studies have aimed to understand the contributions of somatic mutation rates to mutational processes and clonal composition changes in human tissues with age and disease ( Blokzijl et al., 2016 ; Lee-Six et al., 2019 ; Martincorena et al., 2015 ; Welch et al., 2012 ). New data from comparative analyses of somatic mutagenesis across species revealed mutational processes that were shared across mammals, suggesting that somatic mutation rates are evolutionarily restricted and may contribute to aging ( Cagan et al., 2022 ). Interestingly, differences in mutational rates are also seen during development, whereby somatic mutation rate is higher in fetal cells relative to post-natal stages ( Bae et al., 2018 ; Baron & van Oudenaarden, 2019 ; Ju et al., 2017 ; Kuijk et al., 2019 ; Mitchell et al., 2022 ; Osorio et al., 2018 ; Park et al., 2021 ; Spencer Chapman et al., 2021 ; Vanneste et al., 2009 ). For instance, fetal liver, intestine and hematopoietic stem cells accumulate mutations five times faster than their adult counterparts ( Bae et al., 2018 ). Similarly, fetal mitotic neuronal progenitor cells mutational rate was shown to be many-fold higher compared to that in post-natal tissues ( Abascal et al., 2021 ; Bae et al., 2018 ; Luquette et al., 2021 ). It is highly probable that the high somatic mutational rate in fetal tissues is due to increased mitotic activity during embryonic development compared to later stages. Further attempts to characterize mutational rates in adult somatic cells showed tissue-specific patterns, whereby mutations accumulating in tissues varied between 9 substitutions per year in bile ductular cells and up to 56 substitutions in appendiceal crypts ( Moore et al., 2021 ). Of note, and compared to somatic cells, mutation rate is much lower in germ cells whereby female and male germ cells acquire as little as 0.74 and 2.7 mutations per year ( Jónsson et al., 2017 ; Moore et al., 2021 ; Rahbari et al., 2016 ). When reproduction is accomplished, energy invested in somatic maintenance starts to wane ( Kenyon, 2010 ; Longo, Mitteldorf, & Skulachev, 2005 ), and aging manifests as a constellation of stochastic events occurring in somatic cells ( Kenyon, 2010 ). These events include but are not limited to mutations affecting DNA damage repair pathways ( Tiwari & Wilson 3rd., 2019 ) or tightly controlled gene-regulatory mechanisms in the cells, thereby increasing transcriptional heterogeneity among cells in isogenic populations, a phenomenon known as transcriptional noise ( Urban & Johnston, 2018 ; Vijg & Dong, 2020 ). While accruing somatic mutations in tissues could lead to widespread cellular dysfunction, particularly with increased mutational burden ( Martincorena & Campbell, 2015 ), the majority of somatic alterations are benign. One study by Robinson et al. showed that individuals with defective DNA proofreading due to germline mutations in POLE/POLD1 do not exhibit overt features of premature aging or early onset of age-related, non-neoplastic disease, and many survive into the late decades of the standard human lifespan, despite bearing a high mutation somatic burden starting at early embryogenesis and a high predisposition to familial cancers ( Robinson et al., 2021 ). All in all, given the heterogeneity of cellular malfunction caused by high somatic mutational burden, there is an urgent need to better elucidate the mechanisms tightly controlling the consequences of progressive accumulation of mutations in normal-appearing tissues and throughout life. Whether somatic alterations were caused by endogenous or exogenous mutagenic insults, backtracking the consortium of mutations that accumulated over the lifespan of a cell and that were passed on to their progeny – potentially leading to the rise of different clones – can be indicative of the specific initiating event. Indeed, somatic mutations leave a characteristic genomic imprint — a mutational signature — which is defined by the type of DNA damage and DNA repair processes that result in deletions, insertions, base substitutions, or structural alterations ( Helleday, Eshtad, & Nik-Zainal, 2014 ). Indeed, specific mutational processes are already known to leave characteristic gene expression signatures in the mutational catalogs of cancer cells ( L. B. Alexandrov, et al., 2013 ; Alexandrov, Nik-Zainal, Wedge, Campbell, & Stratton, 2013 ; Helleday et al., 2014 ; Nik-Zainal et al., 2012 ). Studies have also demonstrated specific mutational patterns in normal somatic cells. Clock-like mutational signatures (SBS1 and SBS5) which were also described in most adult tissues, characterize the earliest embryonic mutations ( Ju et al., 2017 ; Park et al., 2021 ; Spencer Chapman et al., 2021 ). During development, it has been shown that specific mutation patterns in adult intestinal cells were evident in fetal intestinal cells as early as 13 weeks after fertilization ( Kuijk et al., 2019 ). In contrast, mutation patterns of liver or hematopoietic stem cells are different before versus after birth ( Hasaart et al., 2020 ; Kuijk et al., 2019 ). SBS18 was highly evident in fetal liver stem cells and fetal neural progenitors; this signature is known to be associated with oxidative stress-induced mutagenesis ( Bae et al., 2018 ; Kucab et al., 2019 ; Pilati et al., 2017 ). Therefore, a better understanding of mutational signatures of normal tissues at different stages of life is highly warranted to fill major gaps in our understanding of how normal cells evolve during aging or progress to cancer. Irrespective of how clones arose, their mode of recognition of other clones and their battle to avoid selective elimination, a change in survival that is dependent on differences between cells remains the main distinctive feature of cell competition. Elucidating the molecular mechanism of cell competition has been extremely informative as mutations in oncogenes and tumor suppressor genes ( Hanahan, 2022 ) often underscore cell competition. Decoding cellular competition in normal cells is key to improving our understanding of underlying mechanisms that could be leveraged for early prevention and treatment of various cancers ( Baker, 2020 ). The molecular rules that control cell competition and fate in normal tissues are yet to be fully elucidated. Along the same lines of super-competitors in Drosophila melanogaster with homologous mammalian oncogenes, it is conceivable to infer that mammalian tumor cells may be super-competitors when compared to neighboring normal cells, and the outcome of this competition is tumor expansion. Tumor-suppressive roles of cellular competition ought not to be underrated as well ( Baker, 2020 ), as epithelia may distinguish and expel potentially transformed single cells in a process termed “epithelial defense against cancer” ( Yamauchi & Fujita, 2012 ). In the following section, we will summarize snapshots of competing clonal communities that have been reported in normal solid tissues, and which hint towards mechanisms that enable the fittest clones to expand and eliminate their less “fit” neighbors. Using RNA sequencing databases from more than 6700 histologically healthy samples encompassing 29 tissue types from 500 individuals, a study by Yizhak et al. found multiple somatic mutations associated with cancer, as well as macroscopic clones of mutated cells in normal tissues. The number of somatic mutations, which was reportedly the highest in skin and esophageal epithelia, was positively associated with age and cell proliferation rates. Unsurprisingly, the skin and lungs were among the tissues with the highest mutational burden, owing to their frequent exposure to environmental mutagens such as UV and smoke, respectively ( Yizhak et al., 2019 ). Using mathematical models, another study found that more than half of somatic mutations found in cancer were already present in the founder clone – i.e., the normal cell from which the cancer was initiated ( Tomasetti et al., 2013 ). Thus, it is likely that these somatic mutations would have been present in the normal somatic cells, whether or not the cells progressed to give rise to subsequent tumors ( Fiala & Diamandis, 2020 ). High mutational burdens in normal somatic cells of the skin were further demonstrated in another study which found that 18 to 32% of skin cells from healthy individuals carry driver mutations of cutaneous squamous cell carcinomas and are under strong positive selection (at a density of ~140 driver mutations per square centimeter), while still exhibiting normal physiological functions ( Martincorena et al., 2015 ). Fowler et al. demonstrated that the selection of mutant clones in normal human skin varies with body site, which paints a patchwork-like portrait of mutant cells. Intriguingly, the variation in cancer risk between sites substantially exceeded the differences in the densities of mutant clones across the patches. Thus, it became more tempting to refute the hypothesis that carcinogenesis is directly attributed to the mutations observed in the normal patches ( Fowler et al., 2021 ). Clonal competition and maintenance of physiological integrity are tightly linked, but the mechanism governing these dynamics is far from being a one-sized shirt that fits all. In one study, p53 mutant progenitors in the murine epidermis outcompeted wild-type cells due to their highly proliferative dynamics. Yet, this effect was transient and physiological homeostasis was later re-established. Intriguingly, while exposing those mutant clones to low-dose UV light drove a short-term expansion of the p53 mutant clones, long-term exposure generated other mutant and superiorly fitter clones that outcompeted the short-lived expansion of p53 mutant cells ( Murai et al., 2018 ). Further investigation of morphologically normal tissues has led to unexpected and interesting findings. For instance, approximately 1% out of 2000 normal colorectal crypts in middle-aged individuals were found to carry a driver mutation ( Lee-Six et al., 2019 ). In fact, the incidence of somatic mutations in normal colonic crypts was higher than that in colorectal cancer (1 in 375,000 and 1 in 3 million chance for an adenoma and carcinoma to form, respectively), even though the mutational burden is higher in the latter. This alludes to the notion that the neoplasms are rare incidences resulting from a pervasive process of alterations that occur in morphologically normal colorectal epithelium ( Lee-Six et al., 2019 ). Moreover, mutations in proto-oncogene and tumor suppressor driver genes ( KRAS, PIK3CA, PTEN, ARID1A , and TP53 ) were found in 19 out of 24 women with endometriosis, a benign condition of the female reproductive system which rarely leads to cancer ( Anglesio et al., 2017 ; Suda et al., 2018 ). Tp53 mutations, albeit at low frequency, were also found in peritoneal fluid and blood of cancer-free women ( Krimmel et al., 2016 ). Those results are in line with findings from another study interrogating endometrial cancer development from uterine lavage fluid and identifying cancerous driver genes in more than 50% of cancer-free women ( Nair et al., 2016 ). By whole genome sequencing, Moore et al. also showed that the burden of driver mutations in cancer genes and occurring in normal endometrial glands decreases with parity and increases with age ( Moore et al., 2020 ). Cell clones carrying driver mutations tend to appear early on in life and may progress to colonize the endometrial epithelium ( Moore et al., 2020 ). In the context of exposure to occupational or environmental carcinogens, few studies have outlined possible clonal mechanisms in normal tissues. In one study, Lawson et al. revealed a rich landscape of mutational processes and clonal selection in normal urothelium. The authors found extensive heterogeneity across clones and individuals, suggesting exposure to a wide range of mutagens, including smoking, a known risk factor for bladder cancer ( Lawson et al., 2020 ). Yoshida et al. interrogated the effects of tobacco smoke on the normal human bronchial epithelium ( Yoshida et al., 2020 ). Unsurprisingly, tobacco smoking increased mutational burden in normal bronchial epithelial cells. Like urothelial tissues, significant cell-to-cell and inter-individual heterogeneity was noted. It was also shown that quitting smoking enables mitotically quiescent cells, which have evaded tobacco-induced mutagenesis, to renew the bronchial epithelium ( Yoshida et al., 2020 ). Somatic mutations in normal esophageal tissues have been especially a hot topic of investigation. Martincorena et al. found several mutations in cancer-associated genes in esophageal tissues from healthy donors. Of note, those genes harbored far more mutations in normal somatic esophageal tissues relative to healthy skin. Furthermore, many of those mutations were positively selected, underwent increased cell proliferation, and subsequently formed dominant esophageal cell clones ( Martincorena et al., 2018 ). It was also noted that despite being seen in cancer genomes, the majority of those mutations, such as the ones affecting the classical oncogene NOTCH1 , are not particularly evident in esophageal carcinomas ( Martincorena et al., 2018 ). Of note, esophageal tissues have a naturally high mutational frequency, which could partially explain the above observation ( Higa & DeGregori, 2019 ; Martincorena et al., 2018 ). Exposure to numerous mutagens through human diet could also account for high mutational burden in the esophagus. In both normal esophageal and skin tissues, mutations in NOTCH1, TP53, NOTCH2, FAT1, NOTCH3 , and ARID1A were reported ( Martincorena et al., 2018 ). While TP53 is the most common mutated gene in esophageal squamous carcinoma (Kim J et al. Nature 2017), it is far less prevalent when compared to NOTCH1 in normal esophageal tissues ( Martincorena et al., 2018 ). Similar findings were observed by Yokoyama et al. which reported many age-related mutational signatures (predominantly in NOTCH1 ) in esophageal tissues from healthy donors ( Yokoyama et al., 2019 ). Clones carrying driver mutations arise multifocally during early childhood and expand in size and number as they age. Eventually, and in the extreme elderly, these clones replace the entire esophageal lining. Interestingly, mutations in NOTCH1 and PPM1D were found to be exceedingly over-represented in normal appearing esophageal epithelial cells, relative to mutations in esophageal cancer ( Yokoyama et al., 2019 ). Altogether, these findings beg a need for deeper investigation of the “rules of the battle” that govern clonal competition in normal tissues. Studies done on normal tissues, such as the ones described above, can elucidate how competing clones expand and eliminate their less-fit neighbors. Yet, much less is known on how dynamic clonal competition in the normal epithelium may impinge on early tumorigenesis, perhaps mostly due to the technical complexity inherent to tracing the rise of competing clones and their fates with time. As described above, patterns of shared alterations between normal-appearing tissues in cancerized fields and tumors of the same tissue type propose a high degree of relevance of mutations observed in normal tissues to possible pathologic transformation. A handful of studies utilized elegant models to try to deconstruct mutational landscape in normal tissues and clonal competition dynamics that might precede, or even prevent, tumor formation. One study showed that competition between normal pancreatic cells and those carrying the key mutation driving human pancreatic tumors, Kras G12D , resulted in elimination of Kras G12D cells and restoration of tissue homeostasis ( Batlle & Wilkinson, 2012 ). The authors further pinpointed that this clearance is dependent on functional EPHA2, a receptor tyrosine kinase that regulates cell proliferation, survival, and cell-cell adhesion at tissue boundaries ( Batlle & Wilkinson, 2012 ). In contrast, persistence of Kras G12D cells accelerated the appearance of premalignant lesions. In another report, Martins et al. showed that the process of cell competition itself is a tumor suppressor in the thymus ( Martins et al., 2014 ). Murine T lymphocytes turnover from bone-marrow-derived progenitors depended on the natural cell competition between “young” bone-marrow-derived and “old” thymus-resident progenitors, which execute differential gene expression programs ( Martins et al., 2014 ). Abrogation of this cell competition was shown to cause self-renewal of progenitor cells, upregulation of the transcriptional master regulator Hmga1 (known to be overexpressed in stem cells and in multiple cancers, such as human T-cell acute lymphoblastic leukemia, ( Di Cello et al., 2013 )), transformation, and development of T-cell acute lymphoblastic leukemia ( Martins et al., 2014 ). To better understand the processes and rules of clonal competition, Colom and colleagues investigated mutant clones in the esophageal epithelium of mutagen-treated mice. This landmark study uncovered an anti-tumorigenic role of clonal competition in histologically normal esophageal tissues ( Colom et al., 2021 ). Following exposure to diethylnitrosamine (DEN) (a mutagen found in tobacco smoke), the murine esophagus became a patchwork of mutant clones with high mutational burdens, while still maintaining normal esophageal structure and function. Hundreds of micro-tumors were detected as early as 10 days post-exposure, with most disappearing as soon as they emerged. Persistent micro-tumors later developed into malignancy. A closer look revealed that 10-day post exposure to DEN, tumors were polyclonal, in contrast to the monoclonal tumors observed at 12-months, thereby suggesting that clonal competition drives the selection of specific mutant clones. The authors show that clonal competition is the main mechanism for elimination of “loser” micro-tumors, whereby only micro-tumor clones carrying additional advantageous mutations would persist and expand. Interestingly, this was observed irrespective of the immune system’s activity, as the phenomenon occurs in both immunodeficient and wild-type controls and to the same extent. Furthermore, lineage tracing in transgenic mouse models revealed strong clonal competition that evolved with time. Many mutant clones harboring mutations in Notch1, Notch2 and Trp53 were under positive selection. Further investigation showed that for the elimination of unfit clones to occur, “loser” micro-tumors were surrounded by histologically normal epithelium composed of mutant clones with Notch1 mutations being the most common. This suggests that loss of functional Notch in the normal epithelium protects from early tumorigenesis caused by exposure to DEN. This is further validated by exposing DEN-treated mice to dibenzazepine (DBZ) – a Notch signaling inhibitor, thereby increasing the relative fitness of all cells. Indeed, DBZ treatment significantly decreased tumor loss, leading to higher tumor survival as it effectively eliminated competition between tumor cells and their surrounding epithelium. The authors speculated that the expansion of mutant clones depends on the “fitness” of the surrounding tissues, whereby the competitive advantage of a mutant clone is dependent on the genotype of its neighboring tissues. Thus, the clones will continue to expand and eliminate less fit wild-type cells and until they encounter other mutant clones of equivalent fitness, at which point the clones revert to homeostasis and clone growth is restricted. They further reveal more complex scenarios, whereby that “triumphant” clones were able to expand more when they bordered less-fit cells, relative to when they were surrounded by other equally-fit and “winner” clones ( Clyde, 2020 ; Colom et al., 2020 ). All in all, this study elegantly highlights the role of expanding mutant clones in outcompeting newly formed tumors, and that tumors are able to persist if they are armed with a mutation that confers a competitive advantage over their neighbors ( Colom et al., 2021 ).

As we age, somatic mutations accumulate in response to endogenous and/or exogenous factors ( Milholland, Auton, Suh, & Vijg, 2015 ; Risques & Kennedy, 2018 ). Some of these mutations will modulate cellular phenotypes (e.g., enable faster growth) and lead to development of early lesions that could further evolve into cancer, that is, if the conditions favor development of tumors ( Di Giacomo et al., 2017 ; Lenz et al., 2021 ; Robert et al., 2018 ). As a consequence of this evolutionary model of cancer development, tumors are not “monolithic”, meaning that the tumor cells themselves are not genetically identical but rather consist of genetically distinct cellular clones, or a group of identical cells that share a common ancestor (Mel Greaves & Maley, 2012 ; Naugler, 2010 ; Parsons, 2018 ). The accumulation of somatic mutations during cycles of cell division has been thoroughly studied in the context of tumor development ( Hao & Wang, 2016 ; Takeshima & Ushijima, 2019 ). Yet, our focus on tumor tissues at advanced stages limits our knowledge of the earliest stages of cancer evolution and overlooks valuable clinical opportunities for early detection and prevention of malignancy. As such, there has been an increasing interest in decoding the genomic landscape of normal tissues in stages that precede cancer formation, which for the most part constitute an unknown history of the tissue that cannot be directly discerned by simply looking at the present-day tumor. There is a strong need for an in-depth understanding of the “secret” history of the clones that lead to the checkerboard pattern observed in cancer. Mathematical modeling has helped identify evolutionary dynamics producing this pattern by characterizing clonal mutations which are “public”, meaning that they are found in all cancer cells of a particular tumor, versus subclonal mutations, which are “private”, low in frequency, and exist in some but not all tumor cells, thereby implying that the pattern of the latter could provide a window into the secret history of the ensuing clones ( Dentro et al., 2021 ; Errico, 2015 ; McGranahan & Swanton, 2017 ; Tomasetti, Vogelstein, & Parmigiani, 2013 ). These concepts are also elegantly portrayed in a number of epithelial tissues that undergo a phenomenon called “field cancerization’, whereby cancers were found to arise from within a clonal region of normal-appearing epithelium ( Braakhuis, Tabor, Kummer, Leemans, & Brakenhoff, 2003 ; Slaughter, Southwick, & Smejkal, 1953 ) ( Fig. 1 ). Intact histological and normal cellular function were no longer grounds for dismissing the potential presence of processes that could underlie tumorigenic mutation and transformation. As a result, there has been a growing interest in understanding mechanisms that control the rise of mutant clones in normal tissues, and their tendency to compete for space and survival in tissues. Clonal dynamics, including the rise of clones, their competition and expansion, were found to be tightly linked to the inception of mutations in normal cells and the fates dictated by these alterations ( De Dominici & DeGregori, 2021 ; Olmeda & Ben Amar, 2019 ; Park et al., 2021 ). Since mutations are transmitted to progeny, clones are likely to arise and compete with one another within normal somatic tissues. Competition results in elimination of less “fit” cells, allowing others to expand, physically breach through epithelial space, and be crowned the “winner” or “triumphant” clone ( D’Ambrogio, Hill, & Hogan, 2022 ; Naxerova, 2020 ). Even though such “battlefields” have been frequently observed in normal tissues, only few clones progress to develop neoplasms; the likely overall turnout remains re-establishment of tissue homeostasis, despite the high mutational burden ( Clavería & Torres, 2016 ; D’Ambrogio et al., 2022 ; Di Gregorio, Bowling, & Rodriguez, 2016 ; Kajita & Fujita, 2015 ). Therefore, characterizing the clones, the relationships between their mutations and their fate, and how their competition may impinge on the transition from normal to cancerous tissues, is crucial to improve our search for targets for early detection and prevention, and thus, to abrogate tumor progression. Despite its clinical significance, investigating mutations in normal tissues requires innovative approaches that will enable current sequencing modalities to detect rare and subtle changes in the genome. Recent advances have enabled researchers to detect mutations in normal tissues for example, by studying in vitro-expanded clones, in-depth sequencing of microscopic biopsies that carry a limited number of clones, utilizing sequencing platforms with high capacity for error correction, sequencing DNA from both strands (duplex sequencing) to detect rare mutations, or decoding the genome at the single-cell level ( Fiala & Diamandis, 2020 ; R. Li et al., 2021 ). Mapping of somatic mutations accumulating in normal somatic cells, as well as longitudinal tracing of those alterations in aging tissues, became more achievable. Mutation accumulation was also studied in the context of exposure to exogenous factors such as smoking and UV exposure, and in inflammatory states such as inflammatory bowel diseases ( Martincorena et al., 2015 ; Nanki et al., 2020 ; Yoshida et al., 2020 ). Much of these findings have been catalogued in public databases specific for somatic mutations in human normal samples ( Miao, Li, Wang, Zheng, & Cai, 2019 ; Sun, Wang, Maslov, Dong, & Vijg, 2021 ). In this review, we will provide a brief history of field cancerization and its implications on understanding early stages of pathogenesis and deviation from the pathologically “normal” state. The review will provide an overview of how mutations accumulating in normal tissues can lead to patchwork of mutated cell clones that compete while maintaining an overall state of functional homeostasis. The review also explores the role of clonal competition in directing the fate of normal tissues and summarizes multiple mechanisms which have been linked to cancer development. Finally, we highlight the importance of understanding mutations in normal tissues and the role of clonal competition (in both the epithelium and the microenvironment) and their significance in exploring new approaches to combatting cancer.