Rare diseases are disorders that affect less than one case in 2000 people, i.e., only a small percentage of the population. However, there are more than 6000 known rare diseases, affecting over 300 million people worldwide (Boycott et al., 2017; The Lancet Diabetes Endocrinology, 2019). In 80% of the cases, the origin of a rare disease is one or multiple disadvantageous inherited variations of the genome (Wang et al., 2021). These are present in all cell types of the affected individual. Nevertheless, most rare diseases, which are not already prenatally lethal, are rather tissue specific. Pediatricians are more likely confronted with rare diseases than healthcare specialists from other disciplines, as those diseases frequently present symptoms early in life. Rare diseases are often referred to as “orphan diseases,” since in comparison to common non-communicable diseases, such as cardiovascular diseases and type 2 diabetes, there is less research and development of therapies for them. Fanconi anemia (FA) belongs to a small group of rare diseases that are investigated more intensively than most others (Velleuer and Carlberg, 2020). This is also the result of significant contributions from patient organizations in the United States, Germany and many other countries (Wu, 2013).

In general, evidence-based medicine aims to make optimal medical decisions by integrating the experience of a clinician with data from the individual patient and available scientific information on the respective disease (Akobeng, 2005). The latter information often derives from randomized clinical trials involving large numbers of cases and controls. Those trials are the source for the construction of statistical models, i.e., to quantify mathematical relationships between non-random variables measured from the study participants (Figure 1, left). For common diseases, there is no problem identifying a sufficiently large number of cases to achieve acceptable statistical power of the applied statistical model, e.g., reflected by the p-value. However, this approach cannot be used for rare diseases due to the small number of cases. An alternative approach is to study a few individuals in very high detail by collecting longitudinal samples for many biological parameters (Wheatley et al., 2021) (Figure 1, right). For example, multi-omic analyses provide many thousands of data points per individual, such as genome-wide DNA methylation, histone modifications and gene expression. These data, together with information on biochemical and regulatory pathways from public databases, such as KEGG (Kyoto Encyclopedia of Genes and Genomes) (Kanehisa et al., 2017), Wikipathways (Pico et al., 2008) and SPOKE (Scalable Precision Medicine Open Knowledge Engine) (Himmelstein and Baranzini, 2015), can then be used to construct multi-level, mechanistic dynamical computational models (Aldridge et al., 2006; Harris et al., 2019; Niarakis and Helikar, 2021). Importantly, mechanistic models differ from statistical models in that they encode existing biological knowledge, often obtained from decades of published experimental studies, into a formal, physics-based mathematical representation that mimics the real living system. Mechanistic models do not require experimental data for their construction, but they do require it to constrain the values of model parameters, which can be numerous. Indeed, the difficulty of estimating values of large numbers of adjustable parameters has been the primary barrier to the widespread utilization of mechanistic models in human health and disease applications. However, the situation is beginning to change. With recent advances in parameter estimation (Eydgahi et al., 2013; Shockley et al., 2018), model analysis and model selection methodologies (Beik et al., 2023), the construction of large-scale, executable models of whole cells, whole tissues and whole patients may now be within reach. Indeed, efforts to develop such detailed computational models, termed “medical digital twins” (Laubenbacher et al., 2021; Masison et al., 2021), are underway. These models can act as virtual platforms for identifying novel therapeutic targets and for designing treatment and preventative protocols to aid clinicians improving individual patient outcomes. In this Perspective article, we introduce the concept of mechanistic modelling as a clinical decision support tool in FA, using the example of the multi-step tumorigenesis of squamous cell carcinoma (SCC) in FA individuals.

FA is a rare disease (1 case in 300,000 persons) that was first described nearly 100 years ago by Guido Fanconi (Lobitz and Velleuer, 2006). The clinical characteristics of FA are: i) congenital malformations including absent radius, thumb hypoplasia, disturbed skin pigmentation, as well as inner organ abnormalities most frequently found in the renal and cardiac system (Fiesco-Roa et al., 2019), ii) progressive bone marrow failure already at childhood age (Alter, 2017; Dufour, 2017), and iii) dramatically increased risk of developing cancers, such as acute myeloid leukemia (Alter et al., 2018) and SCC, especially of the head and neck, in early adulthood (Kutler et al., 2003). FA individuals have defects in the molecular machinery of detection and repair of interstrand crosslinks (ICLs) and DNA double-strand breaks (DSBs), which are mostly due to the biallelic inheritance of recessive pathogenic variants in a subset of at least 20 FANC genes (FANCA, FANCC, FANCD1 (BRACA2), FANCD2, FANCE, FANCF, FANCG, FANCI, FANCJ (BRIP1), FANCL, FANCM, FANCN (PALB2), FANCO (RAD51C), FANCP (SLX4), FANC (ERCC4), FANCS (BRCA1), FANCT (UBE2T), FANCU (XRCC2), FANCV (MAD2L2), FANCW (RFWD3)) (Wang and Smogorzewska, 2015). Moreover, variants in the FANCB gene are inherited in an autosomal recessive X-linked manner, whereas the FANCR (RAD51) shows an autosomal dominant inheritance pattern and can also be spontaneously mutated (Meetei et al., 2004; Ameziane et al., 2015; Wang and Smogorzewska, 2015). FANC genes encode for proteins that maintain genomic integrity during DNA replication, i.e., their inactivation leads to accumulation of DSBs and genomic instability (Ceccaldi et al., 2016). However, patients with identical variants, such as siblings, often show significant differences in their clinical presentation, i.e., there are more factors than the mutated FANC genes contributing to the disease (Tischkowitz and Hodgson, 2003; Dufour, 2017).

To date, hematopoietic stem cell transplantation is the only curative treatment option for the hematological complications of FA (Gluckman, 2015) and the main reason for improved life expectancy of young FA individuals (Bonfim et al., 2016). In addition, treatment with supra-pharmacological doses of testosterone analogs, such as Oxymetholone, Danazol and others, can stabilize declining blood counts and even improve them (Scheckenbach et al., 2012; Rose et al., 2014; Paustian et al., 2016; Calado and Cle, 2017). Non-transplanted and, in particular, transplanted FA individuals have a several 100-fold increased risk for developing SCC, especially of their oral mucosa but also in their pharynx, larynx, esophagus, anus, and vulva (Alter et al., 2018), even without the main risk factors like alcohol and tobacco exposure. For adult FA individuals, developing SCC is the most life-threatening complication. Due to their dysregulated DNA repair machinery, FA patients cannot tolerate standard chemo-radiation therapies and treatment side effects are difficult to predict (Lin and Kutler, 2013; Kutler et al., 2016). This makes non-surgical systemic therapeutical options very limited. Therefore, detecting SCC at an early stage and eliminating it surgically is, at present, the best way to prolong the lives of FA individuals (Velleuer et al., 2020). Ultimately, prevention of disease progression should be the goal but is still far away from the clinical routine.

Based on its clinical and cellular phenotype, FA can also serve as a cellular model for the study of general molecular functions and physiological aspects, like aging, as well as other non-communicable diseases occurring in the general population. In that respect, the study of FA has had a major impact on the molecular understanding of breast/ovarian cancer (Howlett et al., 2002). Moreover, FANC genes are frequently mutated or dysregulated in sporadic cancers (Del Valle et al., 2020), as well as in childhood cancers (Pouliot et al., 2019). Nevertheless, the enormous and quite specific cancer risk of SCC for FA individuals is poorly understood from a mechanistic standpoint.

During tumorigenesis, the cells of all types of solid cancers arising in adults undergo a multi-step process from a healthy, non-transformed cell to low-grade and high-grade dysplasia, carcinoma in situ, and invasive cancer (Carlberg and Velleuer, 2021) (Figure 2A). The early stages of this process are reversible, but when a “point of no return” is reached, an aggressive carcinoma forms, which via the spreading of metastases will eventually lead to patient mortality. In cancer patients in the general population, this multi-step tumorigenesis process takes decades, but in FA individuals it can take only months to a few years.

During tumorigenesis, transformed cells acquire a conserved set of cancer hallmarks, including self-sufficiency in growth signals, insensitivity to antigrowth signals, and tissue invasion and metastasis (Hanahan and Weinberg, 2000). These are altered functional capabilities, the accumulation of which allows malignant cells to survive, proliferate, and disseminate. The hallmarks are outcomes of specific cell fate-controlling regulatory pathways, which in turn are affected by cancer driver gene mutations (Vogelstein et al., 2013). Environmental factors, like lifestyle decisions on diet, physical activity, and smoking, modulate these pathways. In addition, germline mutations may accelerate tumorigenesis like in the Li-Fraumeni syndrome (Carlberg and Velleuer, 2021). In contrast, 100% of all cancers arising in FA individuals (here termed “FA cancers”) are based on heredity of a defective FA/BRCA pathway of DNA repair. Therefore, genome instability and mutation prominently affects other physiological processes, which can result in the emergence of additional hallmarks, such as polymorphic microbiomes, tumor-promoting inflammation, deregulating cellular metabolism, activating invasion and metastasis, senescent cells, avoiding immune destruction, and non-mutational epigenetic reprogramming. These eight hallmarks of FA cancers (Figure 2B) summarize clinical, cellular and molecular observations in the field (Webster et al., 2022) (Box 1). They overlap largely with the latest version of 14 hallmarks of cancer (Hanahan, 2022). However, while in the general population the order in which the cancer hallmarks are established, as well as the underlying mechanisms, varies significantly, in FA cancers the hallmark “genome instability and mutation” always emerges first. For example, we illustrate five real cases of tumorigenesis in FA individuals (Figure 2C; Box 1). Note that these also demonstrate that in FA cancers there is variability in the order of appearance of subsequent hallmarks.

Cancer progression is a dynamical process ranging from early, mostly clinically asymptomatic, to late stages that are difficult to reverse. Each stage is characterized by a particular configuration (emergent behavior) of the tissue, such as infiltration of immune cells, the microbiome, cell cycle speed and the grade of cell dysplasia (Figure 3A). Disease onset and aggravation emerge from the dynamic interplay between hallmarks of SCC in FA (Figure 2B). The hallmarks are connected mechanistically by complex, multi-level regulatory networks that under homeostatic conditions maintain a healthy phenotype (Figure 3B). At the cellular level, the stratified epithelium underlying the epithelial barrier function interplays with the oral microbiome and the infiltration of immune cells, which together shape the micro-environment of tumor cells. At the sub-cellular level, genetic factors such as mutations in FANC genes, copy number variations and epigenetic reprogramming, together with micro-environmental perturbations, such as chronic inflammation and exposure to pathogens or a disturbed microbiome, can lead to altered cell fate decisions. These factors can be intrinsically disturbed, e.g., by aldehydes, but also influenced by lifestyle decisions, such as physical exercise, antibiotic treatment, immunosuppressants or chemotherapy. The complex interplay between these processes makes prevention but also optimal treatment protocols and individual patient assessments extremely challenging without a dynamical computational framework as an aid.

Based on the hallmarks of FA SCC and known underlying cellular and subcellular regulatory networks, we developed a blueprint for modelling SCC development in FA individuals (Figure 3C). Biological processes comprising the blueprint include: i) microbial interactions in the oral cavity (pink inset), ii) circulating immune cells and immune response (orange inset), iii) metabolites affected by food intake or endogenously generated within a cell (yellow inset), iv) DNA damage sensing and repair (red inset), v) epigenetic reprogramming (green inset), vi) stratified epithelial dynamics (brown inset), and vii) loss of epithelial function through EMT (epithelial-mesenchymal transition) (dark blue inset). Together, these tissue level processes are sensed and integrated by the cells and their inner regulatory networks (black inset), resulting in (viii) stress response and xi) cell cycle progression or senescence, x) leading finally to cell fate decisions, including survival, proliferation, cell death/apoptosis. The blueprint amounts to a preliminary wiring diagram connecting these processes and will act as a template for an executable computational model the dynamical multi-level dynamical network underlying the tumorigenesis in FA individuals.

The construction, calibration and validation of mechanistic mathematical models require a constant dialogue between the mathematical/computational implementation and analysis of the model, with the experimental and clinical data. As a first step, all available relevant empirical data describing the biological phenomenon are gathered from the literature together with in-house measurements from clinical, animal and in vitro studies. These data can be complemented using publicly available databases, such as String-DB (Szklarczyk et al., 2021), KEGG (Kanehisa et al., 2017) and the Human Cell Atlas (Regev et al., 2017). In principle, all types of data, from low-to high-throughput, single-timepoint to dynamic and mean-field to single-patient-single-cell resolution, can be used (Figure 4A).

After cleaning, filtering and pre-processing the data using standard statistical and bioinformatic methods (Sinha, 2014; Sinha, 2015), regulatory networks at the cellular- and tissue-level scales are assembled from the data and visually represented using the Systems Biology Markup Language (SBML) (Keating et al., 2020). The networks are translated into mechanistic mathematical models using a chosen modeling formalism that depends on the scale and resolution of the data. Multi-scale mathematical models can be modularly constructed and assembled using a variety of formalisms from dynamical systems (de Jong, 2002), including Boolean networks for genetic regulatory networks, systems of nonlinear ordinary differential equations (ODEs) for signaling networks and interactions between cell types in the tissue, compartmental differential equations or agent based models to explicitly model spatial coupling between different cell types, stochastic differential equations to simulate population-level distributions of cell markers and delay-differential equations for multi-level regulatory loops that control tissue homeostasis. In general, when constructing a model for a specific disease affecting a particular tissue (e.g., FA-related SCC in the oral mucosa), a good starting point is to identify mathematical models previously proposed and experimentally validated, which can then be extended and adapted to reproduce specific experimental and clinical observations. For example, mathematical models of loss of epithelial homeostasis, originally developed for atopic diseases (Domínguez-Hüttinger et al., 2017), could be extended to include SCC in FA-specific hallmarks. The quantitative models are then calibrated with a training set of experimental data, using global parametric optimization algorithms (Balsa-Canto et al., 2012; Shockley et al., 2018; Tsiantis et al., 2018). This allows one to adapt the mathematical models to specific experimental conditions, as well as to integrate scattered experimental data into a formal and coherent framework that articulates all the individual regulatory players that are typically described in isolation, to understand how they give rise to different clinical manifestations. Finally, model validation is achieved by ensuring that the model can reproduce an additional set of empirical data (the validation set) that was not used for the calibration (Figure 4B).

Next, the calibrated and validated models are analyzed extensively. For this, the nominal conditions, i.e., the calibrated model, are perturbed, e.g., by systematically altering the magnitude of the individual regulatory interactions (changing parameter values), or by structurally altering the different regulatory interactions (changing the equations). The output of the model, e.g., the “phenotype”, is then collected. Examples of such perturbations-to-response mappings are robustness analysis, sensitivity analysis and bifurcation analysis. A robustness analysis tells us which fraction of variations results in a given phenotype (Domínguez-Hüttinger et al., 2017). Sensitivity analysis weights each individual interaction (parameter) in terms of its contribution to the change in the model output (Zi, 2011). In bifurcation analysis, the model output is assessed as one (or more) parameters are gradually changed (Kuznetsov, 2004). Abrupt health-to-disease transitions mathematically correspond to qualitative changes in the model, and they occur typically at bifurcations. The value(s) of the parameter(s) at which such a bifurcation occurs, known as a bifurcation (set), is particularly interesting because it represents the critical threshold of a perturbation that can be tolerated by the regulatory structure. Furthermore, the appearance of these bifurcation sets can often be anticipated by early warning signals, such as an increased variance in recovery times of the system (Bargaje et al., 2017). Thus, early warning signals of bifurcating systems can be used to improve early detection strategies for abrupt disease transitions (Figure 4C).

Finally, once the mathematical model has been exhaustively calibrated, validated and analyzed, it can be used for designing and optimising personalised treatment strategies that consider the specific disease stage and patient characteristics (Fey et al., 2015). For this, one starts by identifying the potential targets of the intervention strategy, e.g., growth hormone receptor inhibitors that reduce excessive proliferation of malignant cells, and modifying the model accordingly. Next, analysis of the extended model, i.e., the system without treatments plus the dynamics of the treatments, can be performed (Figure 4C). For example, one can use bifurcation analysis to systematically explore how a given treatment affects the overall virtual population of patients (e.g., considering the natural variations that occur in a population due to polymorphisms) by looking at how the bifurcation sets shift under a specific treatment, such as cetuximab (an epidermal growth factor receptor inhibitor). With a similar line of reasoning, it is possible to use bifurcation analysis to explore how different treatment combinations affect the phenotype of a given patient. Here, again, it is particularly relevant to look for bifurcation sets that separate the qualitatively different clinical phenotypes because these curves correspond to all the minimal treatment combinations that can effectively trigger a disease-to-health transition (Tanaka et al., 2018). Besides bifurcation analysis, other techniques, such as model predictive control (Christodoulides et al., 2017), can also be applied to maximize treatment efficiency while minimizing duration, dosing, and negative side effects (Figure 4D). In all stages of this modelling pipeline, model predictions must be verified by comparing them to empirical data.

As a simple but illustrative example of the model construction process, we present a small biochemical model of the first few steps of ICL detection and repair by the FA/BRCA pathway (Niraj et al., 2019; Semlow and Walter, 2021). During DNA replication, the presence of an ICL causes the replication fork to stall. This stressed fork is detected by the protein FANCM, which binds to the branched DNA structure caused by replication fork arrest and recruits the FA “core complex” to the damage site (Figure 5A). The FA core complex is composed of three protein subcomplexes, each of which is composed of three proteins (Figure 5B): AG20 is a complex of the proteins FANCA, FANCG, and FAAP20 (FA core complex associated protein 20); BL100 is a complex of FANCB, FANCL, and FAAP100; and CEF is a complex of FANCC, FANCE, and FANCF. For simplicity, we have chosen to represent in the model each of these subcomplexes as distinct molecular species that reversibly bind to each other to form the FA core complex. The complete computational model includes these subcomplex binding reactions, the binding of FANCM to the ICL, and binding of the FA core complex to ICL-bound FANCM (Figure 5C). By defining parameter values for the rates of each of these individual reactions, the model becomes executable, i.e., the populations (or concentrations) of the constituent species can be simulated over time (Figure 5D). In this way, hypotheses regarding the effects of mutations (changing parameter values) and/or adding external perturbagens (e.g., drugs) can be explored in silico.

The tumorigenesis process (Figure 6A) results in a heterogenous composition of tumors, i.e., each tumor contains cells in various stages of the transformation process to aggressively metastasizing cells. Importantly, tumors are not only composed of malignant proliferating cells, but also by multiple cell types, thus making the tumor mass a complex ecosystem that includes immune cells of multiple types (B cells, T cells, macrophages, etc.), tumor-associated fibroblasts, endothelial cells (Anderson and Simon, 2020) and even microbes, including bacteria and fungi (Zong et al., 2023). At the same time, a tumor is not only composed of cells, but also of extracellular matrix and secreted factors that can signal messages among cells (Anderson and Simon, 2020). If malignant tumors from individuals with FA are to be characterized and this information used for accurate model building, all these factors must be accounted for. In this respect, high throughput multi-omics technologies can leverage the components of the tumor of interest, generating data in multiple modalities that need to be integrated and potentially exploited for discovering novel biomarkers and therapeutic targets for individuals with FA.

Of note, classical DNA sequencing, RNA sequencing (RNA-seq), and protein detection technologies are not able to deconvolute and deconstruct the above-mentioned complex composition of a given tumor, since they use the bulk content of the tumor or tissue and are, therefore, constrained to detect the mean expression of molecules, or the presence of a predominant DNA sequence, thus losing information of minor cell populations or incipient emergent cellular clones (Stark et al., 2019). However, we are witnessing the appearance, development, and refinement of multiple technologies with the capacity to resolve the cellular heterogeneity of tumors. Among these technologies, one of the most popular is single-cell RNA-seq (scRNAseq), which has given rise to a growing number of datasets from liquid and solid tumors (after tumor dissociation), as well as healthy tissue, leading to a compendium of single-cell-resolution gene expression atlases of multiple tissues and organs (Yuan et al., 2019). Although scRNAseq is a technology that has revolutionized the resolution at which we analyze cell populations and tissues, it still lacks a critical component, i.e., preservation of tissue architecture in its original context (Stark et al., 2019).

In the context of FA cancer, we are interested in the implementation of technologies that, in a multi-omics fashion, will generate single-cell resolution data but will prevent tissue disaggregation and, therefore, maintain tissue architecture. The latter implies the preservation of cellular neighborhoods and cell-cell interactions, which are lost when the tissue is disaggregated. These technologies are known as spatial omics and include spatial transcriptomics, spatial proteomics, and spatial genomics, which combine molecular characterization with spatial resolution (Akhoundova and Rubin, 2022). The aim of these spatial resolution technologies is to assign omics information to spatial locations in the tissues, reaching cellular and subcellular resolution. Spatial genomics assigns DNA sequencing information, including copy-number variants and somatic mutations; spatial transcriptomics provides information on the number of transcripts of a certain gene per region; and spatial proteomics provides relative amounts of protein concentrations (Akhoundova and Rubin, 2022). The data obtained by these multi-omics technologies are highly dimensional in nature and require potent computational tools for their analysis. Although intense research is underway for improving all spatial omics technologies, the most developed are spatial transcriptomics and spatial proteomics. These technologies will allow for the detection and quantification of cell populations of interest, the discovery of new cell populations, the comparison of the abundance of cell populations across the carcinogenic progression and the quantitative and qualitative description of infiltrating immune cells (Pelka et al., 2021; Karimi et al., 2023). These technologies have the capacity to compensate for the lack of resolution of bulk sequencing analyses, which has hampered the detection of premalignant clones at early stages in bone marrow failure in FA (Rodriguez et al., 2021).

Here, we use as an example a hypopharynx cancer from a 41-year-old woman with FA. The hematoxylin- and eosin-stained tumor sample shows multistage carcinogenesis, ranging from low-grade dysplasia (yellow) to high-grade dysplasia (orange) and invasive carcinoma (red) (Figure 6B). This type of formalin-fixed paraffin-embedded (FFPE) sample can be used for exploration and information retrieval using one or multiple of the multi-omics technologies discussed here (Box 2). If, e.g. tissue-cyclic immunofluorescence (tCycIF) is used, multiple sequential pictures of the tissue stained with fluorescent antibodies will be acquired and stitched. The composite image that is generated must first be segmented using artificial intelligence-based programs, such as ASHLAR (Pelka et al., 2021), which recognize every cell nucleus and apply single-cell-level segmentation of the tumor (Figure 6C, upper left panel). For every cell, we can feature-extract the expression of every marker of interest and proceed to non-supervised machine learning-based algorithms, such as uniform manifold approximation and projection (UMAP) (Figure 6C, upper right panel), which generate clusters of cells based on the similarity of their expressed markers. This allows the separate visualization of cell populations (Becht et al., 2018), such as cancer and immune tumor-infiltrating cells. After feature extraction, we can explore the expression of markers of interest in every tumor population or across the tumor progression, e.g. the proportion of proliferating cells (Figure 6C, lower left panel), the relative expression of p53 (Figure 6C, lower right panel) or other markers of interest.

Studies on SCC prevention in FA are limited by the small number of individuals with the condition, who are spread around the world. In addition to regular histopathological diagnosis of oral cancer development and cytology-based screening methods (Velleuer et al., 2020), reliable molecular markers are limited. Moreover, the scarcity of genotype-phenotype associations in FA makes it highly likely that each patient will respond in an individual way to drug treatments and/or lifestyle changes. Since robust predictive in vitro and in vivo FA models are lacking, drug screening and testing cannot be generalized for all FA individuals. For example, in vitro analysis of radiation sensitivity of fibroblasts from FA individuals does not correlate with the clinical response of the same patient to radiotherapy (Marcou et al., 2001; Alter, 2002) and the amount of chromosomal breaks found in lymphocyte cultures does not correlate with the severity of the disease, e.g., bone marrow function of the individuum. These issues motivate the effort to create multi-level, dynamical computational models of FA that can aid clinicians in tailoring therapies to each specific FA patient. Models of this type have been termed “medical digital twins” (Laubenbacher et al., 2021; Masison et al., 2021).

Although a consensus definition of a medical digital twin does not yet exist, the concept of a digital twin is common in engineering disciplines (Tao and Qi, 2019). Often referred to as “industrial digital twins,” these models are computational replicas of complex devices or processes, such as jet engines or wind turbines, that are used to diagnose technical problems and guide interventions. Industrial digital twins are typically composed of multiple, interconnected computational models of the constituent components of an engineered system. Critically, this integrated “template” model of the base processes of the engineered system in question is subsequently tuned, or “calibrated,” to a specific instance of that system, e.g., a particular jet engine, using performance data collected from sensors in real time. It is this “twinning” process, involving consistent feedback from real-time data streams, that differentiates a multi-level, computational model of a dynamical system from a true digital twin (An et al., 2022). Construction of digital twins for medical and clinical applications has been receiving increased interest lately (Laubenbacher et al., 2021; Masison et al., 2021). However, biological systems are far more complex than engineered systems, making their practical implementation much more challenging. Nevertheless, there have been successful applications of medical digital twins for the treatment of type 1 diabetes (Kovatchev, 2019) and pediatric cardiac patients (Shang et al., 2019). Furthermore, it is important to note that medical digital twins differ from alternative approaches gaining popularity in biomedical sciences, such as statistical and machine-learning models (Swanson et al., 2023), in that they are based on a mechanistic understanding of the underlying biological system. As such, they are not constrained by the confines of the experimental data on which they are constructed, which in FA is sparse.

The utility of a FA medical digital twin will be to aid clinicians in determining best courses of action for individual patients in both the prevention and treatment of malignant tumors. The template model for an FA medical digital twin will comprise the biological processes mentioned previously, including microbiome interactions, DNA damage sensing and repair, EMT, cell cycle progression, and cell death, among others (Figure 3). Calibrating the template model to individual FA patients will be challenging and require collecting spatially resolved, single-cell resolution multi-omics data, epigenome profiling, and metagenomics of the oral microbiome, from patients at regular intervals, e.g., every 3 months in accordance with clinical care guidelines for FA individuals with potentially premalignant lesions in the epithelial tissue. Additional patient data, such as blood draws, genetic screens, and oral swabs, together with standard data from electronic health records, can also be integrated into the calibration data stream. Once the model is personalized in this way, it will be possible to test, in silico, numerous preventive and/or therapeutic options before applying them clinically to FA patients. Furthermore, medical digital twins should be flexible and extensible, able to grow in precision and predictive power as new knowledge is accrued and experimental data sets are generated. In this way, the FA digital twin will develop together with the patient and their clinician, ultimately forecasting, with high accuracy and precision, responses to novel personalized interventions and therapies.

However, the here suggested use of mechanistic modelling has a few limitations. Often it is difficult to estimate the values of large numbers of adjustable parameters, which makes the use of mechanistic models in human health and disease a challenge. In addition, most physicians are not used to apply computationally generated recommendations in the general routine of patient diagnosis and care. Furthermore, potentially identifiable data of human subjects might make it difficult to exchange data across country borders.

For FA cancers, especially oral SCC, the use of multi-level dynamic mechanistic modelling provides a new perspective on early-stage diagnosis and decision support for the treatment of this rare disease. Such an approach is critical, since classical statistical models, using case studies and controls, cannot be applied due to the dearth of large patient groups. As such, we aim to build accurate computational models of tumorigenesis in a limited but representative number of FA patients. These mechanistic models will utilize pre-existing public knowledge on biochemical and regulatory pathways together with our knowledge of the life and disease course of more than 750 FA individuals, which will be essential for distinguishing the tumorigenesis process of FA cancer from that of regular cancer. In this way, our mechanistic models of FA cancer will take specific characteristics of this rare disease into account. Using longitudinal information about the lifestyles of FA individuals over years, together with multi-omics data at the genomic, transcriptomic, and proteomic levels, will lead to the construction of individual-specific models, or digital twins, that can be used to develop personalized treatment options. This approach has the potential to revolutionize the way FA individuals are treated clinically. Since multi-omic data is often used to build the digital twin, ethical approval needs to be obtained. Although recommendations of physicians in the context of disease prevention may not need ethical approval per se, but we strongly suggest institutional review board authorization, in particular when there is the aim to publish the findings.