Insights into the Molecular Mechanisms of Genetic Predisposition to Hematopoietic Malignancies: The Importance of Gene–Environment Interactions

The systematic use of comprehensive genomic approaches to study human disease over the past decade has led to the discovery of numerous germline and somatic (see Glossary at the end) DNA variants that elevate an individual's risk of developing cancer. Although historically germline predisposition has been well described for solid tumors, the recognition of its importance for hematopoietic malignancies, such as leukemia, lymphoma, or myeloproliferative neoplasms, has been increasing steadily. Furthermore, in contrast to the long-standing view that germline predisposition preferentially affects young people and causes solid tumors at ages below those seen in the general population, emerging data from the study of hematopoietic malignancies indicate that deleterious germline alleles can increase cancer risk throughout the entire age range of life. Indeed, the mutations driving hematopoietic malignancies in adults do not arise only as somatic events occurring later in life, but frequently, they may be inherited or occur at the very early stages of life as de novo variants that appear at the time of fertilization or occur postzygotically leading to somatic mosaicism.

The fact that predisposition might already exist at birth but not manifest as an hematopoietic malignancy until a later age prompts important questions about what mechanisms drive a cell harboring a predisposing deleterious variant to progress to a latent premalignant state and eventually to an overt malignant clone. As relates to childhood B-cell acute lymphoblastic leukemia (B-ALL), it has long been proposed that delayed exposure to infection during infancy or exposure to a specific pathogen might initiate an abnormal immune response that then triggers malignant transformation of susceptibly lymphoid precursors (1, 2). Although this has been challenging to prove conclusively in humans, preclinical mouse models have demonstrated that development of B-ALL requires an external postnatal stimulus, often an immune stress (see below) to spur the acquisition of additional somatic mutations leading to leukemia (3, 4). Similarly in adults, acquired somatic mutations accumulate during an individual's lifetime in all organs, but these are most easily accessed and measured in the peripheral blood, the only tissue where the stem cell progeny are pooled and mixed together. Thus, deep sequencing of an individual's peripheral blood gives a view of the acquired mutations that have occurred in the entire stem cell pool, something not feasible in any other tissue. This form of somatic mosaicism within the hematopoietic compartment is now referred to as clonal hematopoiesis (CH). Thus, some hematopoietic malignancies in adults arise as a consequence of germline predisposition, which may itself increase the risk for CH, whereas other hematopoietic malignancies arise purely as a consequence of somatic mutational events. Inflammatory and other environmental stressors likely promote the expansion of these clones and acquisition of additional somatic oncogenic mutations that ultimately give rise to hematopoietic malignancies.

The relative influence of genetic versus environmental factors may vary across different types of hematopoietic malignancies and demographic groups. A logical consequence of this perspective posits that the combined effects of host genetic factors, inflammatory bone marrow and systemic milieus, possible perturbations of host immune surveillance, and acquired genetic somatic mutations can drive the formation of hematopoietic malignancies and likely influence solid tumor risk as well. In this regard, a better understanding of the factors that promote acquisition of oncogenic second hits in premalignant clones as well as host immune surveillance mechanisms that curtail cancer progression is critical to design novel strategies to slow or even inhibit hematopoietic malignancies from developing. In this Perspective, we discuss the latest evidence for the roles of genetic predisposition and environmental factors in hematopoietic malignancy development and highlight directions for future research.

Germline predisposition to cancer has been recognized for decades, initially through the observation of rare familial clustering of cancer. Some of the earliest descriptions of familial leukemia appeared early in the 20th century, with reports of kindreds in which multiple close relatives were affected by hematopoietic malignancies (5–7). Despite this recognition, the first true leukemia predisposition gene was not identified until 1999, with the discovery of RUNX1 (also known as CBAF2; ref. 8) as the cause of a familial platelet disorder with predisposition to myeloid malignancy. Since this initial seminal discovery, many additional genes have been associated with increased susceptibility to hematopoietic malignancies with several recent reviews providing comprehensive details on these conditions (9–13). By and large, most of these conditions follow an autosomal-dominant pattern of inheritance in which passage of a single altered gene copy is sufficient to confer cancer risk. However, some autosomal-recessive conditions have also been described. Overall, the genes affected in these conditions regulate critical cellular processes, such as cell differentiation, DNA damage recognition and/or repair, apoptosis, ribosome function, and signal transduction. Furthermore, the oncologic phenotypes of hematopoietic malignancy–predisposing conditions appear to correlate with the expression pattern of the encoded proteins, with mutations in transcription factors expressed during hematopoietic development predisposing primarily to B-ALL (e.g., PAX5, ETV6, IKZF1) or MDS/AML (e.g., CEBPA, RUNX1), whereas germline alleles impacting genes that are expressed more broadly predispose to hematopoietic malignancies as well as solid cancers (e.g., TP53) and cause other non-oncologic manifestations (e.g., NF1, Fanconi anemia genes). Recently, data have also emerged linking certain germline predisposition variants to development of CH (e.g., TERT, RUNX1, GATA2, POT1), which can then progress to full-blown leukemia (9–18), and recent genome-wide association studies indicate that common genetic variants also play an important role in CH and myeloproliferative neoplasms (19, 20).

Despite the advances made, there are several aspects of hereditary hematopoietic malignancy predisposition that remain poorly understood. For example, for some genes (e.g., PAX5, SAMD9/SAMD9L, RAS family genes in Noonan syndrome), leukemia occurs earlier in life (21–23), whereas for others, leukemia occurs with a bimodal age distribution (e.g., ETV6), in which B-ALL occurs during childhood and AML in adulthood (24) or most often in adulthood (e.g., DDX41; ref. 25; Fig. 1). These findings suggest that there may exist distinct developmental stages during which hematopoietic stem/progenitors are most susceptible to cancer formation. Furthermore, the penetrance for hematopoietic malignancy (HM) development, when known, is highly variable between conditions with biallelic mutations in mismatch repair genes highly penetrant for lymphoid, but not myeloid, malignancies early in life (26), whereas deleterious germline variants in DDX41 and ANKRD26 lead primarily to myeloid malignancies, often later in adulthood (25, 27). Recently, a familial CH syndrome linked to deleterious POT1 variants has been reported to be associated with a range of benign and malignant solid neoplasms due to long telomere length, extended cellular longevity, and genomic instabliity (18). Finally, progression to leukemia often follows distinct patterns depending on the underlying germline predisposition. For most cases, the germline variant alone is insufficient to cause leukemogenesis. Thus, it is common to observe loss or mutation of the second gene copy within clonal cells. However, the patterns of acquired somatic mutations within leukemic cells are often distinct depending on the germline condition. For example, for germline SAMD9/SAMD9L, nonrandom monosomy 7 (leading to loss of deleterious germline allele) can cause leukemic progression, whereas UPD7q, or somatic SAMD9/SAMD9L, mutations are protective events associated with clinical remission (13, 22). In germline DDX41-associated hematopoietic malignancies (HMs), loss of chromosome 5q or acquired point mutation of DDX41, typically the R525H allele, is common along with additional leukemogenic somatic hits in ASXL1, CUX1, TP53, DNMT3A, and TET2 (14, 25). These latter observations suggest that disease progression is influenced not only by the gene affected in the germline but also by the timing/type of acquired somatic mutations and possibly the cell in which those changes occur (Fig. 2A). Altogether, these data beg the question as to what spurs or selects for the acquisition of second hits, and why do only some clones transform to a full-blown malignancy? This conundrum illustrates the need to understand better the interplay between host genetic factors and extrinsic exposures and/or environmental factors that cooperate to promote malignant transformation. In the next sections, we take advantage of recent advances in our understanding of hematopoietic malignancies to discuss the transition from premalignant to full-blown malignant state, highlighting specific opportunities and vulnerabilities that serve as starting points for development of novel strategies to prevent the formation of hematopoietic malignancies and possibly other cancers.

Despite genetic alterations that raise disease risk for B-ALL being present in >5% of healthy children, or in >10% of older people in the case of CH (4, 25), the majority of these predisposed individuals never develop the associated disease, thus suggesting that cellular context and/or environmental factors and “gene/environment” interactions play a crucial role in determining clonal expansion. In certain hematopoietic malignancies, the germline genetic risk allele may be the primary causal factor (e.g., deleterious germline 5′-end CEBPA variants, which have near 100% penetrance for developing a myeloid malignancy). In contrast, for other hematopoietic malignancies, environmental exposures (e.g., radiation exposure among survivors of the Chernobyl disaster, toxin exposure among survivors of the 9/11 World Trade Center attacks) are the major contributor to hematopoietic malignancy development (28, 29). As is the case with most cancers, the specific external factors that trigger the conversion from preleukemic to leukemic cells are not yet fully understood. A better understanding of these factors and their underlying mechanisms may provide insights into novel therapeutic and/or preventive strategies to modulate the risks associated with genetic susceptibility to hematopoietic malignancies, ultimately improving human health.

The contribution of environmental exposures to leukemogenesis has been studied most extensively in the context of childhood B-ALL. Because the age peak for childhood B-ALL occurs between 2 to 5 years, it was postulated that postnatal infection exposure causes immune stress and increases the risk of developing B-ALL in vulnerable children (4). The predisposing mutations associated with increased B-ALL risk often affect genes encoding transcription factors crucial for B-lymphocyte development (e.g., PAX5, ETV6, IKZF1), thus impacting cell differentiation and rendering preleukemic cells vulnerable to transformation upon exposure to immune stress (4). However, the mere presence of preleukemic cells does not lead to leukemia in the majority of the cases, either in mice or in humans, therefore implying that the existence of preleukemic cells alone is insufficient to cause leukemia and that an external trigger, such as an environmental factor or an additional susceptibility gene mutation, is necessary for tumorigenesis. Until recently, evidence was lacking to demonstrate conclusively that infections could trigger the progression of premalignant cells to B-ALL. However, studies using mouse models (e.g., Pax5^+/− germline heterozygous or ETV6-RUNX1-transgene expressing) provide evidence that B-ALL develops only after animals are exposed to common infections, with a penetrance that is remarkably similar to that observed in humans bearing similar mutations (30, 31). Moreover, the acquired somatic mutations associated with progression to B-ALL in these models match those observed in human leukemic blasts (Fig. 2B). Thus, these mouse models of B-ALL provide systems that faithfully mirror the human condition, enabling further exploration of the mechanisms promoting tumorigenesis. However, this is not universally true, particularly with some mouse models of CH and progression to myeloid malignancy (32).

As mentioned, there is a second level of action of this gene–infection interaction, in which leukemia-predisposing mutations can progress to B-ALL using distinct mechanisms, even when exposed to the same leukemia-promoting environment, such as infections (4). For example, when Pax5^+/− mice exposed to pathogens develop leukemia, the emerging B-ALL blasts commonly contain mutated Jak3, whereas mice expressing Etv6–Runx1 develop B-ALL blasts harboring alterations affecting genes of the lysine demethylase family (30, 31). This implies that distinct vulnerabilities to B-ALL resulting from respective initiating mutations may be gene- or mutation-specific despite undergoing similar progression stages and being triggered by comparable secondary exposures. (Fig, 2B). Altogether, whereas the preclinical results are promising, more research is needed to determine the specific mechanisms by which immune stressors promote leukemia development in humans who are at increased genetic risk for B-ALL.

As with childhood ALL, deleterious germline variants may also drive the development of hematopoietic malignancies in adults with the affected genes changing as does the age of cancer diagnosis (Fig. 2A). Similarly, evidence suggests that specific stressors drive the expansion of mutant clones in adults (32). For example, infection and inflammation trigger oncogenesis in individuals with CH with mutations in specific genes, such as TET2, DNMT3A, ASXL1, and JAK2, whereas the expansion of cells with mutations in TP53 or PPM1D is facilitated by genotoxic stressors such as chemotherapy. These results indicate that environmental stressors contribute to mutation-specific clonal expansion in adults. Furthermore, when exposed to environmental stressors, mutant clones like TET2 or PPM1D in CH generate higher levels of IL6, leading to inflammation, which could explain the link between CH and various inflammation-mediated diseases (33–37). Mice lacking Tet2 develop myeloid cell expansion that is completely dependent on an IL6-mediated systemic inflammatory milieu that is caused by translocation of gut microbes into the systemic circulation (36, 38). Similar results have been seen recently in nonhuman primates (39). Indeed, macrophages with TET2 mutations as well as individuals with TET2-mutant CH have increased IL6 levels (33, 40). Whether the gut microbiome plays a similar role in the expansion of CH clones in people remains to be seen.

As discussed above, similar environmental stressors can lead to clonal expansion in childhood B-ALL as well as adult HMs (Fig. 1), an observation that could provide valuable insights into the molecular origins of these tumors. However, one key question that needs to be addressed is: If the key step in the progression from preleukemia to a full-blown malignancy is the interaction between the preleukemic cell and an environmental stressor, why is B-ALL primarily a disease of children, whereas CH with progression to MDS/AMLS is a condition that predominantly affects adults (Fig. 1)?

There are at least two possible explanations for this difference. First, B-ALL occurs at an age coinciding with the peak period of B-cell maturation and rapid expansion (41). During this process, recombinase-activating gene (RAG) enzymes become activated and are responsible for many of the secondary genetic mutations that drive B-ALL (42). In contrast, aging hematopoietic stem cells often exhibit in homing to the bone marrow niche, impaired lymphocytic differentiation, and an increased propensity for myeloid lineage priming, which can contribute to the development of CH and myeloid malignancies (43, 44). Second, the age at which the hematopoietic malignancy manifests may serve as a proxy for the underlying biological pathway leading to disease (see Fig. 2A), or, in other words, different germline gene mutations manifest, causing hematopoietic malignancies at different ages. This hypothesis has been supported by recent studies of patients with MDS, which revealed the presence of germline- predisposing mutations in patients of all ages (45), suggesting that oncogenic changes affecting different developmental hematopoietic compartments might determine differential susceptibility to transformation explaining age-linked disease specificity and disease phenotype. In this context, the influence of the cell of origin where the oncogenic mutation first exerts its action is also paramount, because not all development stages are equally susceptible to the preleukemic potential of each given predisposing gene (46, 47). These aspects further suggest that the predisposing mutations themselves, and their cellular context, may be more influential than the timing of gene/environment interaction in promoting leukemia development. Additional research is needed to resolve why some predisposing genes are associated with lymphatic leukemias, whereas others predominantly cause myeloid cancers.

Several groups have sought to delineate the frequency with which deleterious germline variants cause bone marrow failure disorders, including MDS and aplastic anemia, with these disorders representing those prone to malignant transformation. In children diagnosed with primary MDS (after exclusion of classical bone marrow failure syndromes) approximately 8% have underlying SAMD9/SAMD9L syndrome and 7% have GATA2 deficiency, for a total of 15% with known common pediatric MDS predisposition syndromes (22). Additional germline mutations may occur in a small subset (∼5%) in genes like RUNX1 (48). Importantly, the recent discovery of SAMD9/SAMD9L mutations in pediatric MDS suggests that additional novel predisposition genes likely remain to be uncovered in this population. When diagnosed between the ages of 18 and 40 years, around 15% to 19% of individuals with MDS/acute myeloid leukemia (AML) arising from preceding MDS and aplastic anemia have candidate germline genetic risk variants, typically in genes controlling DNA repair and telomere biology (49). Further studies are needed to clarify the frequencies of definitive pathogenic germline mutations in adults, taking into account emerging knowledge on germline predisposition.

In a more recent study of paired peripheral blood samples from patients with MDS across the entire age range and their related allogeneic hematopoietic cell transplant (HCT) donors, the frequency of these alleles is at least 7% (45). Importantly, although the highest frequency of positive findings was in those diagnosed younger than 20 years old, deleterious germline alleles were seen above 5% in every age decile, with different genes identified across the age range (45). In contrast to long-held views, pathogenic/likely pathogenic (P/LP) variants in DDX41 cause hematopoietic malignancies almost exclusively in older adults, at the same age as the general population, challenging our theories about how germline alleles drive cancer (refs. 25, 50; Fig. 2A). Analyzing a similar cohort of 732 patients diagnosed with severe aplastic anemia who underwent HCT, 16.5% of individuals had causative LP/P germline variants, most commonly in TINF2, RUNX1, MPL, MECOM/EVI1, or ERCC6L2 (51). Thus, the age at which MDS/AA is diagnosed is linked to the underlying biological pathway(s) driving hematopoietic malignancies, with variants in genes encoding transcriptional regulators common in children, whereas DNA repair and telomere biology genes dominate adulthood diseases, and DDX41 is frequent in the elderly.

Because the incidence of CH rises with age, CH-associated clones play a larger role in development of HMs in adults than in children, and the inflammation associated with CH is thought to play a major role in HM pathogenesis, development of other malignancies, atherosclerotic cardiovascular diseases, and a variety inflammatory conditions (52–55). Recently, single-cell multi-omic studies demonstrated that chronic inflammation enhances the fitness of TP53-mutant clones promoting development of myeloproliferative neoplasms (56, 57). Similarly, inflammatory triggers have been associated with childhood leukemia pathogenesis (58–62). Together, these data reveal undercharacterized features and vulnerabilities of precancerous cells during early leukemogenesis, paving the way for precision cancer prevention. In people, TET2 and DNMT3A mutations are associated with high ARG1 expression, and increased arginase activity is detected in bone marrow mononuclear cells of low-grade MDS and chronic myelomonocytic leukemia (63). Data from the CANTOS trial suggest that patients with TET2-associated CH had fewer major adverse cardiovascular events while taking canakinumab, which neutralizes IL1β, compared with patients without CH (64), suggesting that targeting inflammation may reduce CH-associated disease. In fact, it has been recently shown that providing a less inflammatory context confers a lower clonal advantage to mutant clones and reduces the chances for preleukemic cells to progress and transform into frank leukemias (61, 65).

An area of active research currently is understanding the pathogenesis by which an individual progresses from having a deleterious germline variant to developing an overt hematopoietic malignancy. Many disorders appear to involve the development of CH (16, 66–72), but whether those clones are direct precursors to the hematopoietic malignancy or merely represent individuals at increased risk is not yet clear. At the extremes of the age spectrum, germline predisposition disorders demonstrate unique pathogenesis: SAMD9/SAMD9L in the young and DDX41 in the elderly (Fig. 2A). Each will be described below.

Sterile alpha motif domain-containing protein 9 (SAMD9) and its paralogue SAMD9L are located next to one another on chromosome 7q21 and demonstrate an unusually high propensity for self-correction by somatic reversion (13, 22, 73–75). Deleterious germline variants in these two genes are dominant, generally missense, and gain-of-function variants that drive refractory cytopenia of childhood, often manifesting as thrombocytopenia or pancytopenia, and MDS with monosomy 7/del(7q) preferentially in individuals within the pediatric age group (13). A unique aspect of myeloid malignancies that develop in these individuals is the nonrandom loss of the chromosome 7 containing the pathogenic SAMD9/SAMD9L variant (13). Patients can present with a variety of additional phenotypes, including those involving the immunologic (e.g., severe infections), endocrine (e.g., adrenal hypoplasia, primary adrenal insufficiency), intestinal (e.g., enteropathy, reflux, achalasia), genital (e.g., 46XY females, bifid shawl scrotum, testicular dysgenesis, intra-abdominal/inguinal testes, clitoromegaly), and neurologic (e.g., spinocerebellar ataxia, dysmetria, nystagmus, white matter abnormalities, and loss of Purkinje cells) systems (13, 22, 73–75).

P/LP germline DDX41 variants are the most common alleles that cause myeloid malignancies in adults, but they do so with a skewed age spectrum that challenges our long-held views about germline predisposition (25, 50). Several aspects of this disorder are quite unique: Specific variants are quite common in certain populations; male P/LP germline variant carriers progress to malignancy more common than females; and when progression occurs, acquired mutations in the normal copy of the gene occur frequently, particularly the R525H allele (25, 50). The skewed age distribution suggests that this disorder follows an unusual disease pathogenesis, which is currently unclear. Although DDX41 is a member of the DEAD-box RNA helicase family, the protein interacts with double-stranded DNA and RNA:DNA hybrids (76), where it acts as a regulator of splicing, rRNA and snoRNA processing, and the innate immune system (77–80). Interestingly, DDX41 germline mutations are present with a high prevalence (1 in 129) in the general population, with different variants associated with distinct ancestry groups and varying risks of developing MDS/AML (81, 82). Strong evidence that P/LP germline DDX41 variants induce an inflammatory milieu comes from the clinical observation that even when transplanted with wild-type donors, these individuals experience extreme graft-versus-host disease after allogeneic HCT unless posttransplant cyclophosphamide is used (83). Future scientific studies will likely reveal the unique underlying pathophysiology by which myeloid malignancies develop in these individuals.

Technical advances have enabled widespread germline evaluation, which is performed increasingly now for patients with hematopoietic malignancies without initial suspicion of germline predisposition because many of the predisposition genes are not associated with preexisting organ anomalies or specific phenotypes. Moreover, in many cases, family history of hematopoietic malignancies or cancer may be negative or unknown. Multigene panel testing using next-generation sequencing (NGS) is typically employed as a first-line test to identify deleterious germline variants, including single-nucleotide variants (SNV), small insertions, and deletions in genes implicated in HM predisposition. These panels are designed to capture gene-coding regions and splice sites, and for some genes like GATA2 and ANKRD26, also their regulatory regions and promoter, respectively. Accurate identification of an underlying germline predisposition allele requires testing DNA from nonhematopoietic cells to avoid confounding somatic variants. For healthy individuals, germline DNA is commonly obtained from venous blood, saliva, or buccal swabs. However, in people with HMs, such specimens often contain somatic mutations obscuring accurate germline assessment. Moreover, somatic reversion leading to disappearance of germline allele can occur in some syndromes (e.g., SAMD9/SAMD9L), further confounding analysis. Thus, best practices entail utilizing cultured skin fibroblasts or hair bulbs as the germline DNA source. Since culturing fibroblasts is associated with procedural barriers and delays, buccal swabs or peripheral blood may be used initially for faster turnaround, with confirmatory testing from cultured fibroblast DNA. Another common route leading to diagnosis of germline predisposition is somatic leukemia gene panels that incidentally identify variants in germline predisposition genes. These panels routinely include several predisposition genes such as GATA2, RUNX1, ETV6, CEBPA, and TP53. Positive somatic panel results are then followed by confirmatory germline testing using skin fibroblasts.

Importantly, NGS-based testing is typically not designed to detect copy-number variants (CNV). Therefore, comprehensive germline testing should incorporate methodologies capable of identifying both CNVs and SNVs. Approaches best suited for CNV detection include comparative genomic hybridization, single-nucleotide polymorphism arrays, and whole-genome sequencing (WGS). WGS is being adopted increasingly as a standard clinical genetic testing modality given its advantages: (i) its rapidly decreasing cost, which will soon reach parity with whole exome sequencing; (ii) its ability to detect all classes of genomic variants, including SNVs, CNVs, and structural genomic alterations; and (iii) the ability to reanalyze the data in the future for the identification of new germline predisposition genes. However, the breadth of WGS-derived data increases the proportion of uncertain findings that may prolong the turnaround time for obtaining results. Data interpretation must be performed by certified personnel following the ACMG/AMP guidelines (84), taking into consideration ClinGen variant curation rules, which are being established and optimized successively for classifying the pathogenicity of variants identified in hematopoietic malignancy predisposition genes (85).

Identification of individuals with germline pathogenic variants informs management across the lifespan. Carriers should undergo surveillance according to existing guidelines for associated non-oncologic manifestations and be referred to appropriate specialists. Comprehensive surveillance for hematopoietic malignancies (including repeat bone marrow testing, cytogenetics, and somatic mutation sequencing) can uncover syndrome-specific changes arising in the transformed cells of patients with germline predisposition. For example, asymptomatic carriers with germline ERCC6L2 mutations should undergo hematologic surveillance including detection of somatic TP53 mutations that can progress to AML (86). From a discovery perspective, such studies can shed light on mechanisms of malignant transformation and uncover novel strategies to prevent or treat hematopoietic malignancies.

For individuals already diagnosed with hematopoietic malignancies, identification of an underlying predisposition assists in determining prognosis, selection of therapy including tailored chemotherapy regimens, and planning HCT, as exemplified in the following scenarios: (i) patients with Fanconi anemia will require modified conditioning regimens for HCT and comprehensive post-HCT solid tumor surveillance to account for the underlying DNA repair defect (87); (ii) individuals with GATA2-related MDS need timely HCT as the only curative approach, whereas prolonged chemotherapy should be avoided to minimize risk of death from infections (88); (iii) patients with AML resulting from germline CEBPA mutations will likely respond favorably to chemotherapy but have a high risk of relapse or development of a new primary AML and thus require close monitoring (89); and (iv) patients with germline DDX41-associated AML experience more indolent disease courses and excellent outcomes following chemotherapy compared with AML without DDX41 mutations (90). These examples demonstrate how knowledge of the underlying germline syndrome informs prognosis, treatment decisions, and surveillance.

Research into germline susceptibility for hematopoietic malignancies has entered an era of rapid advances. Novel sequencing technologies and standardized variant interpretation approaches are unveiling genetic contributions across all ages, allowing rapid diagnosis and individualized management of patients and affected relatives. Disease modeling studies have provided fundamental insights into gene–environment interactions enabling progression to cancer. Precancerous cells are now well-established players in cancer development and hold promise for improved cancer diagnosis, disease monitoring, and prevention. The abundance of these mutant clones contrasts with the relatively low incidence of leukemias and illustrates the need to understand better the effect of these somatic mutations on hematopoietic cells and the role of environmental factors in cooperating to promote malignant transformation. Most research on environmental factors influencing CH has depended on mouse models kept in clean, or even sterile, conditions. This is a limitation in itself, because it does not replicate the natural environmental conditions associated with hematopoietic malignancy development in humans. There are many potential environmental perturbations whose effects in malignant progression still remain poorly understood. Nevertheless, the inflammatory milieu facilitated by autoimmunity, infections, and the microbiota and their roles in promoting the development of cancers should be prioritized since existing evidence implicates them in the tumorigenic process. Clinical and translational research studies will be vital to enhance our knowledge of these gene–environment interactions. In addition, we still need to explore how changes in the environment due to systemic therapies or lifestyle alterations might impact disease risk. Therefore, to understand fully how environmental factors might influence hematopoietic malignancy development, animal models with a specific genetic predisposition in which hematopoietic malignancies emerge without further genetic manipulation will likely be critical to reveal underlying mechanisms. Although mouse models are powerful tools to address these issues, there is also a clear need for human models, including use of primary cells, xenografts, and induced pluripotent stem cells, to capture key features not present in mice, allowing investigation of germline variants, particularly noncoding variants, that are unique to humans. Finally, the application of next-generation sequencing technologies like single-cell RNA sequencing, along with proteomics and advanced cytometry, such as multicolor quantitative confocal cytometry or mass cytometry, should enable us to identify the intricate molecular mechanisms (e.g., inflammatory biomarkers, immune stressors, and the microbiome) that control preleukemic cell behavior at the single-cell level. We anticipate that understanding the mechanisms that regulate the interactions between these preleukemic cells and immune/inflammatory stressors will equip us with novel strategies to prevent hematopoietic malignancies.

De Novo Mutation: a genetic mutation that arises spontaneously in an individual, generally at conception, rather than being inherited from their parents. They are not present in the parental germ cells, but are present in the individual's germ cells, and therefore can be inherited by that person's offspring.

Germ Cells: haploid cells that form the zygote: in females, eggs; in males, sperm.

Germline Predisposition: the presence of one or more altered gene copies (also called genetic variants, gene alterations, or mutations) that increase an individual's risk of developing a particular disease or condition. They are present in all the cells of the body and, therefore, are also found in the person's germ cells and can be passed down to the person's offspring; can either be inherited from the person's parents or de novo. See also Germline Variant.

Germline Variant: a genetic DNA variant (also called a gene alteration or mutation) that is present in all cells of an individual's body, including somatic and germ cells. They can be inherited from parents and can be passed on to offspring. See also Germline Predisposition.

Postzygotic Mutation: (see also Somatic Variant and De Novo Mutation) a genetic mutation that occurs after zygote formation during embryonic development in the cells of the developing fetus or later in life in the cells of an adult. Therefore, contrary to Germline Variants or Zygotic Mutations, they are not present in all the cells of the organism.

Somatic Cells: diploid cells that make up the organism; distinct from the haploid germ cells.

Somatic Variant: any DNA mutation or alteration that occurs in nonreproductive cells of the body, known as somatic cells. Therefore, they cannot be passed on to the offspring. See also Postzygotic Mutation and De Novo Mutation.

Somatic Mosaicism: also known as clonal mosaicism; occurs when the somatic cells of the body are of more than one genotype, due to the appearance of postzygotic mutations that result in various genetic lines (clones). Somatic mosaicism is not generally inheritable as it does not generally affect germ cells.

Zygotic Mutation: any DNA mutation that occurs in or is present at zygotic formation, in which the first cell is formed by fertilization of the egg by sperm, resulting in the formation of a new individual.

L.A. Godley reports other support from UptoDate, Inc outside the submitted work. M.W. Wlodarski reports other support from Partnership for Health Analytic Research, LLC outside the submitted work. No other disclosures were reported.

Research at C. Cobaleda's laboratory was partially supported by Ministerio de Ciencia e Innovación/AEI/FEDER (PID2021–122787OB-I00) and a Research Contract with the “Fundación Síndrome de Wolf-Hirschhorn o 4p-.” Institutional grants from the “Fundación Ramón Areces” and “Banco de Santander” to the CBMSO are also acknowledged. L.A. Godley receives funding from NIH/NCI U24CA258118, Edward P. Evans Foundation, NIH/HLHBI via Oregon Health & Science University R01HL155426, Leukemia & Lymphoma Society, NIH/NCI U01CA257666, and RUNX1 Research Program. K. Nichols receives funding from the American Lebanese Syrian Associated Charities (ALSAC), R01CA241452 from the NCI, R21AI113490 from the National Institute of Allergy and Infectious Diseases, Histiocytosis Association, and Cures within Reach. M.W. Wlodarski receives funding from ALSAC, Edward P. Evans Foundation, and The Vera and Joseph Dresner Foundation. Research in ISG group is partially supported by FEDER and by SAF2015–64420-R MINECO/FEDER, UE; by RTI2018–093314-B-I00 MCIU/AEI/FEDER, UE; by PID2021–122185OB-I00 MCIU/AEI/FEDER, UE; by Junta de Castilla y León (UIC-017, CSI001U16, CSI234P18, CSI144P20, and CSI016P23); and by the Fundacion Unoentrecienmil (CUNINA project). C. Cobaleda and I. Sánchez-García have been supported by the Fundación Científica de la Asociación Española contra el Cáncer (PRYCO211305SANC).