Cafe-au-lait spots as a clinical sign of syndromes

Several studies describe the frequent association of cafe-au-lait spots with neurofibromatosis. However, many other genetic diseases might be associated with the presence of café-au-lait spots. Several genetic diseases are rare. In most cases, syndromes present themselves as a set of signs and symptoms that may present varied penetrance, therefore largely reducing the percentage of final diagnosis. Exploration of clinical symptomatology is essential for the understanding and diagnosis of syndromes. In this review, we conduct an extensive literature search looking for research that investigated diseases that may be present simultaneously with the cafe-au-lait spots. A total of 60 genetic diseases were found, all of them rare. These syndromes were evaluated based on their most relevant features and described in a summary of the typical, general, and head and neck findings. The available OMIM number, mode of inheritance, chromosome, mutated genes, and affected proteins were also listed. The considerable variety of diseases associated with the presence of cafe-au-lait spots and the fact that many of these conditions affect various organ systems with diverse phenotypic presentations is a diagnostic and therapeutic challenge. The objective of this study was to provide health professionals with an instrument containing a broad spectrum of genetic diseases coincident with the presence of cafe-au-lait spots in order to facilitate the differential and final diagnosis of these syndromes.


Introduction
Cafe-au-lait spots (CALS) or cafe-au-lait macules (CALM) are characteristically well-defined lesions, with a homogeneous light brown or medium to dark brown spots in dark-skinned people, that might be found all over the body except for the scalp, palms and soles (Hamm, Emmerich & Olk, 2019). Morphologically, they have been described as oval-shaped and with smooth edges or irregular contours, ranging in size from 0.2 cm to 30 cm in diameter, being smaller in young children since they increase proportionally to the body surface (Fistarol & Itin, 2010;Shah, 2010;Hamm et al., 2019). Histologically, they present increased melanin content in both melanocytes and basal keratinocytes, with giant melanosomes also being observed (Shah, 2010). They should be distinguished from lentigo (small pigmented spots with clearly defined edges, varying in size from 2 to 20 mm, usually smaller than 1 cm, that might occur anywhere on the skin) and nevus (a congenital or acquired usually highly pigmented area on the skin, flat or raised), clinical entities that alone will not be analyzed in this study (Fistarol & Itin, 2010). CALM may be present at birth or develop in the first years of life, which occurs in most cases (Hamm et al, 2019).
Isolated CALM is a common finding (10-36% of healthy people) with no clinical significance when dissociated from other findings (Rivers et al., 1985;Hamm et al, 2019). However, the presence of multiple CALMs, large segmental CALM, other cutaneous anomalies, associated facial dysmorphism or unusual findings on physical examination, may suggest the possibility of an associated genetic disease and should be promptly investigated (Shah, 2010).
Several steps are involved in determining skin color, such as lineage specification from embryonic neural crest cells (melanoblasts), melanoblast migration to skin of the embryo; proliferation and survival of the melanocytes in the basal layer of the epidermis; biogenesis of the melanosomes in the melanocytes; production of melanin granules in the melanosomes; translocation of melanosomes from the perinuclear region to the peripheral region of the melanocytes; transfer of the melanosomes from the melanocytes to the keratinocytes; and translocation of the transferred melanin granules from the peripheral region to the supranuclear region of the keratinocytes (Cichorek, Wachulska, Stasiewicz & Tymińska, 2013;Oiso, Fukai, Kawada & Suzuki, 2013). In parallel, a complex melanogenic paracrine network between the mesenchymal and epithelial cells regulates the processes involved in determining skin color after birth (Picardo & Cardinali, 2011;Oiso et al., 2013).
Multiple genes encode component proteins or signaling pathway regulators that control these paracrine network and the melanogenic growth factors which play a crucial role in the control of physiological and pathological skin pigmentation (Picardo Research, Society andDevelopment, v. 10, n. 9, e14310917607, 2021 (CC BY 4.0) | ISSN 2525-3409 | DOI: http://dx.doi.org/10.33448/rsd-v10i9.17607 3 & Cardinali, 2011). In this category are the KITLG gene, which encodes the Kit ligand (ligand for the receptor-type proteintyrosine kinase KIT) and the proto-oncogene c-KIT encoding the receptor tyrosine-protein kinase KIT, which activate the Ras/mitogen-activated protein kinase (RAS/MAPK) signaling pathway (Picardo & Cardinali, 2011;Oiso et al., 2013;Zhang, Li & Yao, 2016). Kit signaling plays an important role in a variety of physiological processes that occur in many cell types in the body, such as hematopoietic stem cells, mast cells, melanocytes, and germ cells (Ronnstrand, 2004). The RAS family comprises genes expressed in several types of normal cells: H-RAS, N-RAS, and K-RAS, which plays an important role in intracellular signaling pathways and in the regulation of functions such as cell cycle control, differentiation, growth, and cell senescence (Boquett & Ferreira, 2010). A large group of diseases associated with CALM result from germline mutations in these associated genes.
This article provides a critical review of the literature on genetic syndromes associated with CALM. In addition, the genotypic and phenotypic alterations of the identified diseases are described.

Methodology
We conducted a bibliographical research from September 2020 to January 2021, looking for studies that investigated diseases that can present CALM as a clinical manifestation. The compendium of human genes and genetic diseases OMIM (Online Mendelian Inheritance in Man) was used as the main source of data for the identification of associated syndromes.
Through a critical analysis of the literature, this narrative review was designed to provide a comprehensive understanding of the syndromes associated with CALM, in order to contribute to health professionals, facilitating the differential and final diagnosis of these syndromes. These objectives are in alignment with the reported by Oates & Harris (2015) on the importance of this scientific methodology, either to inform practice and to provide a comprehensive understanding of what is known about a topic.
The genotypic and phenotypic manifestations, as well as the dysmorphic changes in the head and neck of the identified syndromes, were obtained from the OMIM and Orphanet (Online database of rare diseases and orphan drugs) databases, and the book Gorlin's Syndromes of the Head and Neck (Hennekam, Allanson, Krantz, & Gorlin, 2010). Then, the search was expanded to other databases, such as PubMed (https://pubmed.ncbi.nlm.nih.gov/) and Virtual Health Library (VHL) (www.bvsalud.org). No limits were established regarding the date of the published works.

Results
Several genetic syndromes can be associated with CALM, all of them rare in occurrence. In the research conducted at OMIM, a total of 60 associated genetic diseases were identified. These syndromes were classified into subsections based on their most relevant features and described in a summary of typical, general, and head and neck findings (Table 1). Among the 60 diseases identified, 40 of them had frequent and significant changes in pigmentation or skin formation. Multiple CALMs are part of the clinical manifestation of at least 29 of these syndromes. In most other syndromes, the presence of CALM was simply an occasional finding, with a total number of affected patients too small to establish any overall rate of involvement, or its presence is not relevant compared to other more evident clinical characteristics.

OMIM (617506)
Noonan Syndrome-like disorder with or without juvenile myelomonocytic leukemia Typical: facial dysmorphism, wide spectrum of cardiac disease, reduced growth, variable cognitive deficits, ectodermal and musculoskeletal anomalies. General: CALM, lymphedema; thin skin; hypotonia; delayed psychomotor development (mild); language delay; increased susceptibility to juvenile myelomonocytic leukemia; joint laxity, cubitus valgus; cryptorchidism; pectus excavatum, widely spaced nipples; congenital heart defects, aortic stenosis, mitral insufficiency. Neurological disorders were observed in 32 types of diseases that present CALM, which may include a wide spectrum of changes, such as cognitive impairment, mental retardation, developmental delay, multiple aneurysms, cerebellar cortical degeneration, cerebellar ataxia, seizures, central nervous system malformations, brain tumors, hyperactivity, autistic spectrum disorders, peripheral nerve tumors, and other disturbances. Short stature was a characteristic observed in 28 of the identified genetic syndromes. Other relevant clinical findings observed in CALM-associated syndromes included predisposition to cancer (observed in 16 of 60 types of syndromes) and endocrine disturbances. Changes in the head and neck assessment were found in 52 syndromes linked to the presence of CALM.

OMIM (613563)
The OMIM number, mode of inheritance, chromosome, mutated genes, and affected proteins for the 60 identified diseases that may exhibit CALM in their clinical presentation are listed in Table 2 (grouped according to mutated gene classification). The most common mode of inheritance for syndromes with CALM is autosomal dominant, occurring in 68.3% (41 of 60) of these genetic disorders. An autosomal recessive mode of inheritance was detected in 21.6% (13 of 60) of the syndromes, with the remainder being X-linked cases, isolated cases, or unestablished patterns of inheritance.

Discussion
Genetic disorders account for approximately 80% of all rare diseases (Giugliani et al., 2019). All syndromes associated with the presence of CALM listed here are rare. There is no single definition for rare diseases. Within health systems, rare diseases have been defined based on the criteria of prevalence or number of affected individuals. According to the European Union, rare diseases are defined as those that affect less than 1 in 2,000 people (Giugliani et al., 2019). For the World Health Organization, a rare disease is one that affects up to 65 people in 100,000 individuals or 1.3 people in every 2,000 individuals.
In the United States, legislation defines rare diseases strictly according to prevalence, specifically as "any disease or condition that affects fewer than 200,000 people in the United States". Brazil follows the same definition of rare diseases adopted by the World Health Organization (Alawi, 2019;Martelli, 2019).
Most of these diseases that present multiple CALMs are part of the developmental diseases known as RASopathies.
These diseases, which are associated with germline genetic modifications, comprise a group of clinically and genetically related diseases that present mutations and deletions associated with protein-coding genes that lead to activation and/or dysregulation of the Ras/mitogen-activated protein kinase (RAS/MAPK) pathway (Tajan, Paccoud, Branks, Edouard, & Yart, 2018).
Defects at any stage of neural crest cell development, such as migration, proliferation, cell-to-cell interaction, differentiation or growth, are associated with the pathophysiology of neurocutaneous syndrome or phakomatoses (Sarnat & Flores-Sarnat, 2005;Gursoy & Erçal, 2018). This group includes pathologies with different genetic mechanisms (Sarnat & Flores-Sarnat, 2005;Klar, Cohen & Lin, 2016). So we have neurofibromatosis type I that exhibits mutations in the NF1 gene, which encodes neurofibrin, a negative regulator of RAS signaling that is also expressed in migrating neural crest cells during early fetal development; neurofibromatosis type II, where mutations occur in the NF2 gene that encodes a 595 amino acid protein, named Merlin, a negative Schwann cell regulator whose impairment allows Schwann cells to proliferate excessively; tuberous sclerosis complex, where mutations in the TSC1 and TSC2 genes are responsible for the pathogenesis of the disease, leading to overactivation of the mTOR pathway, which plays an essential role in normal cell growth, proliferation, and survival (Gursoy & Erçal, 2018). They usually involve inherited conditions, but spontaneous mutations can occur. As a common feature in the group, all diseases represent neurocristopathies and therefore share a common ectodermal embryologic origin, include abnormalities in the tissues of ectodermal origin, especially skin, eyes, and central nervous system (Reith, 2013). However, it is significant to emphasize that the neural crest is also important as an inducer of many tissues in craniofacial development and other mesodermal structures (Sarnat & Flores-Sarnat, 2005). Neurofibromatosis type I, neurofibromatosis type II, tuberous sclerosis, ataxiatelangiectasia, Peutz-Jeghers syndrome, McCune-Albright syndrome and Cowden syndrome 1 are diseases belonging to this group that have CALM as a phenotypic manifestation.
In addition to these classifications, localized or general melanotic hyperpigmentation may be part of the clinical presentation of many other congenital systemic disorders that result from ubiquitous protein defects and/or basal cell processes (Baxter & Pavan, 2013). This suggests that melanocytes are a cell type with high sensitivity to such disorders. A representative example is Fanconi anemia, a genetically heterogeneous disorder that affects DNA repair, characterized by different phenotypes that affect all organ systems (Baxter & Pavan, 2013).
Among all characteristics of these hereditary syndromes associated with CALM, predisposition to tumors is one of the most important, considering the high levels of cancers associated with these syndromes, with many of these cancers presenting in childhood (Walsh et al., 2017). The incidence of specific types of cancer in the carriers of the germline mutations is dramatically high compared to the general population, considering that the rate of a simple somatic allelic loss is exponentially greater than the independent mutation of two alleles within the same cell (Elissen, 2016). Malignant tumors are the most common cause of death in individuals with some of these familial tumor syndromes (Brems, Beert, Ravel & Legius, 2009).
The trend to develop tumors, which was observed in 16 of the 60 syndromes listed in this study, suggests a common underlying genetic basis. In this line, in neurofibromatosis type 1, the NF1 acts as a tumor suppressor gene (Origone et al., 2003). In this condition, a germline pathogenic variant and a somatic mutation lead to homozygous inactivation of the NF1 gene, resulting in a partial or total interruption of neurofibromin activity causing increased intracellular RAS signaling and abnormal cell proliferation (Origone et al., 2003). Different mechanisms are involved in the somatic inactivation observed in these hereditary syndromes, such as intragenic mutations (eg, nonsense, missense, frameshift, splice-site mutations, small insertions, and deletions), loss of heterozygosity, and hypermethylation of the promotor (Brems et al., 2009).
Another example is the genetically proven constitutional mismatch repair deficiency syndrome (CMMRD), a disease with multiple CALMs and other features of NF1 that are also part of the clinical findings. It is speculated that the remaining NF1 signals in patients with CMMRD result from post-zygotic mutations of the NF1 gene that may occur more frequently than normal in the population due to an accelerated rate of NF1 mutation in cells without a functional MMR system. However, it is also possible that CALM and other NF1 resources in these patients represent "isolated" skin manifestations (Maertens et al., 2007;Wimmer et al., 2017). Ataxia telangiectasia, Bloom syndrome, Nijmegen rupture syndrome and Fanconi anemia are among the most common DNA repair diseases and may present with CALM (Walsh et al., 2017). Pathogenic germline mutations in genes encoding proteins key in DNA repair and telomere biology result in the characteristic physical findings observed in patients with these hereditary disorders and in a high risk of cancer associated with these syndromes (Walsh et al., 2017).

Conclusion
The presence of CALM during a clinical evaluation of a patient should always be critically assessed by the healthcare professional, who must be aware of the possibility of an associated genetic syndrome. A detailed clinical evaluation should be performed by the physician to identify signs and symptoms that indicate the presence of any systemic disease.
If a genetic syndrome is suspected, given the genetic complexity and phenotypic heterogeneity of diseases that present CALM, the large number of overlapping features in similar diseases, and the incurable nature of these conditions, advanced testing is needed to distinguish between these syndromes, to provide genetic counseling to families, establish the prognosis and available therapeutic measures, monitor the potential risk to prevent complications.
It is important to note that, although the vast majority of genetic syndromes meet the criteria for a rare disease, it is estimated that there are between 6,000 and 8,000 different types of rare diseases described, with consequently millions of people affected by these diseases worldwide. The specific diagnosis of the genetic syndrome that affects a given patient has a great impact on their life, both in terms of clinical guidance and early treatment, and in the emotional sense, as it provides some comfort by obtaining an explanation for their symptoms, in addition to the possibility of obtaining support from groups of people affected by the same disease. Thus, research that systematizes knowledge on a given topic and organizes clinical signs and symptoms into a coherent system, as done in this study, can provide valuable tools in the process of building clinical reasoning and can add value to clinical practice, contributing to improved clinical outcomes.