Langerhans cell histiocytosis (LCH) has historically evolved in its classification from a primary immune dysregulatory disorder to what current evidence supports as a dendritic cell neoplasm with an immune-inflammatory component. A key part of the classification of LCH as a neoplasm has been the identification of BRAF V600E mutations in 35% to 60% of cases. Tumor protein p53 (TP53) and RAS mutations have also been identified, albeit in less than 2% of reported cases. Of note, over 50% of patients with another dendritic cell disease, Erdheim-Chester Disease, have also been shown to have BRAF V600E mutations. Although the BRAF mutations have not been shown to be associated with extent of disease, they may still provide a target for a molecularly guided approach to therapy. In cases of LCH in which no BRAF mutations were identified, there was evidence for activation of the RAS-RAF-MEK-extracellular signal-regulated kinases (ERK) pathway, suggesting that similar to other tumors, this pathway may be therapeutically exploitable. Anecdotal responses have been reported in a few patients with LCH and Erdheim-Chester Disease to vemurafenib, a BRAF V600E inhibitor. Although these results pave the way for careful, prospective clinical testing, selection of the optimal groups in which to test such inhibitors, alone or in combination, will be critical based on the toxicity profile thus far observed in adults with melanoma and other BRAF mutated tumors.
Unlike brain tumors and melanoma, LCH and some of the related syndromes such as Erdheim-Chester Disease, has existed in a sort of Twilight Zone of ambiguity regarding its origin and whether it represents a neoplastic or immunoreactive disorder. The diverse clinical characteristics, including the sometimes spontaneous self-regression, along with the clearly defined inflammatory, granulomatous histology have led to the conclusion that LCH is a primarily immunologic dysregulatory disease. However, a series of cell and molecular investigations, culminating in the identification of distinct mutations of the BRAF gene and activation of the ERK pathway in lesional cells has led to the conclusion that LCH is a clonally derived neoplasm associated with intense expression of inflammatory cytokines and chemokines, similar in many ways to other neoplasms that are capable of eliciting inflammatory responses.1 This has resulted in LCH being termed an “inflammatory myeloid neoplasm.”2 This understanding has important therapeutic implications.
THE PATH TO PATHWAYS
Key to understanding the basis for a disease is to define its cell of origin and what drives that cell to become pathologic. Further, to understand the LCH cell of origin, one needs to place it in the context of normal cell lineage maturation.
Although the different clinical manifestations of LCH result in a nomenclature characteristic of the Tower of Babel,3 in 1941, Farber proposed that the different manifestations of LCH labeled as different disease shared similar radiographic bone findings and pathology.4 In 1953, Lichtenstein unified this concept by lumping the different disorders under the name Histiocytosis X.5 With emerging data demonstrating the presence of distinctive endocytic vesicles termed Birbeck granules as a characteristic of Langerhans cells, Nezelof et al proposed that Histiocytosis X was likely a type of tumoral transformation of the Langerhans cell.6 Subsequent studies determined that the LCH cell demonstrated maturational immaturity that could in part be overcome in vitro with an altered cytokine environment.7 In 2010, Allen et al reported an RNA expression pattern analysis that demonstrated overlap of expression of many transcripts in both Langerhans cells and LCH cells, but that the LCH cell transcript pattern most closely resembled a myeloid dendritic cell (DC).8 These results suggested that the LCH was derived from a precursor cell at one or more stages of bone marrow-derived myeloid maturation. However, another possible explanation is that the expression of transcripts characteristic of myeloid precursors by LCH cells is a result of chromatin reprogramming as a result of either inflammatory or aberrant molecular changes, similar to what takes place in epithelial-mesenchymal transition (EMT) of solid tumors.9 Thus, from this point of view, LCH may derive from resident LCs or precursors that then can proliferate and transition out of the epidermis to other anatomic sites. Consistent with this hypothesis is the loss of E-cadherin in LCH, similar to that observed in neoplastic EMT.10 Altered expressions of key chemokines and their receptors that regulate LC trafficking, especially chemokine receptors such as CCR6 and CCR7, as well as such key surface molecules, such as very late activation antigen 5 (VLA-5) and l-selectin, are consistent with such an EMT-like phenomenom.11-13 Of potential additional relevance is the demonstration that LCH cells express both the Notch ligand Jagged 2 and its receptor, suggesting an aberrant transcriptional program.14 Further work defining the LCH initiating cell as well as the nature of the pathologic cell should have important implications for therapy, similar to that of other neoplastic disorders.
Investigation into the cell of origin of LCH has paralleled work trying to understand the biologic underpinnings of the disease. Evidence for the clonal nature of LCH arose from exploitation of markers identifying maternal versus paternal X chromosome inactivation on flow cytometric sorted LCH cells and on unsorted lesions.15,16 On note the World Health Organization system of classifying hematopoietic disorders classified LCH as a clonal, neoplastic proliferation of pathologic Langerhans cells.17 The deregulated expression of nonmutant TP53, p21, and p16 as well as B-cell lymphoma 2 (BCL2), KI-67, RAS, c-MYC, and reduced caspase 3 suggest aberrant responses promoting cell cycle arrest while concomitantly supporting proliferation and survival, again similar to what can be observed in some cancers.18-20 Further, the expression of FAS and FAS ligand by LCH cells has been postulated to play a role in the spontaneous regression of lesions, suggesting an endogenous mechanism for disease regression rather than an immune-mediated rejection.21
The demonstration of a single case of an unbalanced t(7;12) translocation in LCH cells22 along with more subtle chromosomal gains and losses23 were intriguing, but subsequent studies were not able to reproduce similar changes.24 However, telomere length shortening, a finding characteristic of various cancers, was reported in LCH cells, and suggested that a survival mechanism to circumvent shortened telomeres could be at play in the pathologic LCH cell.25,26
Driver mutations are those that lead to the initiation and progression of cancers, whereas modifying mutations function more in altering the physiological responses of tumor cells.27 In 2010, Badalian-Very et al, using a 115 cancer gene subset of the OncoMap panel, reported the presence of oncogenic BRAF V600E mutations in 57% of cases of LCH.28 The mutations that were present were not associated with the extent of disease but did correlate inversely with the age of patients. A frequency of less than 2% of potentially transforming mutations was also noted in the genes encoding TP53, KRAS, and mesenchymal-epithelial transition factor (MET). The mutations involved single alleles, which is consistent with their dominant oncogenic nature. Of further importance was the observation that the RAS-RAF-MEK-ERK (MAPK) pathway was activated in essentially all of the cases using detection of phosphorylated forms of key signal transducers. Several subsequent reports have confirmed these findings.
A case of congenital LCH with a benign clinical course was reported.29 The BRAF V600E mutations have also now been reported in some cases of pulmonary LCH, historically considered to be a purely reactive disease linked to cigarette smoking in adults.30 Of special interest has been the observation of BRAF V600E mutations in the disease biopsy specimens from 54% of patients with Erdheim-Chester Disease, a rare form of histiocytosis, thus suggesting a possible common origin with LCH.31,32 Other histiocytic disorders, such as Rosai-Dorfman Disease, were not found to harbor BRAF mutations.28 Importantly, the detection of BRAF V600E mutations in LCH cells with different degrees of maturation, bringing the discussion back to whether LCH derives from a bone marrow precursor or an LC with altered transcription.33 Of course, both possibilities may be correct, depending on the heterogeneity of LCH in different patients.
PATHWAY DIRECTED THERAPY: DRUGS, DREAMS, AND DILEMMAS
The identification of drug-targetable V600E mutated BRAF in LCH raises the immediate question of whether patients will respond to such inhibitors (Fig. 1). The report of three patients with LCH and Erdheim-Chester Disease (two also had evidence of LCH) responding to the BRAF V600E inhibitor, vemurafenib, is potentially interesting, although these patients were not part of a clinical trial and follow-up was relatively short.34
A. Normal signaling of the RAS-RAF-MEK-ERK pathway generated by binding of ligand to a tyrosine kinase (TK) receptor resulting in phosphorylation (small green diamond) of the TK subsequently leading to activation of RAS and its downstream signaling targets including RAF and PI-3K followed by activation of ERK and then various key cell survival regulators. B. In LCH or other BRAF V600E-mutated neoplasms, the activation of BRAF (monomer) leads to constitutive activation of ERK without the need for dimerization or RAS upstream signaling. The linkage to the Notch pathway is unclear but appears to be active in at least some tumor types. Dashed lines have less certainty as does the presence of a question mark. Rx means possible treatment opportunities.
Several critical factors should be considered in the development of clinical trials with BRAF inhibitors for patients, particularly children, with histiocytic disorders. As is noted in multiple trials with BRAF inhibitors, not all neoplasms with BRAF V600E mutations respond similarly.35 This suggests the key need to define the cellular and molecular context in which the BRAF mutation resides. The prerequisite determination of the type of the RAF mutation is also critical, as not all mutations respond to inhibition, and some may be stimulated by inhibitors directed to the BRAF V600E mutation, providing the possibility of stimulating tumor progression.36 In addition, multiple feedback loops and resistance mechanisms rapidly develop to overcome initial sensitivity to BRAF inhibition.37,38 To this end, combination drug regimens, such as the use of BRAF and MEK inhibitors to positive clinical end in patients with metastatic melanoma, are likely to be more effective.39,40 Further, the incidence of dermatologic toxicity, particularly the development of squamous cell cancers and new onset melanoma/dysplastic nevi, in patients treated with BRAF inhibitors, raises the need to careful selection of patients.41,42 For instance, initial trials may need to focus on adults and/or children with more refractory and progressive disease.
The activation of the RAS-RAF-MEK-ERK pathway nevertheless provides new opportunities for understanding LCH and related disorders, as well as treating afflicted patients. For instance, it has been known for many years that anti-inflammatory agents, such as indomethacin, steroids, and tumor necrosis factor (TNF)-alpha inhibitors can result in positive clinical responses in patients with LCH.43,44 It is possible that modulating the ERK pathway may have advantageous effects on microenvironment interactions in such inflammatory neoplasms.45,46 Furthermore, there is evidence in other tumor systems of a linkage of ERK pathway activation and stabilization of c-MYC and Notch pathway-mediated inflammatory reactions.14,47-50
The presence of BRAF V600E mutations may also provide an important marker to follow in terms of assessing response to treatments as well as minimal residual disease for both pathway directed clinical trials and in patients treated with conventional chemotherapy. The evidence that a large percentage of patients with LCH do not harbor BRAF V600E mutations strongly suggests the presence of additional mutations possibly part of, or linked to, the ERK pathway, but are yet to be identified. Some of these mutations may turn out to be cooperating mutations to overcome the often senescence phenotype associated with BRAF activation, as is observed in melanocytes.51,52 Whether such mechanisms are functional in LCs or their precursors remain to be discovered and possibly therapeutically exploited.52-54
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