Current insight into the molecular mechanisms of lymphangioleiomyomatosis

Abstract

Lymphangioleiomyomatosis (LAM) is a rare, progressive lung disease that predominantly affects women, particularly during their reproductive years. It is characterized by the proliferation of abnormal smooth muscle-like cells that infiltrate the lungs, leading to cystic destruction of lung tissue and a decline in respiratory function. The pathogenic defect in LAM occurs due to mutations in the tuberous sclerosis complex genes, resulting in failure to suppress the mTOR pathway activation, which drives abnormal cell proliferation and lymphangiogenesis. Although mTOR inhibitors, such as sirolimus, have improved clinical management by slowing disease progression, they are not curative and do not reverse existing lung damage. Recent research has expanded our understanding of LAM pathogenesis by revealing substantial genetic and cellular heterogeneity within LAM lesions. Beyond mTOR dysregulation, pathways involving estrogen signaling, metabolic adaptation, and immune evasion contribute to disease development. These insights open new avenues for treatment. A better understanding of these pathways will pave the way for more durable and individualized treatments for LAM.

Graphical abstract

Graphical Abstract.

Clinical Relevance.

Lymphangioleiomyomatosis (LAM) remains a progressive disease despite mTOR inhibition, underscoring the need for novel therapeutic strategies. Growing recognition of the complex immunological landscape within LAM lesions, including mechanisms of immune evasion and tumor-immune cell interactions, highlights the potential for immunomodulatory approaches as adjunctive or alternative treatments. Targeting immune pathways alongside mTOR inhibition may provide more durable and individualized clinical benefit.

Introduction

Lymphangioleiomyomatosis (LAM) is a rare neoplastic lung disease that affects women in their reproductive years. Lymphangioleiomyomatosis involves the accumulation of abnormal smooth muscle-like cells in lungs, destroying pulmonary architecture and forming cystic lesions.¹ The disease presents as dyspnea, pneumothorax, chylothorax, and respiratory failure in advanced stages. Recent research suggests LAM is more prevalent than previously thought, with studies showing approximately 23.5 cases per million adult women.² Diagnosis delays occur due to nonspecific features.³

Lymphangioleiomyomatosis exhibits neoplastic characteristics, including proliferation, apoptosis resistance, metastasis-associated gene expression, immune evasion, and stem cell properties, due to increased mTOR activity.⁴ Single-cell transcriptomics has revealed heterogeneity in LAM cells and identified molecular subtypes affecting disease progression and treatment response, highlighting the need for personalized therapy.⁵

Lymphangioleiomyomatosis pathogenesis arises predominantly from loss-of-function mutations in TSC2, which encodes tuberin, a negative regulator of mTOR complex 1 (mTORC1) signaling. While TSC1 and TSC2 function together as part of the tuberous sclerosis complex, pathogenic TSC1 mutations are rare in sporadic LAM. Loss of TSC2 results in failure to suppress mTORC1 activity, leading to uncontrolled cell growth and disease pathology.^6,7 Mechanistic target of rapamycin complex 1 dysregulation enhances cell migration and angiogenesis, worsening disease progression.¹ This has led to targeted therapies like sirolimus, which reduces lung function decline and tumor burden.⁸

Recent advancements have identified new biomarkers improving LAM diagnosis and monitoring. Vascular endothelial growth factor D (VEGF-D), a lymphangiogenic factor, indicates disease severity and treatment response.⁹ These biomarkers enhance diagnostic accuracy and provide insights into disease mechanisms. Discovery of novel biomarkers remains critical for diagnostic precision and revealing therapeutic targets for this complex condition.

Despite these advancements, LAM biology remains poorly understood, particularly its cellular origins, progression mechanisms, and immune-hormonal interactions. The rarity and heterogeneity of the disease challenge treatment development. However, genome-wide association studies (GWAS) and single-cell sequencing studies have revealed genetic contributors and subtypes, advancing personalized therapeutic approaches.

This review explores molecular and genetic mechanisms of LAM pathogenesis, including TSC1/TSC2 mutations, mTOR pathway dysregulation, hormonal influences, and genetic factors.

Genetic drivers of LAM

Lymphangioleiomyomatosis is genetically determined, and mutations of the TSC1 and TSC2 genes are the main pathogenic causes. Recent studies have also identified additional genetic players, including NR2F2 and described the heterogeneity of LAM cells. Here, we discuss these genetic drivers and their relevance to disease progression and treatment.

TSC2 as the primary genetic driver of LAM

Loss-of-function mutations in TSC2 are the predominant genetic abnormality in LAM and lead to dysregulated signaling through the mTORC1, a central regulator of cellular growth and metabolism. While hamartin and tuberin normally act to restrain mTORC1 activity, pathogenic TSC1 mutations are rare in sporadic LAM. Loss of TSC2 results in failure to suppress mTORC1 signaling.⁶ Mechanistic target of rapamycin complex 1 activation causes cell proliferation in tumors and affects protein synthesis, autophagy, and metabolism, leading to LAM’s hallmark manifestations of cyst formation and lung destruction.^10,11

Inoki et al.¹² show the TSC1-TSC2 complex integrates Wnt and AMPK pathway inputs. Targeting this mechanism in TSC mutations demonstrates a link between cellular energy metabolism and LAM pathogenesis.¹² This dysregulation stimulates LAM cell proliferation and impairs adjacent tissues, contributing to disease manifestations. The study of 261 TSC patients by Kozlowski et al.¹³ found that large rearrangements like deletions and duplications of the TSC2 gene are frequent alongside point mutations. These findings align with Murphy et al.,¹⁴ who identified similar TSC2 gene alterations in sporadic LAM. These studies reveal common genetic pathways in TSC and LAM, highlighting the need for comprehensive genetic testing. Although mTORC1 inhibitors like sirolimus stabilize lung function, they cannot reverse tissue damage, necessitating new therapeutic approaches.

Recent genomic and single-cell sequencing studies reporting lower TSC1/TSC2 mutation detection rates likely reflect tumor heterogeneity, mosaicism (where only a subset of cells harbor pathogenic mutations, as documented in LAM cases with low-level TSC2 mosaicism^15,16), low neoplastic cell content, and technical limitations of contemporary sequencing approaches, rather than absence of TSC pathway involvement.

Emerging genetic contributors

Recent GWAS have implicated additional genetic factors in LAM pathogenesis beyond TSC1 and TSC2 mutations. Kim et al.¹⁷ reported associations between NR2F2/RTF1 variants and LAM-related phenotypes. NR2F2 encodes a nuclear receptor involved in vascular development and smooth muscle differentiation, suggesting its dysregulation could affect LAM cellular behavior. This discovery suggests alternative molecular targets for therapeutic intervention beyond the mTOR pathway. Huang et al.¹⁸ examined mutations in sporadic LAM cases through genomic profiling of 22 patients, identifying 67 variants across 43 genes, with 68.2% TSC2 mutation rate. A novel VEZF1 mutation (c.A659G) was identified in 4 participants and was associated with a milder phenotype, although this observation is based on a small cohort and requires further validation. These findings indicate sporadic LAM may arise from mutations in non-TSC2 genes, with genetic profiles potentially influencing clinical outcomes. Cho et al.¹⁹ studied TP53 codon 72 polymorphism in angiomyolipomas, LAM, and TSC patients, examining its role in tumorigenesis through Notch-Nodal signaling. This polymorphism affects cellular stress responses and apoptotic stimuli, potentially enhancing TSC-associated neoplasm proliferation and promoting tumorigenesis. Further research is needed to understand how TP53 variants influence LAM progression and treatment outcomes.

Heterogeneity of LAM cells

Lymphangioleiomyomatosis is characterized by a lack of ­uniformity, and its genomic landscape reflects this complexity. Guo et al.⁵ identified a unique LAMcore cell population with a specific genetic signature using single-cell transcriptomics. Lymphangioleio­myomatosis core cells drive disease pathology through their characteristics and cross talk with other cells in the microenvironment. This cellular heterogeneity highlights the need for personalized therapies, as different subpopulations may respond differently to mTOR inhibitors. Espín et al.²⁰ explored LAM cells’ cancer-like features, emphasizing their genetic and phenotypic heterogeneity. Their study identified subpopulations with differential expression patterns in hormone signaling, metabolism, proliferation, and stemness, including markers like lumican and YB1.

Olatoke et al.²¹ reported a HOX-PBX gene network critical for LAM cell survival using single-cell multiomic analysis. Their results suggest disrupting HOX-PBX interactions could delay disease progression. Epigenetic modifications also regulate LAM progression. Research showed tuberin and hamartin’s abnormal expression in breast cancer correlates with poor outcomes.²² TSC1 gene silencing in LAM cells is linked to promoter methylation. Beyond DNA methylation, studies indicate HDAC inhibition can suppress proliferation in TSC2-deficient cells,²³ suggesting chromatin remodeling as a therapeutic target. The genetic factors in LAM, particularly TSC1/2 mutations, highlight the disease’s complexity and need for customized treatments. Future studies should validate these genetic insights and assess their therapeutic implications (Figure 1).

Figure 1.

Cellular and molecular heterogeneity in lymphangioleiomyomatosis (LAM). Lymphangioleiomyomatosis is driven by a complex interplay of genetic, hormonal, and microenvironmental factors, contributing to the heterogeneity of the disease. Genetic mutations drive the abnormal proliferation of LAM cells. These cells are further shaped by hormonal signaling and immune interactions, leading to extracellular matrix (ECM) remodeling and disease progression.

Recent single-cell and spatial transcriptomic studies also provide insight into the broader cellular context of LAM lesions. In addition to LAM cells, these analyses highlight the presence of fibroblasts and other stromal cell populations within affected tissue, suggesting a role for the local microenvironment in disease progression. While the precise cellular origin of LAM cells remains an area of ongoing investigation, interactions between LAM cells and surrounding stromal cells are increasingly recognized as contributing to extracellular matrix remodeling and paracrine signaling within LAM lesions.^24,25

Dysregulation of molecular pathways

Dysregulation of molecular pathways is key to LAM pathogenesis. Central to this loss of regulation is the hyperactivation of the mTOR pathway induced by mutations in TSC1 and TSC2. Hormonal signaling and other molecular cascades, including the MAPK and histamine pathways, also play a role in disease progression. Here, we discuss the involvement of these pathways in LAM and their relevance to targeted therapies.

The mTOR pathway

The mTOR pathway is a key player in the pathogenesis of LAM, as it controls cell growth, proliferation, and metabolism. TSC1-TSC2 normally inhibits mTORC1 activity in healthy cells to prevent inappropriate cellular behaviors. In LAM, however, mutations in TSC1 or TSC2 lead to failure to suppress mTORC1 signaling and promote the aberrant proliferation of smooth muscle-like cells.Mechanistic target of rapamycin activation also appears to control angiogenesis and lymphangiogenesis—2 processes involved in the systemic manifestations of LAM. Vascular endothelial growth factor D, a lymphangiogenic factor, has been widely studied as a biomarker of severity and treatment response in LAM.⁹ Elevated levels of VEGF-D in the blood correlate with disease severity and can be slightly modulated through mTORC1-dependent signaling, reinforcing the pivotal role of mTOR dysregulation in maintaining and advancing the disease.

McCormack et al.⁷ demonstrated the usefulness of the mTOR inhibitor sirolimus in stabilizing lung function and reducing angiomyolipoma size in patients with LAM. These findings led to the adoption of mTORC1 inhibitors as first line therapy for LAM. Yao et al.⁸ showed that health benefits persisted on prolonged follow-up for patients treated with sirolimus, showing reductions in cystic lesions and improvements in pulmonary function. Beyond its therapeutic effects, mTOR dysregulation has systemic downstream effects on various pathways. Although mTOR inhibitors improve clinical outcomes in LAM, their effects are largely cytostatic, and disease progression typically resumes following treatment discontinuation. At a molecular level, this limited efficacy is supported by evidence that several downstream processes in TSC2-deficient cells are rapamycin-insensitive, including regulation of extracellular matrix remodeling, DNA damage responses, and metabolic pathways. These findings indicate that incomplete suppression of mTOR-dependent outputs and persistence of mTOR-independent signaling programs may contribute to the limited durability of mTOR inhibitor responses.^26,27

Tuberin (gene symbol TSC2) inhibits the activation of 2 important downstream effectors of mTORC1, p70 S6 kinase, and ribosomal protein S6. These proteins are hyperphosphorylated due to loss-of-function mutations in TSC2, enhancing DNA synthesis and proliferation of LAM cells.⁶ These findings elucidate the molecular changes that facilitate the pathogenic growth and survival of LAM cells and establish a mechanistic link to the significant role of mTOR signaling in disease development.

Another downstream effector of mTOR, previously studied in relation to cardiomyopathy, has also been recognized for its potential involvement in LAM pathogenesis. Wang et al.²⁸ identified αB-crystallin (HSPB2) as a molecular chaperone induced by hyperactive mTOR signaling in cardiomyocytes in a model of mTOR-driven cardiomyopathy. Although this work focused on cardiac tissue, it suggests that HSPB2 may contribute to cellular stress adaptation in contexts of sustained mTOR dysregulation. These findings highlight an interaction between molecular chaperones and mTOR signaling in the cellular stress response; however, the relevance of HSPB2 in LAM remains indirect and has not been functionally validated in LAM models.

Alesi et al.²⁹ highlighted transcription factor EB (TFEB) as a transcription factor that governs lysosomal biogenesis. Under normal conditions, mTORC1 phosphorylates TFEB at the lysosomal surface, a process that depends on its recruitment by Rag GTPases, particularly the active RAGC/D heterodimer. This phosphorylation retains TFEB in the cytoplasm. Loss of TSC2 can lead to dysregulated mTORC1 signaling and altered Rag GTPase activity, particularly involving RAGC, which may impair proper TFEB phosphorylation and localization. Consequently, TFEB translocates to the nucleus and activates a transcriptional program that promotes lysosomal biogenesis and metabolic reprogramming. This contributes to aberrant cell growth and hyperproliferation. While Rheb has long been recognized as the primary driver of mTORC1 hyperactivation following TSC2 loss, failure to restrain Rheb activity in the absence of TSC2 further amplifies mTORC1 signaling, promoting uncontrolled cell proliferation. However, a more recent work from the same research group³⁰ indicated that TFEB also contributes to sustained mTORC1 activity due to increased lysosomal biogenesis. This creates a positive feedback loop, whereby TFEB activation enhances the lysosomal pathway, which in turn further activates mTORC1 signaling. These findings suggest that TSC2-deficient cells rely on activated mTORC1 driven by Rheb as well as on TFEB-mediated lysosomal adaptations for survival, indicating that lysosome biogenesis may represent a potential therapeutic target in LAM.

A related, yet equally significant, regulator of mTORC1, calcium and integrin-binding protein 2 (CIB2), was studied by Sethna et al.³¹ They demonstrated that CIB2 binds Rheb in a manner that maintains it in an inactive state, resulting in decreased mTORC1 activity. This regulatory pathway governs growth, metabolism, and autophagy. Nevertheless, an imbalance in CIB2 function can lead to Rheb hyperactivation and mTORC1 overactivation, inhibiting autophagy. These alterations further enhance cellular proliferation and may contribute to LAM pathogenesis. Restoring CIB2 function or employing adenoviral vectors to mimic CIB2-dependent Rheb inhibition may serve as a targeted therapeutic approach to counteract severe, uncontrolled mTORC1 activation in LAM.

Nonetheless, challenges remain in the use of mTOR-targeted therapies. Liu et al.³² found that rapamycin pretreatment upregulates miR-29b in mTORC1-deprived TSC2-deficient cells. This microRNA, which directly targets and downregulates the tumor suppressor retinoic acid receptor β (RARβ), promotes cell growth. RARβ, in turn, interacts with the tumor suppressor ING4. Attenuated activity of ING4 upon loss-of-function of RARβ promotes LAM cell proliferation and tumorigenic potential. Consequently, the miR-29b/RARβ/ING4 axis builds a negative feedback loop that antagonizes the tumor-suppressive effects of mTOR inhibition, which may reduce the therapeutic effect of rapamycin. These findings highlight the intricate pathogenesis of LAM (Figure 2).

Figure 2.

Mechanistic target of rapamycin complex 1 (mTORC1) signaling dysregulation in TSC-deficient LAM cells. Loss of the TSC complex leads to failure to restrain Rheb activity, resulting in mTORC1 hyperactivation. This drives LAM cell growth and alters multiple signaling pathways. CIB2 normally restrains Rheb activity; however, its dysfunction contributes to mTORC1 overactivation. Inhibition of mTORC1 by rapamycin induces miR-29, which suppresses RARβ expression, thereby reducing the activity of the tumor suppressor ING4. This negative feedback loop may diminish the therapeutic effects of rapamycin. In parallel, mTORC1 regulates lysosomal biogenesis via TFEB and modulates the cellular stress response via HSPB2. Together, these pathways illustrate the complexity of LAM pathogenesis and the limitations of mTOR-targeted therapies. Abbreviations: CIB2 = calcium and integrin-binding protein 2; HSPB2 = heat shock protein family; LAM = lymphangioleiomyomatosis; RARβ = retinoic acid receptor; TFEB = transcription factor EB.

Hormonal influences on mTOR and LAM progression

The significant female predominance in LAM has long suggested the potential role of hormonal signaling in disease pathogenesis. This has prompted numerous studies to investigate the effects of estrogen on relevant pathways. The exacerbation of LAM during pregnancy and its worsening with exogenous estrogen exposure support the hypothesis that estrogen drives disease pathophysiology.^33-35

Yu et al.³⁶ provided early evidence that estradiol and tamoxifen stimulate the growth of LAM-associated angiomyolipoma cells by activating both genomic and nongenomic signaling pathways. These pathways include the mTOR and MAPK cascades, which synergistically drive cell proliferation and enhance survival. Beyond its proliferative effects, estrogen also facilitates pulmonary metastasis and enhances LAM cell survival.³⁷ Further investigations revealed that mTOR and estrogen-ERK2 signaling pathways converge to drive LAM progression. Estrogen activates ERK2 signaling, which, in turn, synergistically interacts with mTOR, amplifying LAM cell proliferation, survival, and invasiveness.³⁸ This hormone-mTOR cross talk emphasizes the complexity of LAM pathogenesis and its dependence on estrogenic signaling.

Yamnik and Holz³⁹ examined the molecular cross talk between estrogen and mTOR signaling and discovered that estrogen receptor activity is modulated by the MAPK/RSK and mTOR/S6K1 pathways. They showed that phosphorylation of the estrogen receptor induced via these pathways activates transcription and drives cell proliferation.

Alayev et al.⁴⁰ further found that mTORC1 can directly phosphorylate and activate estrogen receptor α, also confirming the role of mTORC1 in estrogen-mediated signaling and cell survival. This study further illustrates the importance of estrogen-mTOR cross talk in promoting LAM cell survival and propagation.

Importantly, in addition to its involvement in proliferation, estradiol drives metabolic reprogramming in LAM cells, providing them with the adaptive capacity to survive under stress conditions and maintain tumor growth. Sun et al.⁴¹ showed that estradiol acts to reactivate Akt, which upregulates glucose metabolism through the pentose phosphate pathway in mTORC1-hyperactive cells. This metabolic reprogramming enhances cell survival under oxidative and metabolic stress, providing LAM cells with a biological advantage in challenging environments.

Parallel oncogenic pathways

While mTOR signaling remains the primary focus of LAM research, accumulating evidence suggests that several parallel oncogenic pathways also contribute to disease progression. These pathways control cell proliferation, survival, invasion, metabolism, and immune interactions, making the molecular landscape of LAM more complex and challenging for designing targeted therapies.

One emerging pathway is histamine signaling, which has been linked to LAM cell proliferation and tissue remodeling. Herranz et al. demonstrated that mast cells, a major source of histamine, accumulate at the periphery of LAM nodules, where they interact with LAM cells and fibroblasts to promote microenvironmental changes.^42,43 Inhibiting mast cell activation has therefore been proposed as a potential strategy to disrupt these interactions and slow disease progression. These findings extend the molecular landscape of LAM and provide new avenues for biomarker and drug development.

The Notch pathway also plays a key role in LAM pathogenesis. Ma et al.⁴⁴ showed that mTOR regulates cell differentiation through a STAT3/p63/Jagged/Notch cascade, implicating STAT3 hyperactivation as a central event. Additionally, increased expression and activity of STAT3 in TSC2-deficient cells promote excessive proliferation and survival.⁴⁵ Beyond this, Notch1 has been found to directly activate Rheb, sustaining multipotency in TSC2-deficient cells and preventing differentiation.⁴⁶ Furthermore, another study revealed that TP53 polymorphisms altered the Notch-Nodal signaling axis, a pathway strongly linked to cellular plasticity and differentiation, suggesting that genetic variation may shape LAM cell behavior and disease severity.¹⁹

Another recently identified oncogenic driver is Gαq signaling, which acts in concert with mTORC1 to promote pathways governing proliferation, survival, and migration. Blocking Gαq signaling in preclinical models markedly reduced neoplastic behavior, positioning this pathway as a promising therapeutic target.⁴⁷

MicroRNAs (miRNAs) have emerged as important modulators of tumorigenesis in LAM. Liu et al.⁴⁸ identified miR-124 as a LAM cell growth modulator, showing that it directly targets retinoid X receptor alpha (RXRα) in TSC2-deficient cells. RXRα is a common heterodimer partner in retinoid signaling pathways, and its inhibition by miR-124 can disrupt RXRα-dependent transcriptional regulation associated with cell differentiation, growth, and apoptosis.

Taken together, histamine, Notch, Gαq, and miRNA signaling represent parallel pathways that extend beyond mTOR, reinforcing the concept of LAM as a multifaceted disease. Their involvement not only broadens the molecular landscape of LAM but also provides potential targets for biomarker development and therapeutic intervention.

Metabolic adaptations

LAM cells display distinct metabolic adaptations that sustain growth under nutrient-limited conditions. One mechanism is macropinocytosis, driven by Vps34, which enables scavenging of extracellular nutrients for anabolic growth.⁴⁹ In parallel, cytokine profiling has identified IL-6 as a driver of proliferation and migration in TSC2-deficient cells; by inducing PSAT1 expression, IL-6 reprograms serine metabolism to support biosynthesis and survival.⁵⁰ Constitutive mTORC1 signaling also enhances glycolysis via HIF1α and c-Myc and upregulates SREBP-dependent lipid and cholesterol biosynthesis, providing energy and membrane components for cell expansion.^51,52

Redox regulation adds another layer of adaptation. Glutaredoxin-1 (Grx1) maintains reactive oxygen species (ROS) homeostasis in TSC2-deficient cells, suppressing Bim-mediated apoptosis. In addition, activation of the COX2/PGE2/ERK axis reinforces proliferation and survival.⁵³ Loss of Grx1 conversely elevates ROS and restores apoptotic signaling, suggesting that redox targeting may reestablish cell death pathways in LAM.

Extracellular matrix remodeling and invasion

The complexity of signaling pathways in LAM has triggered exploration of many interconnected mechanisms to facilitate the understanding of cell invasion and metastatic potential. Mechanistic studies showed that Src kinase activation, osteoprotegerin, and integrin-α1β1 enhance adhesion, survival, and motility of TSC2-deficient cells, including through mTORC1-independent pathways.^54-56 In addition to cell motility, extracellular matrix (ECM) remodeling is another prominent factor involved in LAM progression and contributes directly to cystic destruction of lung tissue. Cathepsin-K, a potent collagenase, is highly expressed in LAM lesions and promotes alveolar wall degradation.⁵⁷ In addition to cathepsin K, several matrix metalloproteinases (MMPs) have been implicated in disease progression.^58,59 Elevated activity of MMP-1, MMP-2, MMP-9, and MMP-14 has been reported in bronchoalveolar lavage fluid, serum, and lung lesions of LAM patients, where they contribute to basement membrane degradation and cyst wall thinning. Recent data also show that not only are MMP-2 and MMP-7 elevated in the serum of LAM and TSC-LAM patients but also their expression in LAM/TSC-deficient cells is regulated by TSC2 and linked to cell migration.^60,61 Importantly, LAM cells themselves produce these proteases, linking ECM destruction directly to the neoplastic population.

Invasiveness is further reinforced by epithelial-to-mesenchymal transition (EMT). Small extracellular vesicles released by LAM cells induce EMT in neighboring epithelial cells, increasing their migratory and invasive capacity.⁶² At the same time, disruption of cell-cycle regulation contributes to uncontrolled growth and invasion: an Rheb–AMPK–p27^Kip1 axis promotes proliferation independent of mTORC1,⁶³ while loss of tuberin disrupts E-cadherin localization, further driving EMT and migration.²²

In addition to local proliferation and ECM remodeling, LAM demonstrates features of a low-grade metastatic neoplasm. Circulating LAM cells have been detected in peripheral blood, urine, and chylous effusions of patients, where they retain defining markers such as smooth muscle actin, human melanoma black 45 (HMB-45), and loss of TSC2 expression.^4,64-66 Their presence indicates that disease progression involves not only localized cystic destruction but also systemic dissemination. Evidence from lung transplantation further supports this: LAM recurs in transplanted lungs, implying that circulating LAM cells can reseed new grafts and drive disease progression.^67,68 Genomic studies have provided further insights into the origin of disseminated LAM cells. Identical TSC2 mutations have been identified in pulmonary and uterine LAM lesions, suggesting that the uterus may serve as a source of circulating neoplastic cells.⁶⁹ This genetic evidence strengthens the concept of LAM as a metastatic-like disease capable of clonal spread. LAM is more than a localized lung disorder.

One of the defining features of LAM is the presence of abnormal lymphatic structures within pulmonary lesions, driven in part by upregulation of lymphangiogenic pathways. Vascular endothelial growth factor-D, a lymphangiogenic factor secreted by LAM cells, is elevated in serum of patients with LAM and serves as a diagnostic and disease-monitoring biomarker.^70,71 Vascular endothelial growth factor-D binds to VEGF receptor 3 (VEGFR-3) expressed on lymphatic endothelial cells, promoting lymphatic vessel formation and remodeling within the lung.^72,73 These changes facilitate dissemination of LAM cells through lymphatic channels. The increasing immunoreactivity of Prox1, lymphatic vessel endothelial hyaluronan receptor 1 (LYVE-1), and VEGFR-3 in LAM biopsies indicates lymphangiogenic remodeling and enrichment of lymphatic endothelial cells within LAM lesions.⁷⁴ Whether these lymphatic endothelial cells arise through recruitment, local expansion, or phenotypic plasticity remains unclear. Together, these findings indicate that VEGF-D–mediated lymphangiogenesis is not merely a diagnostic marker but also plays an active role in the pathobiology and dissemination of LAM. Serum VEGF-D levels decrease following treatment with mTOR inhibitors such as sirolimus and correlate with treatment response; however, VEGF-D primarily reflects disease burden and lymphatic involvement rather than serving as a definitive predictor of therapeutic efficacy.^9,75

Immune microenvironment as a driver of LAM progression

Emerging studies have begun to delineate the role of immune evasion in LAM. Liu et al.⁷⁶ showed that mTORC1 upregulates B7-H3/CD276, an immunologic checkpoint molecule that inhibits antitumor T-cell function. Although predominantly investigated in the context of cancer, these observations suggest that LAM cells might employ immune evasion strategies to evade immune surveillance and continue to grow unchecked, thus providing an avenue for contributing to immune suppression in the LAM microenvironment via mTOR hyperactivation. Supporting this, Osterburg et al.⁷⁷ demonstrated that LAM lesions express ligands for the natural killer (NK) cell-activating receptor NKG2D, specifically ULBP2 and ULBP3. Soluble forms of these ligands were elevated in the serum of LAM patients and correlated with greater lung function decline. This shedding of NKG2D ligands likely impairs NK cell-mediated cytotoxicity by downregulating the NKG2D receptor on NK cells in patients with LAM, further highlighting immune evasion as a pathogenic mechanism in this pathology. LAM cells closely interact with their immune environment. The chemokine CCL2 (MCP‑1) is expressed in approximately 70% of LAM nodules, with its production regulated by loss of TSC2 function, and acts as a chemoattractant for LAM cells, reinforcing local recruitment and expansion of lesions.^78,79 LAM cells also express several chemokine receptors, including CCR2, CCR7, CCR10, CXCR2, CXCR4, and CXCR1, which makes them responsive to these signals. Functional studies have shown that CCL2 directly stimulates the migration of TSC2-deficient LAM cells, indicating that chemokine pathways contribute to their spread.⁷⁸ Recent single-cell and spatial transcriptomic analyses further revealed that LAM cells recruit and polarize macrophages toward an immunosuppressive CD206⁺phenotype through CXCL12/CXCR4 signaling. Reprogramming these macrophages with the peptide RP-182 reduced TSC2-deficient tumor growth in preclinical models, highlighting immune modulation as a potential therapeutic approach in LAM.⁸⁰

Together, these studies emphasize the numerous interdependent signaling networks regulating LAM progression, including cell invasion, migration, proliferation, ECM remodeling, and immune regulation (Table 1, Figure 3).

Table 1.

Molecular pathways involved in LAM pathogenesis and potential therapeutic targets.

Pathway	Key findings	Potential therapeutic targets	References
Histamine signaling	Histamine and mast cells contribute to tissue remodeling, blocking mast cells may be therapeutic.	Mast cell stabilizers, histamine receptor blockers	Herranz et al. (2021)42
Notch pathway	STAT3 hyperactivation and Notch-Rheb loop maintain LAM cell multipotency and tumorigenicity.	Notch inhibitors, STAT3 inhibitors	Ma et al. (2010),44 Cho et al. (2017)46
Gαq signaling	Gαq signaling enhances oncogenic pathways; inhibition reduces neoplastic behavior.	Gαq inhibitors	Trefier et al. (2024)47
MicroRNAs	miR-124 targets RXRα, disrupting retinoid signaling and promoting tumorigenicity.	miRNA-based therapies targeting miR-124/RXRα	Liu et al. (2019)48
Grx1	Grx1 suppresses apoptosis via COX2/PGE2/ERK, enhancing cell survival.	COX2/PGE2/ERK pathway inhibitors	Feng et al. (2023)53
Gene network regulation	HOX-PBX gene network is critical for LAM cell survival; targeting it may reduce progression.	HOX-PBX disruptors	Olatoke et al. (2023)21
Epigenetic modifications	TSC1 gene silencing via promoter hypermethylation contributes to LAM progression.	Epigenetic drugs (DNA methylation inhibitors)	Wang et al. (2017)81
Metabolic adaptations	TSC2 deficiency upregulates macropinocytosis for extracellular nutrient scavenging.	Vps34 inhibitors (targeting macropinocytosis)	Filippakis et al. (2018)49
Cytokine signaling (IL-6)	IL-6 upregulates PSAT1, linking inflammation to metabolic adaptation in LAM cells.	IL-6 inhibitors, PSAT1-targeted therapy	Wang et al. (2021)50
Invasion and metastasis	Src kinase, OPG, and integrin-α1β1 promote invasion, migration, and adhesion.	Src kinase inhibitors, OPG blockade, integrin-targeting drugs	Tyryshkin et al. (2014),54 Steagall et al. (2013),55 Moir et al. (2012)56
ECM remodeling	Cathepsin-K degrades ECM, facilitating lung cyst formation and tissue destruction.	Cathepsin-K inhibitors	Chilosi et al. (2009)57
Cell cycle regulation	Rheb-AMPK-p27 axis promotes proliferation; loss of TSC2 triggers EMT and invasiveness.	AMPK activators, EMT inhibitors	Lacher et al. (2010),63 Barnes et al. (2010)22
Immune modulation	LAM cells evade immune detection through mTORC1-driven B7-H3/CD276 expression and reduced susceptibility to NK cell–mediated cytotoxicity due to altered stress ligand expression.	B7-H3/CD276-targeting immunotherapy, NK cell–activating therapies, restoration of stress ligand expression	Liu et al. (2023),76 Osterburg et al. (2016)77

Abbreviations: ECM = extracellular matrix; Grx1 = glutaredoxin-1; LAM = lymphangioleiomyomatosis; miRNA = microRNA; NK = natural killer; OPG = osteoprotegerin; RXRα = retinoid X receptor alpha.

Figure 3.

Molecular and microenvironmental drivers of lymphangioleiomyomatosis (LAM) progression. Lymphangioleiomyomatosis pathogenesis is driven by the interaction of cell-intrinsic alterations and the surrounding microenvironment. Within LAM nodules, abnormal smooth muscle–like cells interact with fibroblasts, macrophages, and mast cells, while also releasing extracellular vesicles that modulate neighboring cells. These interactions converge on dysregulated metabolic, hormonal, immune, extracellular matrix (ECM)–remodeling, and lymphangiogenic pathways, which collectively drive cystic destruction, invasion, and systemic dissemination.

Conclusions

Lymphangioleiomyomatosis is a complex, multifaceted disease driven by genetic mutations, dysregulated molecular pathways, and interactions with the immune and hormonal systems. The hallmark of TSC, hyperactivation of the mTOR pathway primarily due to TSC2 mutations, has provided a basis for targeted therapy. Sirolimus and other mTOR inhibitors have improved disease management by stabilizing lung function and slowing progression. However, these treatments remain palliative rather than curative, highlighting the need to explore alternative therapeutic strategies.

The genetic landscape of LAM extends beyond TSC1 and TSC2, with genes such as NR2F2 and VEZF1 revealing the disease’s heterogeneity and interpatient variability. Genomic profiling and single-cell multi-omics analyses have revealed heterogeneous LAM cell populations with distinct molecular signatures. This heterogeneity may influence treatment responses, emphasizing the importance of precision medicine based on individual genetics and cellular characteristics. Hormonal modulation, particularly the interplay between estrogen signaling and mTOR hyperactivation, has emerged as a potential therapeutic strategy. Clinical evidence remains inconclusive, with some studies suggesting that anti-estrogen therapies may slow disease progression, although concerns over their impact on premenopausal women have limited their adoption. Further research is needed to identify candidates for hormonal therapy and its role in mTOR inhibition.

While progress has been made in understanding LAM, critical gaps persist. The rarity of this disease limits large clinical trials, and its variability complicates treatment standardization. Although mTOR inhibitors have improved management, their limitations necessitate innovative strategies, including combinatorial approaches and metabolic modulation. From a translational perspective, molecular targets may be prioritized based on their mechanistic relevance to TSC2 loss, validation in human LAM tissue, and their ability to complement current mTOR-based therapies. Future research should integrate technologies such as single-cell sequencing and CRISPR-based gene editing to identify new therapeutic targets.

Author contributions

A.J., M.P.K., and C.M.C. conceived and wrote the manuscript.

Supplementary material

Supplementary material is available at American Journal of Respiratory Cell and Molecular Biology online.

Conflicts of interest

Please see the ICMJE disclosure forms, which have been provided as supplementary material. The authors declare no conflicts of interest related to this work.

Funding

None declared.