Zvida‐Bloch, Tal

Abstract Introduction Systemic light chain (AL) amyloidosis is a clonal plasma cell disorder characterized by the deposition of misfolded immunoglobulin light chains in vital organs, often resulting in irreversible organ damage. While plasma cell-directed therapies have improved clinical outcomes, the molecular mechanisms driving disease pathogenesis remain poorly understood. Our previous studies identified overactivation of the PI3K/AKT signaling pathway in AL amyloidosis (Zvida et al., Blood, 2024; Fishov et al., Cancer Med., 2023), a central regulator of cellular and lipid metabolism. Dysregulated lipid metabolism has been implicated in several hematologic malignancies, yet its role in AL amyloidosis remains largely unexplored. Objectives To characterize the lipidomic landscape in AL amyloidosis compared to multiple myeloma (MM) and healthy controls (HC) in the context of inflammatory signaling. The ultimate goal of this work is to uncover novel metabolic-immunologic targets for therapeutic intervention. Methods Lipidomic profiling was performed by liquid chromatography-mass spectrometry (LC-MS) on plasma samples from AL patients (n=30), MM (n=30), and HCs (n=15). Fatty acid synthase (FASN) expression was validated by qRT-PCR in CD138+ bone marrow (BM) samples. A lipogenesis index was calculated using the ratio of palmitoleic acid to linoleic acid as a surrogate marker of FASN activity. Additionally, RNA-seq was conducted on CD138⁺ BM cells from AL and HC samples (n=8). Cytokine profiling was performed on serum from AL, MM, and HC subjects (n=10 each). Functional assays and lipidomic analysis were conducted in ALMC1 cells using the FASN inhibitor TVB-2640 and the MEK inhibitor cobimetinib. Results Lipidomic analysis of pre-treatment plasma samples revealed a significant increase in the lipogenesis index in AL patients, indicating elevated FASN activity. This was consistent with increased FASN mRNA expression in bone marrow CD138⁺ cells, suggesting both transcriptional upregulation and enzymatic activation. In ALMC1 cells, dose- and time-dependent FASN inhibition with TVB-2640 (0-2 μM, 24-72h) significantly reduced cell viability (IC50=1.3 μM), suggesting that FASN activity supports AL plasma cell survival. In addition, AL patients demonstrated a significant decrease in polyunsaturated fatty acids (PUFAs) levels, including docosahexaenoic acid (DHA), eicosapentaenoic acid (EPA), docosapentaenoic acid (DPA), and linoleic acid (LA), compared to MM and HC. As PUFAs are known suppressors of FASN expression and activity, their depletion may contribute to sustained FASN upregulation and a metabolic shift toward de novo lipogenesis. Pathway enrichment analysis of patient lipidomic data revealed significant enrichment of omega-3/6 fatty acid metabolism, ATP generation, and inflammation-related pathways. Notably, imbalances in omega-3 and omega-6 fatty acids ratio have been associated with heightened inflammatory responses. In parallel, RNA-seq analysis identified several upregulated pathways in AL amyloidosis patients compared to controls, including immune regulation, MAPK signaling, mitochondrial dysfunction, oxidative phosphorylation, and IL-13 signaling. Both IL-13 and MAPK are known to transcriptionally induce FASN and regulate immune responses, providing a possible mechanistic link between inflammation and lipid remodeling. Cytokine profiling revealed elevated levels of IL-1β, IL-13, IL-8, IFN-γ, MIP-1α (CCL3), G-CSF, MCP-1 (CCL2), and IP-10 (CXCL10) in AL serum samples, consistent with a systemic inflammatory phenotype. Pharmacologic inhibition of MAPK signaling (by cobimetinib) in ALMC1 cells led to restoration of PUFA levels and a concurrent reduction in cytokine secretion, suggesting that MAPK activity sustains both the inflammatory and lipogenic phenotype in AL. ConclusionsOur findings identify metabolic-inflammatory axis in AL amyloidosis, in which IL-13 and MAPK signaling may promote fatty acid reprogramming through FASN activation and PUFA suppression. This axis sustains a pro-inflammatory environment that may contribute to disease pathogenesis. Thus, FASN and MAPK may represent promising metabolic-immunologic targets for therapeutic intervention in AL amyloidosis.

Summary Amyloid light‐chain (AL) amyloidosis is a systemic clonal plasma cell disorder characterized by the production and deposition of misfolded immunoglobulin light chains (LCs), resulting in multiorgan dysfunction. Due to its intricate molecular mechanisms and diverse organ involvement, the disease poses significant diagnostic and therapeutic challenges. This review explores the molecular landscape of AL amyloidosis, emphasizing genetic, transcriptomic and proteomic alterations. Key findings include chromosomal abnormalities, somatic mutations, aberrant gene expression, disrupted protein folding pathways and the role of cytokine and chemokine secretion. These factors collectively drive the overproduction and destabilization of amyloidogenic LCs, leading to organ‐specific amyloid deposition, clinical heterogeneity and variable patient outcomes. Despite therapeutic advancements, the disease's complexity challenges the development of effective biological models. Progressing towards personalized therapies requires the development of preclinical models and the identification of biomarkers and molecular data to design targeted interventions. This review highlights the importance of integrating DNA, RNA and protein‐level analyses to deepen the understanding of AL amyloidosis pathogenesis. Such insights are pivotal for improving diagnostics, prognostics and therapeutic strategies, ultimately advancing precision medicine for this challenging disease.

Background: Systemic light chain (AL) amyloidosis is a clonal plasma cell disorder characterized by the deposition of misfolded immunoglobulin light chain products in vital organs, leading to significant organ dysfunction. The molecular mechanisms underlying its pathogenesis remain incompletely elucidated. Identifying the molecular mechanisms and novel therapeutic targets is critical for improving treatment strategies and patient outcomes. Our previous research work identified that the PI3K/AKT signaling pathway is overexpressed in AL amyloidosis (Zvida et al., Blood, 2022, Fishov et al., Cancer Med., 2023). This pathway plays a critical role in regulating cellular metabolism. Specifically activated AKT can phosphorylate and regulate various target proteins involved in metabolic processes. One of the significant targets is fatty acid synthase (FASN), an enzyme responsible for the synthesis of fatty acids. AKT can enhance FASN expression and activity, thereby promoting lipid biosynthesis. Additionally, elevated FASN levels can drive inflammation through increased fatty acid synthesis, which activates inflammatory signaling pathways and enhances cytokine production (Acosta-Martinez et al.,IJMC, 2022). Elevated levels of FASN have been associated with hematological malignancies and cell survival (Vanauberg, et al., Oncogenesis, 2023). However, its role in AL amyloidosis is yet to be elucidated. Aim: To determine the role of FASN in the pathology and inflammatory response in AL amyloidosis, and to evaluate the potential of targeting FASN and the PI3K/AKT pathway for therapeutic intervention. Methods: We profiled FASN expression in CD138+ bone marrow (BM) samples from patients with AL amyloidosis (n=20), Multiple Myeloma (MM) (n=20), Monoclonal Gammopathy of Undetermined (MGUS) (n=8) and Healthy Controls (HC) (n=6), using qRT-PCR. To further explore the relationship between the PI3K/AKT pathway and FASN, we treated ALMC-1 cells with Capivasertib (an AKT inhibitor) and assessed FASN protein levels, cell proliferation by WST-1 assay, cell cycle distribution by flow cytometry, light chain secretion by ELISA, and expression of AKT and its downstream targets via western blot analysis. Furthermore, TVB-2640 (FASN inhibitor) was used to evaluate the cell response to FASN inhibition. Results: Our analysis revealed a significant upregulation of FASN in AL amyloidosis BM samples compared to MM, MGUS, and HCs. Capivasertib treatment resulted in a dose-dependent reduction in FASN protein levels. Additionally, Capivasertib treatment inhibited cell proliferation, induced G1 phase cell cycle arrest, reduced light chain secretion, and decreased AKT protein expression and its downstream targets p4EBP pS6 and pERK. Elevated levels of proinflammatory cytokines, including TNF and IP-10 (CXCL-10), were observed in AL amyloidosis serum samples, suggesting the involvement of immune mechanisms and the PI3K/AKT pathway, potentially induced by FASN. Finally, treatment with TVB-2640 reduced cell viability and decreased the protein expression of LIPIN-1, which is one of the key target genes regulated by FASN. Conclusions: Our findings highlight the significant upregulation of FASN in AL amyloidosis and its regulation by the PI3K/AKT pathway. The observed effects of Capivasertib on FASN expression and cell proliferation, along with the upregulation of inflammatory cytokines, underscore the potential for targeting the PI3K/AKT signaling pathway and FASN in developing more specific therapeutic strategies for AL amyloidosis.