Daratumumab in first-line treatment of patients with light chain amyloidosis and Mayo stage IIIb improves treatment response and overall survival.

Treatment of patients with Mayo stage IIIb light chain (AL) amyloidosis is still challenging, and the prognosis remains very poor. Mayo stage IIIb patients were excluded from the pivotal trial leading to the approval of daratumumab in combination with bortezomib-cyclophosphamide-dexamethasone. This retrospective, multicenter study evaluates the addition of daratumumab to first-line therapy in patients with newly diagnosed stage IIIb AL amyloidosis. In total, data from 119 consecutive patients were analyzed, 27 patients received an upfront treatment including daratumumab, 63 a bortezomibbased regimen without daratumumab, eight received therapies other than daratumumab or bortezomib and 21 pretreated patients or deceased prior to treatment were excluded. In the daratumumab group, median overall survival was not reached after a median follow-up time of 14.5 months, while it was significantly worse in the bortezomib- and the otherwise treated group (6.6 and 2.2 months, respectively) (P=0.002). Overall hematologic response rate at 2 and 6 months was better in the daratumumab group compared to the bortezomib group (59% vs. 37%, P=0.12, 67% vs. 41%, P=0.04, respectively). Landmark survival analyses revealed a significantly improved overall survival in patients with partial hematologic response or better, compared to non-responders. Cardiac response at 6 months was 46%, 21%, 0% in the daratumumab-, bortezomib- and otherwise treated groups, respectively (P=0.04). A landmark survival analysis revealed markedly improved overall survival in patients with cardiac very good partial response vs. cardiac non-responders (P=0.002). This study demonstrates for the first time the superiority of an upfront treatment with daratumumab over standard-of-care in stage IIIb AL amyloidosis.

Cell-intrinsic ceramides determine T cell function during melanoma progression.

Acid sphingomyelinase (Asm) and acid ceramidase (Ac) are parts of the sphingolipid metabolism. Asm hydrolyzes sphingomyelin to ceramide, which is further metabolized to sphingosine by Ac. Ceramide generates ceramide-enriched platforms that are involved in receptor clustering within cellular membranes. However, the impact of cell-intrinsic ceramide on T cell function is not well characterized. By using T cell-specific Asm- or Ac-deficient mice, with reduced or elevated ceramide levels in T cells, we identified ceramide to play a crucial role in T cell function in vitro and in vivo. T cell-specific ablation of Asm in Smpd1fl/fl/Cd4cre/+ (Asm/CD4cre) mice resulted in enhanced tumor progression associated with impaired T cell responses, whereas Asah1fl/fl/Cd4cre/+ (Ac/CD4cre) mice showed reduced tumor growth rates and elevated T cell activation compared to the respective controls upon tumor transplantation. Further in vitro analysis revealed that decreased ceramide content supports CD4+ regulatory T cell differentiation and interferes with cytotoxic activity of CD8+ T cells. In contrast, elevated ceramide concentration in CD8+ T cells from Ac/CD4cre mice was associated with enhanced cytotoxic activity. Strikingly, ceramide co-localized with the T cell receptor (TCR) and CD3 in the membrane of stimulated T cells and phosphorylation of TCR signaling molecules was elevated in Ac-deficient T cells. Hence, our results indicate that modulation of ceramide levels, by interfering with the Asm or Ac activity has an effect on T cell differentiation and function and might therefore represent a novel therapeutic strategy for the treatment of T cell-dependent diseases such as tumorigenesis.

Impact of time to diagnosis on Mayo stages, treatment outcome, and survival in patients with AL amyloidosis and cardiac involvement.

OBJECTIVE: To study the impact of time to diagnosis on cardiac Mayo stages, treatment outcome, and overall survival. METHODS: We retrospectively analyzed 77 consecutive patients diagnosed between 2015 and 2020 with AL amyloidosis and cardiac involvement. Medical history was recorded in standardized form with the help of a questionnaire. RESULTS: Time from onset of symptoms of cardiac failure to diagnosis was correlated with the severity of cardiac involvement in modified Mayo 2004 and revised Mayo 2012 staging systems (rs  = 0.30, 95% CI: 0.07-0.50, P = .007 and rs  = 0.25, 95% CI: 0.01-0.45, P = .03). Patients with advanced Mayo 2004 stages received reduced-intensity regimens and had a lower probability to achieve adequate hematologic- and cardiac response after first-line treatment than patients with early stages (rs  = 0.28, 95% CI: 0.04-0.48, P = .01 and rs  = 0.72, 95% CI: 0.55-0.82, P < .0001) and poorer overall survival (P = .0004). Compared with patients diagnosed within the first year, patients diagnosed after 13-18 or ≥19 months from first symptoms had a 3- to 5 times higher risk of dying. Our data indicate that there is a 12-month window within which the diagnosis of AL amyloidosis needs to be established to avoid early deterioration and death. CONCLUSIONS: Sensitizing physicians and raising awareness for the disease are crucial for timely diagnosis and may improve the outcome of the disease.

Characterization of the small molecule ARC39, a direct and specific inhibitor of acid sphingomyelinase in vitro.

Inhibition of acid sphingomyelinase (ASM), a lysosomal enzyme that catalyzes the hydrolysis of sphingomyelin into ceramide and phosphorylcholine, may serve as an investigational tool or a therapeutic intervention to control many diseases. Specific ASM inhibitors are currently not sufficiently characterized. Here, we found that 1-aminodecylidene bis-phosphonic acid (ARC39) specifically and efficiently (>90%) inhibits both lysosomal and secretory ASM in vitro. Results from investigating sphingomyelin phosphodiesterase 1 (SMPD1/Smpd1) mRNA and ASM protein levels suggested that ARC39 directly inhibits ASM's catalytic activity in cultured cells, a mechanism that differs from that of functional inhibitors of ASM. We further provide evidence that ARC39 dose- and time-dependently inhibits lysosomal ASM in intact cells, and we show that ARC39 also reduces platelet- and ASM-promoted adhesion of tumor cells. The observed toxicity of ARC39 is low at concentrations relevant for ASM inhibition in vitro, and it does not strongly alter the lysosomal compartment or induce phospholipidosis in vitro. When applied intraperitoneally in vivo, even subtoxic high doses administered short-term induced sphingomyelin accumulation only locally in the peritoneal lavage without significant accumulation in plasma, liver, spleen, or brain. These findings require further investigation with other possible chemical modifications. In conclusion, our results indicate that ARC39 potently and selectively inhibits ASM in vitro and highlight the need for developing compounds that can reach tissue concentrations sufficient for ASM inhibition in vivo.

Dead Cells Induce Innate Anergy via Mertk after Acute Viral Infection.

Infections can result in a temporarily restricted unresponsiveness of the innate immune response, thereby limiting pathogen control. Mechanisms of such unresponsiveness are well studied in lipopolysaccharide tolerance; however, whether mechanisms of tolerance limit innate immunity during virus infection remains unknown. Here, we find that infection with the highly cytopathic vesicular stomatitis virus (VSV) leads to innate anergy for several days. Innate anergy is associated with induction of apoptotic cells, which activates the Tyro3, Axl, and Mertk (TAM) receptor Mertk and induces high levels of interleukin-10 (IL-10) and transforming growth factor ß (TGF-ß). Lack of Mertk in Mertk-/- mice prevents induction of IL-10 and TGF-ß, resulting in abrogation of innate anergy. Innate anergy is associated with enhanced VSV replication and poor survival after infection. Mechanistically, Mertk signaling upregulates suppressor of cytokine signaling 1 (SOCS1) and SOCS3. Dexamethasone treatment upregulates Mertk and enhances innate anergy in a Mertk-dependent manner. In conclusion, we identify Mertk as one major regulator of innate tolerance during infection with VSV.

A Novel Visible Range FRET Probe for Monitoring Acid Sphingomyelinase Activity in Living Cells.

Activity of acid sphingomyelinase has been implicated in a number of diseases like acute lung injury, sepsis or metastasis of melanoma cells. Here, we present a sphingomyelinase FRET probe based on FAM/BODIPY dyes for real-time monitoring of acid sphingomyelinase. The probe gives rise to a tremendous increase in fluorescence of the fluorescein FRET donor upon cleavage and we show that this is, to a significant part, due to cleavage-associated phase transition, suggesting a more systematic consideration of such effects for future probe development. The probe allows for the first time to monitor relative sphingomyelinase activities of intact living cells by flow cytometry.