Paper Details

ABSTRACT

Background: MYC overexpression is a hallmark of progression from the precursor plasma cell disorder monoclonal gammopathy of undetermined significance (MGUS) to multiple myeloma (MM) and is a major driver of metabolic reprogramming. Among its metabolic effects is enhanced tryptophan catabolism through the kynurenine pathway, in which tryptophan is converted to kynurenine (Kyn) and subsequently to 3-hydroxy-kynurenine (3HK). Activation of this pathway has been implicated in tumor immune evasion through the generation of immunosuppressive kynurenine-based metabolites in multiple cancers. We therefore investigated whether quantitative alterations in tryptophan and kynurenine pathway metabolites within the bone marrow (BM) microenvironment differentiates MGUS from MM and if these levels are associated with progression risk of precursor plasma cell disorders like SMM to MM.

Methods: BM plasma levels of tryptophan and its downstream kynurenine pathway metabolites, Kyn and 3HK, were quantified by targeted liquid chromatography-tandem mass spectrometry using isotopically labeled internal standards in two independent cohorts of patients with MGUS and newly diagnosed MM (NDMM) (cohort 1: 49 MGUS, 50 NDMM; cohort 2: 50 MGUS, 64 NDMM). An independent cohort of 47 patients with smoldering multiple myeloma (SMM) was evaluated to determine the association between kynurenine pathway metabolite levels and time to progression (TTP) to symptomatic MM. To investigate the biological basis of altered tryptophan metabolism, human myeloma cell lines (HMCLs) were used to assess the effects of MYC inhibition and tryptophan deprivation on kynurenine pathway metabolism. In parallel, ex vivo healthy donor T cells were exposed to varying concentrations of kynurenine pathway metabolites, and T-cell activation, and exhaustion markers were quantified by multiparametric flow cytometry. Statistical analyses were performed using JMP version 18.0.

Results: BM plasma concentrations of tryptophan were significantly lower in patients with NDMM than in those with MGUS in both independent cohorts (cohort 1: 34 ± 1.8 vs. 40 ± 1.8 μM, p = 0.013; cohort 2: 35 ± 1.5 vs. 43 ± 1.7 μM, p = 0.0009). Similarly, kynurenine (Kyn) levels were reduced in NDMM compared with MGUS (cohort 1: 1.73 ± 0.17 vs. 2.28 ± 0.17 μM, p = 0.026; cohort 2: 1.77 ± 0.10 vs. 2.21 ± 0.11 μM, p = 0.004). In contrast, BM plasma 3HK concentrations were significantly increased in NDMM (cohort 1: 0.07 ± 0.01 vs. 0.02 ± 0.01 μM, p < 0.0001; cohort 2: 0.188 ± 0.02 vs. 0.06 ± 0.02 μM, p < 0.0001). Among patients with SMM, a 3HK/Kyn ratio above the cohort median was associated with a significantly shorter time to progression (TTP) to symptomatic MM compared with patients with ratios at or below the median (22 months vs. not reached, p = 0.04). HMCLs (RPMI-8226 and MM1S) recapitulated these metabolic changes by consuming extracellular tryptophan and producing and releasing 3HK into the extracellular milieu. Pharmacologic MYC inhibition reduced tryptophan uptake and both intracellular and extracellular 3HK production, while tryptophan deprivation similarly decreased 3HK production, supporting MYC-dependent regulation of the tryptophan-kynurenine metabolic pathway. Finally, ex vivo exposure of healthy donor CD3? T cells to increasing concentrations of kyn but not 3HK reduced expression of activation markers (CD25 and granzyme B) and increased expression of the exhaustion marker LAG-3 in both CD4? and CD8? fractions, consistent with impaired T-cell function.

Conclusions: Differences in metabolite levels that are associated with enhanced tryptophan metabolism through the kynurenine pathway, resulting in depletion of tryptophan and kyn and accumulation of 3HK, differentiate MGUS from MM. Furthermore, an elevated 3HK/Kyn ratio was associated with a shorter TTP to MM in SMM, while increasing kyn level exposure promoted ex vivo T-cell dysfunction. These findings support the tryptophan-kynurenine pathway as a potential link between metabolic remodeling and immune suppression in the disease progression of precursor disorders to MM. Further mechanistic studies are required to evaluate the tryptophan-metabolic pathway  as a future therapeutic target to prevent disease progression in precursor plasma cells disorders.