Targeting Tryptophan Catabolism in Cancer Immunotherapy Era: Challenges and Perspectives.

Metabolism of tryptophan (Trp), an essential amino acid, represent a major metabolic pathway that both promotes tumor cell intrinsic malignant properties as well as restricts antitumour immunity, thus emerging as a drug development target for cancer immunotherapy. Three cytosolic enzymes, namely indoleamine 2,3-dioxygenase 1 (IDO1), IDO2 and tryptophan 2,3-dioxygenase (TDO2), catalyzes the first-rate limiting step of the degradation of Trp to kynurenine (Kyn) and modulates immunity toward immunosuppression mainly through the aryl hydrocarbon receptor (AhR) activation in numerous types of cancer. By restoring antitumor immune responses and synergizing with other immunotherapies, the encouraging preclinical data of IDO1 inhibitors has dramatically failed to translate into clinical success when combined with immune checkpoints inhibitors, reigniting the debate of combinatorial approach. In this review, we i) provide comprehensive evidences on immunomodulatory role of the Trp catabolism metabolites that highlight this pathway as relevant target in immuno-oncology, ii)ii) discuss underwhelming results from clinical trials investigating efficacy of IDO1 inhibitors and underlying mechanisms that might have contributed to this failure, and finally, iii) discuss the current state-of-art surrounding alternative approaches of innovative antitumor immunotherapies that target molecules of Trp catabolism as well as challenges and perspectives in the era of immunotherapy.

IDO Targeting in Sarcoma: Biological and Clinical Implications.

Sarcomas are heterogeneous malignant mesenchymal neoplasms with limited sensitivity to immunotherapy. We recently demonstrated an increase in Kynurenine Pathway (KP) activity in the plasma of sarcoma patients treated with pembrolizumab. While the KP has already been described to favor immune escape through the degradation of L-Tryptophan and production of metabolites including L-Kynurenine, Indoleamine 2,3 dioxygenase (IDO1), a first rate-limiting enzyme of the KP, still represents an attractive therapeutic target, and its blockade had not yet been investigated in sarcomas. Using immunohistochemistry, IDO1 and CD8, expression profiles were addressed within 203 cases of human sarcomas. At a preclinical level, we investigated the modulation of the KP upon PDL1 blockade in a syngeneic model of sarcoma through mRNA quantification of key KP enzymes within the tumor. Furthermore, in order to evaluate the possible anti-tumor effect of IDO blockade in combination with PDL1 blockade, an innovative IDO inhibitor (GDC-0919) was used. Its effect was first assessed on Kynurenine to Tryptophan ratio at plasmatic level and also within the tumor. Following GDC-0919 treatment, alone or in combination with anti-PDL1 antibody, tumor growth, immune cell infiltration, and gene expression profiling were measured. IDO1 expression was observed in 39.1% of human sarcoma cases and was significantly higher in tumors with high CD8 infiltration. In the pre-clinical setting, blockade of PDL1 led to a strong anti-tumor effect and was associated with an intratumoral inflammatory cytokines signature driven by Ifng but also with a modulation of the KP enzymes including Ido1 and Ido2. IDO1 inhibition using GDC-0919 resulted in (i) a significant decrease of plasmatic Kynurenine to Tryptophan ratio and in (ii) a decrease of tumoral Kynurenine. However, GDC-0919 used alone or combined with anti-PDL1, did not show anti-tumoral activity and did not affect the tumor immune cell infiltrate. In order to elucidate the mechanism(s) underlying the lack of effect of GDC-0919, we analyzed the gene expression profile of intratumoral biopsies. Interestingly, we have found that GDC-0919 induced a downregulation of the expression of pvr and granzymes, and an upregulation of inhba and Dtx4 suggesting a potential role of the IDO pathway in the control of NK function.

Accumulation of an endogenous tryptophan-derived metabolite in colorectal and breast cancers.

Tumor immune escape mechanisms are being regarded as suitable targets for tumor therapy. Among these, tryptophan catabolism plays a central role in creating an immunosuppressive environment, leading to tolerance to potentially immunogenic tumor antigens. Tryptophan catabolism is initiated by either indoleamine 2,3-dioxygenase (IDO-1/-2) or tryptophan 2,3-dioxygenase 2 (TDO2), resulting in biostatic tryptophan starvation and l-kynurenine production, which participates in shaping the dynamic relationship of the host's immune system with tumor cells. Current immunotherapy strategies include blockade of IDO-1/-2 or TDO2, to restore efficient antitumor responses. Patients who might benefit from this approach are currently identified based on expression analyses of IDO-1/-2 or TDO2 in tumor tissue and/or enzymatic activity assessed by kynurenine/tryptophan ratios in the serum. We developed a monoclonal antibody targeting l-kynurenine as an in situ biomarker of IDO-1/-2 or TDO2 activity. Using Tissue Micro Array technology and immunostaining, colorectal and breast cancer patients were phenotyped based on l-kynurenine production. In colorectal cancer l-kynurenine was not unequivocally associated with IDO-1 expression, suggesting that the mere expression of tryptophan catabolic enzymes is not sufficiently informative for optimal immunotherapy.

A new drug candidate (GEMSP) for multiple sclerosis.

GEMSP is a mixture of functional polypeptides: fatty acids linked to poly-L-Lysine (PL), antioxidants linked to PL, free radical scavengers linked to PL, and amino acids linked to PL (patent numbers 6114388 (USA) and 792167 (EU)). In this review, we update the data on this new drug reported in the literature. There is evidence suggesting that GEMSP is a good candidate for the treatment of multiple sclerosis (MS), an inflammatory and neurodegenerative disease of the central nervous system characterized by focal leukocyte inflammation, demyelization and axonal degeneration, resulting in nerve cell dysfunction. Experimental autoimmune encephalomyelitis (EAE) is the main animal model used in the study of MS, a T cell-mediated autoimmune disease of the central nervous system. EAE has many clinical and histopathological similarities to MS. In this model, preclinical studies on GEMSP have demonstrated that the drug strongly inhibits brain leukocyte infiltration and completely abolishes EAE episodes and clinical scores, and it also appears that GEMSP preserves myelin integrity. In general, treatment with the free constituents of GEMSP (not linked to the inert carrier protein) is poorly active against brain leukocyte infiltration in EAE-immunized animals. This means that free molecules (not linked to PL) exert a very poor action on such infiltration and that these molecules are either rapidly incorporated into the metabolism or are degraded. Moreover, with immunocytochemical techniques, it has been demonstrated that one component of GEMSP, the methionine compound, is stored inside the motoneurons of the ventral horn of the spinal cord. However, this component of GEMSP has not been found in the brain. The new candidate for MS therapy has shown no toxicity either in experimental animals or in humans. An open clinical trial in humans has demonstrated that GEMSP is completely safe. In addition, the approved drugs for the treatment of MS exert marked side effects, but no side effects have been reported following the administration of GEMSP. The results obtained at six months of treatment with low doses of GEMSP (0.75 mg/day) in that open clinical trial in humans were as follows: 55% of the patients maintained a stable expanded disability status scale (EDSS) value and 18% of the patients had a decreased EDSS value instead of a normal progression of 0.25 point on the mean EDSS scale. We focus our review on the following topics: 1) EAE models and clinical evaluation; 2) the synthesis of GEMSP; 3) the effects of GEMSP dosage on EAE; 4) the effects of GEMSP on brain leukocyte infiltration; 5) GEMSP inside motoneurons; 6) the role of the components of GEMSP; and 7) GEMSP in MS patients, GEMSP toxicity, and side effects. In conclusion, all the data reported indicate that GEMSP is a new potential drug candidate for the treatment of MS.

Gemals, a new drug candidate, extends lifespan and improves electromyographic parameters in a rat model of amyotrophic lateral sclerosis.

Amyotrophic lateral sclerosis (ALS) is a fatal disease involving selective and progressive degeneration and death of motor neurons. ALS is a multifactorial disease in which oxidative stress, glutamate excitotoxicity, intracellular aggregates, neurofilamentous disorganization, zinc excitotoxicity, mitochondrial damage, neuroinflammation, abnormalities in growth factors and apoptosis play a role. Any therapeutic approach to delay or stop the evolution of ALS should therefore ideally target these multiple pathways leading to motor neuron death. We have developed a combination therapy (Gemals) composed of functional polypeptides (fatty acids, free radical scavengers and amino acids linked to poly-L-lysine), chosen according to their known potentiality for regeneration or protection of neuronal components such as myelin, axon transport and mitochondria. We found that Gemals significantly extended lifespan and improved electromyographic parameters in a SOD1(G93A) rat model. The use of two drug concentrations indicated a possible dose dependence. These initial findings open the way to further investigation necessary to validate this new drug as a candidate for ALS treatment.