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Renal cell carcinomas associated with hereditary leiomyomatosis and renal cell cancer (HLRCC) are notoriously aggressive and represent the leading cause of death among patients with HLRCC. To date, a safe and effective standardized therapy for this tumor type is lacking. Here we show that the engineered synthetic therapeutic enzyme, Cyst(e)inase, when combined with rapamycin, can effectively induce ferroptosis in HLRCC cells in vivo. The drug combination promotes lipid peroxidation to a greater degree than cysteine deprivation or Cyst(e)inase treatment alone, while rapamycin treatment alone does not induce ferroptosis. Mechanistically, Cyst(e)inase induces ferroptosis by depleting the exogenous cysteine/cystine supply, while rapamycin reduces cellular ferritin level by promoting ferritins' destruction via ferritinophagy. Since both Cyst(e)inase and rapamycin are well tolerated clinically, the combination represents an opportunity to exploit ferroptosis induction as a cancer management strategy. Accordingly, using a xenograft mouse model, we showed that the combination treatment resulted in tumor growth suppression without any notable side effects. In contrast, both Cyst(e)inase only and rapamycin only treatment groups failed to induce a significant change when compared with the vehicle control group. Our results demonstrated the effectiveness of Cyst(e)inase–rapamycin combination in inducing ferroptotic cell death in vivo, supporting the potential translation of the combination therapy into clinical HLRCC management.

Supplementary Table from Cyst(e)inase–Rapamycin Combination Induces Ferroptosis in Both In Vitro and In Vivo Models of Hereditary Leiomyomatosis and Renal Cell Cancer

Supplementary Data from Cyst(e)inase–Rapamycin Combination Induces Ferroptosis in Both In Vitro and In Vivo Models of Hereditary Leiomyomatosis and Renal Cell Cancer

Chromophobe (Ch) renal cell carcinoma (RCC) arises from the intercalated cell in the distal nephron. There are no proven treatments for metastatic ChRCC. A distinguishing characteristic of ChRCC is strikingly high levels of reduced (GSH) and oxidized (GSSG) glutathione. Here, we demonstrate that ChRCC-derived cells exhibit higher sensitivity to ferroptotic inducers compared with clear-cell RCC. ChRCC-derived cells are critically dependent on cystine via the cystine/glutamate antiporter xCT to maintain high levels of glutathione, making them sensitive to inhibitors of cystine uptake and cyst(e)inase. Gamma-glutamyl transferase 1 (GGT1), a key enzyme in glutathione homeostasis, is markedly suppressed in ChRCC relative to normal kidney. Importantly, GGT1 overexpression inhibits the proliferation of ChRCC cells in vitro and in vivo, suppresses cystine uptake, and decreases levels of GSH and GSSG. Collectively, these data identify ferroptosis as a metabolic vulnerability in ChRCC, providing a potential avenue for targeted therapy for these distinctive tumors.

The Trp metabolite kynurenine (KYN) accumulates in numerous solid tumours and mediates potent immunosuppression. Bacterial kynureninases (KYNases), which preferentially degrade kynurenine, can relieve immunosuppression in multiple cancer models, but immunogenicity concerns preclude their clinical use, while the human enzyme (HsKYNase) has very low activity for kynurenine and shows no therapeutic effect. Using fitness selections, we evolved a HsKYNase variant with 27-fold higher activity, beyond which exploration of >30 evolutionary trajectories involving the interrogation of >10(9) variants led to no further improvements. Introduction of two amino acid substitutions conserved in bacterial KYNases reduced enzyme fitness but potentiated rapid evolution of variants with ~500-fold improved activity and reversed substrate specificity, resulting in an enzyme capable of mediating strong anti-tumour effects in mice. Pre-steady-state kinetics revealed a switch in rate-determining step attributable to changes in both enzyme structure and conformational dynamics. Apart from its clinical significance, our work highlights how rationally designed substitutions can potentiate trajectories that overcome barriers in protein evolution.

Dynamic motions of enzymes occurring on a broad range of timescales play a pivotal role in all steps of the reaction pathway, including substrate binding, catalysis, and product release. However, it is unknown whether structural information related to conformational flexibility can be exploited for the directed evolution of enzymes with higher catalytic activity. Here, we show that mutagenesis of residues exclusively located at flexible regions distal to the active site of Homo sapiens kynureninase (HsKYNase) resulted in the isolation of a variant (BF-HsKYNase) in which the rate of the chemical step toward kynurenine was increased by 45-fold. Mechanistic pre–steady-state kinetic analysis of the wild type and the evolved enzyme shed light on the underlying effects of distal mutations (10 Å from the active site) on the rate-limiting step of the catalytic cycle. Hydrogen-deuterium exchange coupled to mass spectrometry and molecular dynamics simulations revealed that the amino acid substitutions in BF-HsKYNase allosterically affect the flexibility of the pyridoxal-5′-phosphate (PLP) binding pocket, thereby impacting the rate of chemistry, presumably by altering the conformational ensemble and sampling states more favorable to the catalyzed reaction.

Renal cell carcinomas associated with hereditary leiomyomatosis and renal cell cancer (HLRCC) are notoriously aggressive and represent the leading cause of death among patients with HLRCC. To date, a safe and effective standardized therapy for this tumor type is lacking. Here we show that the engineered synthetic therapeutic enzyme, Cyst(e)inase, when combined with rapamycin, can effectively induce ferroptosis in HLRCC cells in vivo. The drug combination promotes lipid peroxidation to a greater degree than cysteine deprivation or Cyst(e)inase treatment alone, while rapamycin treatment alone does not induce ferroptosis. Mechanistically, Cyst(e)inase induces ferroptosis by depleting the exogenous cysteine/cystine supply, while rapamycin reduces cellular ferritin level by promoting ferritins' destruction via ferritinophagy. Since both Cyst(e)inase and rapamycin are well tolerated clinically, the combination represents an opportunity to exploit ferroptosis induction as a cancer management strategy. Accordingly, using a xenograft mouse model, we showed that the combination treatment resulted in tumor growth suppression without any notable side effects. In contrast, both Cyst(e)inase only and rapamycin only treatment groups failed to induce a significant change when compared with the vehicle control group. Our results demonstrated the effectiveness of Cyst(e)inase–rapamycin combination in inducing ferroptotic cell death in vivo, supporting the potential translation of the combination therapy into clinical HLRCC management.

Cancer immunotherapy restores or enhances the effector function of CD8+ T cells in the tumour microenvironment1,2. CD8+ T cells activated by cancer immunotherapy clear tumours mainly by inducing cell death through perforin-granzyme and Fas-Fas ligand pathways3,4. Ferroptosis is a form of cell death that differs from apoptosis and results from iron-dependent accumulation of lipid peroxide5,6. Although it has been investigated in vitro7,8, there is emerging evidence that ferroptosis might be implicated in a variety of pathological scenarios9,10. It is unclear whether, and how, ferroptosis is involved in T cell immunity and cancer immunotherapy. Here we show that immunotherapy-activated CD8+ T cells enhance ferroptosis-specific lipid peroxidation in tumour cells, and that increased ferroptosis contributes to the anti-tumour efficacy of immunotherapy. Mechanistically, interferon gamma (IFNγ) released from CD8+ T cells downregulates the expression of SLC3A2 and SLC7A11, two subunits of the glutamate-cystine antiporter system xc-, impairs the uptake of cystine by tumour cells, and as a consequence, promotes tumour cell lipid peroxidation and ferroptosis. In mouse models, depletion of cystine or cysteine by cyst(e)inase (an engineered enzyme that degrades both cystine and cysteine) in combination with checkpoint blockade synergistically enhanced T cell-mediated anti-tumour immunity and induced ferroptosis in tumour cells. Expression of system xc- was negatively associated, in cancer patients, with CD8+ T cell signature, IFNγ expression, and patient outcome. Analyses of human transcriptomes before and during nivolumab therapy revealed that clinical benefits correlate with reduced expression of SLC3A2 and increased IFNγ and CD8. Thus, T cell-promoted tumour ferroptosis is an anti-tumour mechanism, and targeting this pathway in combination with checkpoint blockade is a potential therapeutic approach.

Pancreatic ductal adenocarcinoma (PDA) is the third-leading cause of cancer mortality in the US and is highly resistant to classical, targeted, and immune therapies. We show that human PDA cells are dependent on the provision of exogenous cystine to avert a catastrophic accumulation of lipid reactive oxygen species (ROS) that, left unchecked, leads to ferroptotic cell death, bothin vitroandin vivo. Using a dual-recombinase genetically engineered model, we found that acute deletion ofSlc7a11led to tumor-selective ferroptosis, tumor stabilizations/regressions, and extended overall survival. The mechanism of ferroptosis induction in PDA cells required the concerted depletion of both glutathione and coenzyme A, highlighting a novel branch of ferroptosis-relevant metabolism. Finally, we found that cystine depletionin vivousing the pre-IND agent cyst(e)inase phenocopiedSlc7a11deletion, inducing tumor-selective ferroptosis and disease stabilizations/regressions in the well-validated KPC model of PDA.One Sentence SummaryGenetic and pharmacological targeting of cystine import induces pancreatic cancer-selective ferroptosisin vivo.

Ferroptotic cell death and cancer Cell death can occur through different mechanisms, several of which are being explored as potential targets for cancer treatment. One form of cell death that has attracted recent interest is ferroptosis, which is triggered by high intracellular levels of lipid reactive oxygen species. Pancreatic cancer cells have high levels of reactive oxygen species but manage to avoid ferroptosis by importing extracellular cysteine. Studying mice bearing pancreatic tumors, Badgley et al. found that administration of a drug inhibiting cysteine import induced tumor-selective ferroptosis and inhibited tumor growth. Further work will be required to determine whether this therapeutic strategy will be effective in human pancreatic cancer, a tumor type for which new treatments are urgently needed. Science , this issue p. 85

Ferroptosis is a form of regulated cell death involving lethal peroxidation of phospholipids (Hirschhorn and Stockwell, 2018). Recent results reveal that ferroptosis mediates the tumor suppressive activity of interferon gamma secreted by CD8(+) T cells in response to immune checkpoint blockade, suggesting the immune system may function in part through ferroptosis to prevent tumorigenesis (Wang et al., 2019).

Increased tryptophan (Trp) catabolism in the tumor microenvironment (TME) can mediate immune suppression by upregulation of interferon (IFN)-γ-inducible indoleamine 2,3-dioxygenase (IDO1) and/or ectopic expression of the predominantly liver-restricted enzyme tryptophan 2,3-dioxygenase (TDO). Whether these effects are due to Trp depletion in the TME or mediated by the accumulation of the IDO1 and/or TDO (hereafter referred to as IDO1/TDO) product kynurenine (Kyn) remains controversial. Here we show that administration of a pharmacologically optimized enzyme (PEGylated kynureninase; hereafter referred to as PEG-KYNase) that degrades Kyn into immunologically inert, nontoxic and readily cleared metabolites inhibits tumor growth. Enzyme treatment was associated with a marked increase in the tumor infiltration and proliferation of polyfunctional CD8+ lymphocytes. We show that PEG-KYNase administration had substantial therapeutic effects when combined with approved checkpoint inhibitors or with a cancer vaccine for the treatment of large B16-F10 melanoma, 4T1 breast carcinoma or CT26 colon carcinoma tumors. PEG-KYNase mediated prolonged depletion of Kyn in the TME and reversed the modulatory effects of IDO1/TDO upregulation in the TME.

Despite the sustained clinical benefit demonstrated by immune checkpoint inhibitors, a majority of patients derive minimal or no appreciable benefit, indicating the urgent need to incorporate novel immunomodulatory targets and therapeutic strategies. Indoleamine 2,3-dioxygenase 1 (IDO1) and tryptophan 2,3-dioxygenase 2 (TDO2) catalyze the first and rate-limiting step in the immunosuppressive tryptophan/kynurenine pathway and are both upregulated in a number of tumor types. Although small-molecule IDO1 inhibitors are being clinically evaluated in several tumor types, so far they have not demonstrated significant clinical benefits either as a single agent or in combination with immune checkpoint inhibition. We are developing pegylated kynureninase (Kynase), a kynurenine degrading enzyme, to treat a broader population with IDO1 and/or TDO2 expressing tumors. We believe that a more robust antitumor immune response can be achieved by depleting kynurenine, produced by both IDO1 and TDO2, with Kynase, than by inhibiting only IDO1. The human Kynase (HsKYN) has been successfully engineered to exhibit vastly improved catalytic activity and stability toward kynurenine over the wild-type human enzyme. HsKYN achieved durable and near complete plasma kynurenine depletion in mice, rats and non-human primates. HsKYN demonstrated single agent efficacy in CT26, MC38 and B16-IDO syngeneic mouse models. Tumor gene expression analysis using NanoString revealed that HsKYN treatment upregulated T-cell and NK cell activation signature. More importantly, HsKYN significantly increased the tumor-infiltrating CD8 T-cells and their activation/polyfunctionality, and reduced the Treg population. As a direct comparison, the lead IDO1 inhibitor epacadostat did not impose any meaningful effects on the same immune cell populations. Furthermore, HsKYN showed beneficial combination efficacy with anti-PD-1 that was superior to combined Epacadostat / anti-PD-1. Evidence to date suggest that HsKYN is well tolerated in multiple species. Therefore, immunoprofiling, efficacy and safety results strongly support that Kynase is a more effective therapeutic approach than IDO1 inhibition. HsKYN is moving toward clinical development for treatment of cancers where IDO1 and/or TDO2 pathways play a significant immunosuppressive role. Citation Format: Silvia Coma, Jillian Cavanaugh, James Nolan, Jeremy Tchaicha, Karen McGovern, Everett Stone, John Blazeck, Candice Lamb, George Georgiou, Mark G Manfredi, Michelle Zhang. Treatment of IDO1 and TDO2 positive tumors with a kynurenine-degrading enzyme: A highly differentiated approach from IDO1 inhibition [abstract]. In: Proceedings of the Fourth CRI-CIMT-EATI-AACR International Cancer Immunotherapy Conference: Translating Science into Survival; Sept 30-Oct 3, 2018; New York, NY. Philadelphia (PA): AACR; Cancer Immunol Res 2019;7(2 Suppl):Abstract nr B008.

Cancer cells experience higher oxidative stress from reactive oxygen species (ROS) than do non-malignant cells because of genetic alterations and abnormal growth; as a result, maintenance of the antioxidant glutathione (GSH) is essential for their survival and proliferation. Under conditions of elevated ROS, endogenous L-cysteine (L-Cys) production is insufficient for GSH synthesis. This necessitates uptake of L-Cys that is predominantly in its disulfide form, L-cystine (CSSC), via the xCT(-) transporter. We show that administration of an engineered and pharmacologically optimized human cyst(e)inase enzyme mediates sustained depletion of the extracellular L-Cys and CSSC pool in mice and non-human primates. Treatment with this enzyme selectively causes cell cycle arrest and death in cancer cells due to depletion of intracellular GSH and ensuing elevated ROS; yet this treatment results in no apparent toxicities in mice even after months of continuous treatment. Cyst(e)inase suppressed the growth of prostate carcinoma allografts, reduced tumor growth in both prostate and breast cancer xenografts and doubled the median survival time of TCL1-Tg:p53-/- mice, which develop disease resembling human chronic lymphocytic leukemia. It was observed that enzyme-mediated depletion of the serum L-Cys and CSSC pool suppresses the growth of multiple tumors, yet is very well tolerated for prolonged periods, suggesting that cyst(e)inase represents a safe and effective therapeutic modality for inactivating antioxidant cellular responses in a wide range of malignancies.

Arginases catalyze the divalent cation-dependent hydrolysis of l-arginine to urea and l-ornithine. There is significant interest in using arginase as a therapeutic anti-neogenic agent against l-arginine auxotrophic tumors and in enzyme replacement therapy for treating hyperargininemia. Both therapeutic applications require enzymes with sufficient stability under physiological conditions. To explore sequence elements that contribute to arginase stability we used SCHEMA-guided recombination to design a library of chimeric enzymes composed of sequence fragments from the two human isozymes Arginase I and II. We then developed a novel active learning algorithm that selects sequences from this library that are both highly informative and functional. Using high-throughput gene synthesis and our two-step active learning algorithm, we were able to rapidly create a small but highly informative set of seven enzymatically active chimeras that had an average variant distance of 40 mutations from the closest parent arginase. Within this set of sequences, linear regression was used to identify the sequence elements that contribute to the long-term stability of human arginase under physiological conditions. This approach revealed a striking correlation between the isoelectric point and the long-term stability of the enzyme to deactivation under physiological conditions.

The tryptophan/kynurenine pathway has been clinically validated in several tumor types with small-molecule IDO1 inhibitors in combination with checkpoint inhibition. Indoleamine-pyrrole 2,3-dioxygenase 1 (IDO1) and tryptophan 2,3-dioxygenase 2 (TDO2) are upregulated in a number of tumor types, metabolizing tryptophan to form immunosuppressive kynurenine. We are developing Kynureninase (Kynase), a kynurenine depleting enzyme, to treat IDO1 and TDO2 positive tumors. The human Kynase has been successfully engineered to vastly increase catalytic activity and stability toward kynurenine over the wild type enzyme. In mice, Kynase achieved prolonged Kynurenine degradation (≥5 days) in plasma and tumor draining lymph node (TDLN), leading to anti-tumor activity as a single agent and in combination with check point inhibitors in mouse syngeneic tumor models. Kynase demonstrated superior tumor growth inhibition and survival benefit relative to a leading IDO1 inhibitor epacadostat in these models. The effects of Kynase on a number of immune cell types, both in vitro and in vivo, are being investigated. Human Kynase has also shown a favorable PK profile and kynurenine degradation in non-human primates, and Kynase variants are now moving toward development candidate selection for treatment of cancers where both IDO/TDO pathways play a significant immunosuppressive role. Citation Format: Silvia Coma, Jill Cavanaugh, James Nolan, Jeremy Tchaicha, Karen McGovern, Everett Stone, Candice Lamb, Christos Karamitros, John Blazek, Kendra Garrison, George Georgiou, Mark Manfredi, Xiaoyan Michelle Zhang. Targeting the IDO/TDO pathway through degradation of the immunosuppressive metabolite kynurenine [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 3757.

Checkpoint inhibitors have become the cornerstone for new innovation in immune-based oncology therapy. Several orthogonal immune pathways are currently being investigated to relieve suppression or boost activity of the innate and adaptive immune system. The IDO immune-metabolism pathway was recently clinically validated in melanoma in combination with checkpoint inhibition. This opens up a new approach to relieving suppressive mediators and lends credence to the tumor microenvironment containing small molecule metabolites that induce immune tolerance. Both indoleamine-pyrrole 2,3-dioxygenase 1 (IDO1) and Tryptophan 2,3-dioxygenase (TDO) enzymes metabolize tryptophan, forming Kynurenine which binds the aryl hydrocarbon receptor (AHR) in multiple innate and adaptive immune cell types causing a net immunosuppressive effect. Both enzymes are upregulated across many tumor types, however, only the IDO1 enzyme has thus far been addressed in the clinic with small molecule inhibitors. We have postulated that enzyme-mediated depletion of Kynurenine into safe and immunologically inert metabolites can alleviate tumor immunosuppression. We have cloned and characterized several bacterial Kynureninases (KYNase) which preferentially degrade Kynurenine with a >1,000 higher kcat/KM as opposed to mammalian enzymes that cleave 3-OH Kynurenine. We show that PEGylated bacterial KYNases can deplete Kynurenine produced by IDO1+, TDO+ and IDO1/TDO+ dual positive human cancer cells whereas, the IDO1 inhibitor epacadostat or TDO inhibitor 680C91 only selectively inhibited Kyn production in IDO1+ or TDO+ cells respectively. In vivo, a single subcutaneous dose of KYNase in B16F10 tumor-bearing mice was able to deplete Kynurenine in both the plasma and tumors and increase effector T-cells in the tumor. KYNase demonstrated significant tumor growth inhibition and survival benefit either as a single agent or in combination with checkpoint inhibitors (anti-PD1 or anti-CTLA4) in B16F10, CT26 and 4T1 models. Interestingly, KYNase combined with anti-PD1, showed greater efficacy than epacadostat / anti-PD1 combination in CT26 tumor bearing mice. A pharmacologically optimized human KYNase is currently moving toward clinical development for the treatment of cancers where both IDO/TDO pathways play a significant immunosuppressive role through kynurenine production. Citation Format: Michelle Zhang, Everett Stone, Todd A. Triplett, Kendra Triplett, Candice Lamb, Christos S. Karamitros, John Blazek, George Georgiou, Mark G. Manfredi. A novel approach to targeting the IDO/TDO pathway through degradation of the immunosuppressive metabolite kynurenine [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 5570. doi:10.1158/1538-7445.AM2017-5570

Tryptophan oxidation and the ensuing production of Kynurenine (Kyn) and downstream products have been established to play an important role in immunosuppression within the tumor microenvironment (TME). The rate limiting step in Tryptophan oxidation is catalyzed by the IFN-γ inducible IDO-1 enzyme which is upregulated in numerous cancers and also, in some instances via the ectopic expression of TDO, which is normally expressed primarily in the liver. Currently, IDO-1 small molecule inhibitors in combination with checkpoint inhibitor antibodies are being evaluated in >12 clinical trials. However, whether the immune-suppressive effects of Tryptophan catabolism results from its depletion in the TME, or from the accumulation of Kynurenine, is not known. To distinguish between the effects of Trp depletion and Kyn accumulation in the TME we used an engineered Kynureninase (KYNase) enzyme that selectively degrades Kyn into immunologically inert and non-toxic metabolites. Peritumoral injection of PEGylated KYNase completely depletes serum and tumor Kyn levels for up to 72 hrs, while leaving Trp concentrations in both compartments unaffected. In the IDO-1 expressing CT26 colon carcinoma model KYNase treatment as monotherapy resulted in 16% complete and durable responses, accompanied by long-term immunity to tumor re-challenge. KYNase had no effect on tumor growth in IDO-/- mice, nor in RAG-/- or in CD8+ T cell depleted mice. Analysis of the TME demonstrated KYNase treatment resulted in increased accumulation of CD8+ cells, with a greater proportion expressing Granzyme B, undergoing proliferation and permeating the tumor interior. This effect was specific to the TME, as no detectable changes to the immune compartment occurred in other organs examined, nor overt signs of autoimmunity and toxicity. The increase in CD8+ cells was consistent with in vitro data showing that elevated concentration of Kyn directly induces apoptosis of activated CD8+ T cells partially through IL-2 suppression. No changes to the Treg percentages and phenotypes in the TME were observed, indicating that the Treg compartment is not impacted by treatment with KYNase. Cytokine analysis of tumor digests revealed increased IL-2 concentrations, as well as IFNγ and IL-9, in tumors after KYNase treatment. Furthermore, ex vivo stimulation of TIL after KYNase treatment demonstrated that the increase in cytokines in the TME is not solely due to an increase in TIL, but also an increased effector cytokine capacity on a per cell basis. Administration of KYNase together with either αCTLA4 or αPD1 synergistically elicited complete and durable regression of multiple established tumor models. In summary, our data support the hypothesis that Kyn accumulation in the TME rather than Trp depletion is the dominant IDO-mediated immune suppressive mechanism, and that enzyme-mediated Kyn depletion is a promising cancer immunotherapeutic approach. Citation Format: Todd A. Triplett, Kendra Triplett, Everett Stone, Michelle Zhang, Mark Manfredi, Candice Lamb, Yuri Tanno, Lauren Ehrlich, George Georgiou. Immune-checkpoint inhibition via enzyme-mediated degradation of kynurenine [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 5571. doi:10.1158/1538-7445.AM2017-5571

Taurine, the most abundant free amino acid in mammals, with many critical roles such as neuronal development, had so far only been reported to be synthetized in eukaryotes. Taurine is the major product of cysteine metabolism in mammals, and its biosynthetic pathway consists of cysteine dioxygenase and cysteine sulfinic acid decarboxylase (hCSAD). Sequence, structural, and mutational analyses of the structurally and sequentially related hCSAD and human glutamic acid decarboxylase (hGAD) enzymes revealed a three residue substrate recognition motif (X1aa19X2aaX3), within the active site that is responsible for coordinating their respective preferred amino acid substrates. Introduction of the cysteine sulfinic acid (CSA) motif into hGAD (hGAD-S192F/N212S/F214Y) resulted in an enzyme with a >700 fold switch in selectivity towards the decarboxylation of CSA over its preferred substrate, L-glutamic acid. Surprisingly, we found this CSA recognition motif in the genome sequences of several marine bacteria, prompting us to evaluate the catalytic properties of bacterial amino acid decarboxylases that were predicted by sequence motif to decarboxylate CSA but had been annotated as GAD enzymes. We show that CSAD from Synechococcus sp. PCC 7335 specifically decarboxylated CSA and that the bacteria accumulated intracellular taurine. The fact that CSAD homologues exist in certain bacteria and are frequently found in operons containing the recently discovered bacterial cysteine dioxygenases that oxidize L-cysteine to CSA, supports the idea that a bona fide bacterial taurine biosynthetic pathway exists in prokaryotes.