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Background: Tofacitinib is an oral Janus kinase inhibitor recently approved to induce and maintain remission in ulcerative colitis (UC)., Aims: Considering the number of anti-TNF non-responders, this study aims to assess the effectiveness and safety of tofacitinib in a cohort of multi-failure patients with moderate-to-severe UC at 52 weeks., Methods: From January 2021 to March 2023, we performed a prospective multicenter study observing adult patients with moderate-to-severe UC starting tofacitinib after an anti-TNF failure for a 52-week-long period. Effectiveness and safety were assessed in terms of colectomy rate, clinical remission and response, endoscopic remission, steroid-free clinical remission, and rate of adverse events., Results: We included 58 patients with UC with an age of 42 ± 14.4 years, 59% males, 96.6% left-sided or pancolitis, who were failure to a single (65.5%) or more than one anti-TNF (34.5%). Only 6 (10.3%) patients underwent colectomy. Colectomy was clinically associated with the necessity and the number of extra cycles of tofacitinib 10 mg bid at W8 (p = 0.023) and W24 (p = 0.004), and with a higher partial Mayo score at W8 (p = 0.025). At W52, clinical remission, clinical response, and steroid-free clinical remission were 53.4%, 43.1%, and 48.3%, respectively. Of 22 performed colonoscopies at W52, 11 (50%) showed endoscopic remission. Adverse events occurred in 14 (24.1%) patients, but only 2 (3.4%) led to tofacitinib discontinuation., Conclusions: In a real-life setting of patients with anti-TNF refractory UC, tofacitinib has proved to be effective in preventing colectomy and inducing clinical and endoscopic remission at 52 weeks with a good safety profile., (© 2024. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Purpose: To characterize patients with APS and to propose a new approach for their follow-up. Query ID="Q1" Text="Please check the given names and familynames.", Methods: Monocentric observational retrospective study enrolling patients referred to the Outpatients clinic of the Units of Endocrinology, Diabetology, Gastroenterology, Rheumatology and Clinical Immunology of our Hospital for Autoimmune diseases., Results: Among 9852 patients, 1174 (11.9%) [869 (73.9%) female] were diagnosed with APS. In 254 subjects, the diagnosis was made at first clinical evaluation (Group 1), all the other patients were diagnosed with a mean latency of 11.3 ± 10.6 years (Group 2). Group 1 and 2 were comparable for age at diagnosis (35.7 ± 16.3 vs. 40.4 ± 16.6 yrs, p = .698), but different in male/female ratio (81/173 vs 226/696, p = .019). In Group 2, 50% of patients developed the syndrome within 8 years of follow-up. A significant difference was found after subdividing the first clinical manifestation into the different outpatient clinic to which they referred (8.7 ± 8.0 vs. 13.4 ± 11.6 vs. 19.8 ± 8.7 vs. 7.4 ± 8.1 for endocrine, diabetic, rheumatologic, and gastroenterological diseases, respectively, p < .001)., Conclusions: We described a large series of patients affected by APS according to splitters and lumpers. We propose a flowchart tailored for each specialist outpatient clinic taking care of the patients. Finally, we recommend regular reproductive system assessment due to the non-negligible risk of developing premature ovarian failure., (© 2023. The Author(s).)

A lifelong gluten-free diet (GFD) is the only treatment for celiac disease and other gluten-related disorders. Nevertheless, strict adherence to the GFD is often challenging due to concerns about social isolation, risk of gluten contaminations, high cost, poor quality and the taste of gluten-free products. Moreover, although the GFD is effective in achieving mucosal healing, it may lead to dietary imbalances due to nutrient deficiencies over a long period of time. To overcome these issues, several gluten-free wheat flours have been developed to create products that closely resemble their gluten-containing counterparts. Furthermore, given the critical importance of adhering to the GFD, it becomes essential to promote adherence and monitor possible voluntary or involuntary transgressions. Various methods, including clinical assessment, questionnaires, serology for celiac disease, duodenal biopsies and the detection of Gluten Immunogenic Peptides (GIPs) are employed for this purpose, but none are considered entirely satisfactory. Since adherence to the GFD poses challenges, alternative therapies should be implemented in the coming years to improve treatment efficacy and the quality of life of patients with celiac disease. The aim of this narrative review is to explore current knowledge of the GFD and investigate its future perspectives, focusing on technology advancements, follow-up strategies and insights into a rapidly changing future.

Background and Objectives: Our Phase-I parallel-cohort study suggested that managing severe traumatic brain injury (sTBI) in the absence of intracranial pressure (ICP) monitoring using an ad hoc Imaging and Clinical Examination (ICE) treatment protocol was associated with superior outcome vs nonprotocolized management but could not differentiate the influence of protocolization from that of the specific protocol. Phase II investigates whether adopting the Consensus REVised Imaging and Clinical Examination (CREVICE) protocol improved outcome directly or indirectly via protocolization., Methods: We performed a Phase-II sequential parallel-cohort study examining adoption of the CREVICE protocol from no protocol vs a previous protocol in patients with sTBI older than 13 years presenting ≤24 hours after injury. Primary outcome was prespecified 6-month recovery. The study was done mostly at public South American centers managing sTBI without ICP monitoring. Fourteen Phase-I nonprotocol centers and 5 Phase-I protocol centers adopted CREVICE. Data were analyzed using generalized estimating equation regression adjusting for demographic imbalances., Results: A total of 501 patients (86% male, mean age 35.4 years) enrolled; 81% had 6 months of follow-up. Adopting CREVICE from no protocol was associated with significantly superior results for overall 6-month extended Glasgow Outcome Score (GOSE) (protocol effect = 0.53 [0.11, 0.95], P = .013), mortality (36% vs 21%, HR = 0.59 [0.46, 0.76], P < .001), and orientation (Galveston Orientation and Amnesia Test discharge protocol effect = 10.9 [6.0, 15.8], P < .001, 6-month protocol effect = 11.4 [4.1, 18.6], P < .005). Adopting CREVICE from ICE was associated with significant benefits to GOSE (protocol effect = 0.51 [0.04, 0.98], P = .033), 6-month mortality (25% vs 18%, HR = 0.55 [0.39, 0.77], P < .001), and orientation (Galveston Orientation and Amnesia Test 6-month protocol effect = 9.2 [3.6, 14.7], P = .004). Comparing both groups using CREVICE, those who had used ICE previously had significantly better GOSE (protocol effect = 1.15 [0.09, 2.20], P = .033)., Conclusion: Centers managing adult sTBI without ICP monitoring should strongly consider protocolization through adopting/adapting the CREVICE protocol. Protocolization is indirectly supported at sTBI centers regardless of resource availability., (Copyright © Congress of Neurological Surgeons 2023. All rights reserved.)

Purpose: To characterize patients with APS type 4 among those affected by APS diagnosed and monitored at our local Reference Center for Autoimmune Polyglandular Syndromes., Methods: Monocentric observational retrospective study enrolling patients affected by APS diagnosed and monitored in a Reference Center. Clinical records were retrieved and analyzed., Results: 111 subjects (51 males) were affected by APS type 4, mean age at the onset was 23.1 ± 15.1 years. In 15 patients the diagnosis of APS was performed during the first clinical evaluation, in the other 96 after a latency of 11 years (range 1-46). The most frequent diseases were type I diabetes mellitus and celiac disease, equally distributed among sexes., Conclusions: The prevalence of APS type 4 is 9:100,000 people. Type I diabetes mellitus was the leading indicator of APS type 4 in 78% subjects and in 9% permitted the diagnosis occurring as second manifestation of the syndrome. Our data, showing that 50% of patients developed APS type 4 within the first ten years, don't suggest any particular follow-up time and, more importantly, don't specify any particular disease. It is important to emphasize that 5% of women developed premature ovarian failure., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Gatta, Anelli, Cimino, Di Lodovico, Piovani, Zammarchi, Gozzoli, Maltese, Cavadini, Agosti, Delbarba, Pirola, Girelli, Buoso, Bambini, Alfieri, Bremi, Facondo, Lupo, Bezzi, Fredi, Mazzola, Gandossi, Saullo, Marini, Licini, Pezzaioli, Pini, Franceschini, Ricci and Cappelli.)

Lipid nanoparticles have demonstrated utility in hepatic delivery of a range of therapeutic modalities and typically deliver their cargo via low-density lipoprotein receptor-mediated endocytosis. For patients lacking sufficient low-density lipoprotein receptor activity, such as those with homozygous familial hypercholesterolemia, an alternate strategy is needed. Here we show the use of structure-guided rational design in a series of mouse and non-human primate studies to optimize a GalNAc-Lipid nanoparticle that allows for low-density lipoprotein receptor independent delivery. In low-density lipoprotein receptor-deficient non-human primates administered a CRISPR base editing therapy targeting the ANGPTL3 gene, the introduction of an optimized GalNAc-based asialoglycoprotein receptor ligand to the nanoparticle surface increased liver editing from 5% to 61% with minimal editing in nontargeted tissues. Similar editing was noted in wild-type monkeys, with durable blood ANGPTL3 protein reduction up to 89% six months post dosing. These results suggest that GalNAc-Lipid nanoparticles may effectively deliver to both patients with intact low-density lipoprotein receptor activity as well as those afflicted by homozygous familial hypercholesterolemia., (© 2023. The Author(s).)

Background: Most patients with severe traumatic brain injury (sTBI) in low- or-middle-income countries and surprisingly many in high-income countries are managed without intracranial pressure (ICP) monitoring. The impact of the first published protocol (Imaging and Clinical Examination [ICE] protocol) is untested against nonprotocol management., Objective: To determine whether patients treated in intensive care units (ICUs) using the ICE protocol have lower mortality and better neurobehavioral functioning than those treated in ICUs using no protocol., Methods: This study involved nineteen mostly public South American hospitals. This is a prospective cohort study, enrolling patients older than 13 years with sTBI presenting within 24 h of injury (January 2014-July 2015) with 6-mo postinjury follow-up. Five hospitals treated all sTBI cases using the ICE protocol; 14 used no protocol. Primary outcome was prespecified composite of mortality, orientation, functional outcome, and neuropsychological measures., Results: A total of 414 patients (89% male, mean age 34.8 years) enrolled; 81% had 6 months of follow-up. All participants included in composite outcome analysis: average percentile (SD) = 46.8 (24.0) nonprotocol, 56.9 (24.5) protocol. Generalized estimating equation (GEE) used to account for center effects (confounder-adjusted difference [95% CI] = 12.2 [4.6, 19.8], P = .002). Kaplan-Meier 6-month mortality (95% CI) = 36% (30%, 43%) nonprotocol, 25% (19%, 31%) protocol (GEE and confounder-adjusted hazard ratio [95% CI] = .69 [.43, 1.10], P = .118). Six-month Extended Glasgow Outcome Scale for 332 participants: average Extended Glasgow Outcome Scale score (SD) = 3.6 (2.6) nonprotocol, 4.7 (2.8) protocol (GEE and confounder-adjusted and lost to follow-up-adjusted difference [95% CI] = 1.36 [.55, 2.17], P = .001)., Conclusion: ICUs managing patients with sTBI using the ICE protocol had better functional outcome than those not using a protocol. ICUs treating patients with sTBI without ICP monitoring should consider protocolization. The ICE protocol, tested here and previously, is 1 option., (Copyright © Congress of Neurological Surgeons 2022. All rights reserved.)

Background: VERVE-101 is an investigational in vivo CRISPR base-editing medicine designed to alter a single DNA base in the PCSK9 gene, permanently turn off hepatic protein production, and thereby durably lower low-density lipoprotein cholesterol. We test the efficacy, durability, tolerability, and potential for germline editing of VERVE-101 in studies of nonhuman primates and a murine F1 progeny study., Methods: Cynomolgus monkeys were given a single intravenous infusion of a vehicle control (n=10) or VERVE-101 at a dose of 0.75 mg/kg (n=4) or 1.5 mg/kg (n=22) with subsequent follow-up up to 476 days. Two studies assessed the potential for germline editing, including sequencing sperm samples from sexually mature male nonhuman primates treated with VERVE-101 and genotyping offspring from female mice treated with the murine surrogate of VERVE-101 (VERVE-101mu)., Results: Liver biopsies 14 days after dosing noted mean PCSK9 editing of 46% and 70% in monkeys treated with VERVE-101 at 0.75 and 1.5 mg/kg, respectively. This translated into mean reductions in blood PCSK9 (proprotein convertase subtilisin/kexin type 9) of 67% and 83% and reductions of low-density lipoprotein cholesterol of 49% and 69% at the 0.75 and 1.5 mg/kg doses, respectively, assessed as time-weighted average change from baseline between day 28 and up to 476 days after dosing. Liver safety monitoring noted a transient rise in alanine aminotransferase and aspartate aminotransferase concentrations after infusion that fully resolved by day 14 with no accompanying change in total bilirubin. In a subset of monkeys necropsied 1 year after dosing, no findings related to VERVE-101 were identified on macroscopic and histopathologic assessment of the liver and other organs. In the study to assess potential germline editing of male nonhuman primates, sperm samples collected after VERVE-101 dosing showed no evidence of PCSK9 editing. Among 436 offspring of female mice treated with a saturating dose of VERVE-101mu, the PCSK9 edit was transmitted in 0 of 436 animals., Conclusions: VERVE-101 was well tolerated in nonhuman primates and led to 83% lower blood PCSK9 protein and 69% lower low-density lipoprotein cholesterol with durable effects up to 476 days after dosing. These results have supported the initiation of a first-in-human clinical trial in patients with heterozygous familial hypercholesterolemia and atherosclerotic cardiovascular disease.

Gene-editing technologies, which include the CRISPR-Cas nucleases 1-3 and CRISPR base editors 4,5 , have the potential to permanently modify disease-causing genes in patients 6 . The demonstration of durable editing in target organs of nonhuman primates is a key step before in vivo administration of gene editors to patients in clinical trials. Here we demonstrate that CRISPR base editors that are delivered in vivo using lipid nanoparticles can efficiently and precisely modify disease-related genes in living cynomolgus monkeys (Macaca fascicularis). We observed a near-complete knockdown of PCSK9 in the liver after a single infusion of lipid nanoparticles, with concomitant reductions in blood levels of PCSK9 and low-density lipoprotein cholesterol of approximately 90% and about 60%, respectively; all of these changes remained stable for at least 8 months after a single-dose treatment. In addition to supporting a 'once-and-done' approach to the reduction of low-density lipoprotein cholesterol and the treatment of atherosclerotic cardiovascular disease (the leading cause of death worldwide 7 ), our results provide a proof-of-concept for how CRISPR base editors can be productively applied to make precise single-nucleotide changes in therapeutic target genes in the liver, and potentially in other organs.

Globally, intracranial pressure (ICP) monitoring use in severe traumatic brain injury (sTBI) is inconsistent and susceptible to resource limitations and clinical philosophies. For situations without monitoring, there is no published comprehensive management algorithm specific to identifying and treating suspected intracranial hypertension (SICH) outside of the one ad hoc Imaging and Clinical Examination (ICE) protocol in the Benchmark Evidence from South American Trials: Treatment of Intracranial Pressure (BEST:TRIP) trial. As part of an ongoing National Institutes of Health (NIH)-supported project, a consensus conference involving 43 experienced Latin American Intensivists and Neurosurgeons who routinely care for sTBI patients without ICP monitoring, refined, revised, and augmented the original BEST:TRIP algorithm. Based on BEST:TRIP trial data and pre-meeting polling, 11 issues were targeted for development. We used Delphi-based methodology to codify individual statements and the final algorithm, using a group agreement threshold of 80%. The resulting CREVICE (Consensus REVised ICE) algorithm defines SICH and addresses both general management and specific treatment. SICH treatment modalities are organized into tiers to guide treatment escalation and tapering. Treatment schedules were developed to facilitate targeted management of disease severity. A decision-support model, based on the group's combined practices, is provided to guide this process. This algorithm provides the first comprehensive management algorithm for treating sTBI patients when ICP monitoring is not available. It is intended to provide a framework to guide clinical care and direct future research toward sTBI management. Because of the dearth of relevant literature, it is explicitly consensus based, and is provided solely as a resource (a "consensus-based curbside consult") to assist in treating sTBI in general intensive care units in resource-limited environments.

Recent studies have identified somatic ESR1 mutations in patients with metastatic breast cancer and found some of them to promote estrogen-independent activation of the receptor. The degree to which all recurrent mutants can drive estrogen-independent activities and reduced sensitivity to ER antagonists like fulvestrant is not established. In this report, we characterize the spectrum of ESR1 mutations from more than 900 patients. ESR1 mutations were detected in 10%, with D538G being the most frequent (36%), followed by Y537S (14%). Several novel, activating mutations were also detected (e.g., L469V, V422del, and Y537D). Although many mutations lead to constitutive activity and reduced sensitivity to ER antagonists, only select mutants such as Y537S caused a magnitude of change associated with fulvestrant resistance in vivo Correspondingly, tumors driven by Y537S, but not D5358G, E380Q, or S463P, were less effectively inhibited by fulvestrant than more potent and bioavailable antagonists, including AZD9496. These data point to a need for antagonists with optimal pharmacokinetic properties to realize clinical efficacy against certain ESR1 mutants. Significance: A diversity of activating ESR1 mutations exist, only some of which confer resistance to existing ER antagonists that might be overcome by next-generation inhibitors such as AZD9496. Cancer Discov; 7(3); 277-87. ©2016 AACR. This article is highlighted in the In This Issue feature, p. 235 ., (©2016 American Association for Cancer Research.)

Although endocrine therapy is successfully used to treat patients with estrogen receptor (ER) positive breast cancer, a substantial proportion of this population will relapse. Several mechanisms of acquired resistance have been described including activation of the mTOR pathway, increased activity of CDK4 and activating mutations in ER. Using a patient derived xenograft model harboring a common activating ER ligand binding domain mutation (D538G), we evaluated several combinatorial strategies using the selective estrogen receptor degrader (SERD) fulvestrant in combination with chromatin modifying agents, and CDK4/6 and mTOR inhibitors. In this model, fulvestrant binds WT and MT ER, reduces ER protein levels, and downregulated ER target gene expression. Addition of JQ1 or vorinostat to fulvestrant resulted in tumor regression (41% and 22% regression, respectively) though no efficacy was seen when either agent was given alone. Interestingly, although the CDK4/6 inhibitor palbociclib and mTOR inhibitor everolimus were efficacious as monotherapies, long-term delayed tumor growth was only observed when co-administered with fulvestrant. This observation was consistent with a greater inhibition of compensatory signaling when palbociclib and everolimus were co-dosed with fulvestrant. The addition of fulvestrant to JQ1, vorinostat, everolimus and palbociclib also significantly reduced lung metastatic burden as compared to monotherapy. The combination potential of fulvestrant with palbociclib or everolimus were confirmed in an MCF7 CRISPR model harboring the Y537S ER activating mutation. Taken together, these data suggest that fulvestrant may have an important role in the treatment of ER positive breast cancer with acquired ER mutations.

Fulvestrant is an estrogen receptor (ER) antagonist administered to breast cancer patients by monthly intramuscular injection. Given its present limitations of dosing and route of administration, a more flexible orally available compound has been sought to pursue the potential benefits of this drug in patients with advanced metastatic disease. Here we report the identification and characterization of AZD9496, a nonsteroidal small-molecule inhibitor of ERα, which is a potent and selective antagonist and downregulator of ERα in vitro and in vivo in ER-positive models of breast cancer. Significant tumor growth inhibition was observed as low as 0.5 mg/kg dose in the estrogen-dependent MCF-7 xenograft model, where this effect was accompanied by a dose-dependent decrease in PR protein levels, demonstrating potent antagonist activity. Combining AZD9496 with PI3K pathway and CDK4/6 inhibitors led to further growth-inhibitory effects compared with monotherapy alone. Tumor regressions were also seen in a long-term estrogen-deprived breast model, where significant downregulation of ERα protein was observed. AZD9496 bound and downregulated clinically relevant ESR1 mutants in vitro and inhibited tumor growth in an ESR1-mutant patient-derived xenograft model that included a D538G mutation. Collectively, the pharmacologic evidence showed that AZD9496 is an oral, nonsteroidal, selective estrogen receptor antagonist and downregulator in ER(+) breast cells that could provide meaningful benefit to ER(+) breast cancer patients. AZD9496 is currently being evaluated in a phase I clinical trial. Cancer Res; 76(11); 3307-18. ©2016 AACR., (©2016 American Association for Cancer Research.)

The tumor microenvironment is emerging as a key regulator of cancer growth and progression, however the exact mechanisms of interaction with the tumor are poorly understood. Whilst the majority of genomic profiling efforts thus far have focused on the tumor, here we investigate RNA-Seq as a hypothesis-free tool to generate independent tumor and stromal biomarkers, and explore tumor-stroma interactions by exploiting the human-murine compartment specificity of patient-derived xenografts (PDX).Across a pan-cancer cohort of 79 PDX models, we determine that mouse stroma can be separated into distinct clusters, each corresponding to a specific stromal cell type. This implies heterogeneous recruitment of mouse stroma to the xenograft independent of tumor type. We then generate cross-species expression networks to recapitulate a known association between tumor epithelial cells and fibroblast activation, and propose a potentially novel relationship between two hypoxia-associated genes, human MIF and mouse Ddx6. Assessment of disease subtype also reveals MMP12 as a putative stromal marker of triple-negative breast cancer. Finally, we establish that our ability to dissect recruited stroma from trans-differentiated tumor cells is crucial to identifying stem-like poor-prognosis signatures in the tumor compartment.In conclusion, RNA-Seq is a powerful, cost-effective solution to global analysis of human tumor and mouse stroma simultaneously, providing new insights into mouse stromal heterogeneity and compartment-specific disease markers that are otherwise overlooked by alternative technologies. The study represents the first comprehensive analysis of its kind across multiple PDX models, and supports adoption of the approach in pre-clinical drug efficacy studies, and compartment-specific biomarker discovery.

Acquired resistance to PI3K/mTOR/Akt pathway inhibitors is often associated with compensatory feedback loops involving the activation of oncogenes. Here, we have generated everolimus resistance in ER+ breast cancer cells and in long-term estrogen deprived (LTED) models that mimic progression on anti-estrogens. This allowed us to uncover MYC as a driver of mTOR inhibitor resistance. We demonstrate that both everolimus resistance and acute treatment of everolimus can lead to the upregulation of MYC mRNA, protein expression and, consequently, the enrichment of MYC signatures as revealed by RNA sequencing data. Depletion of MYC resulted in resensitization to everolimus, confirming its functional importance in this setting. Furthermore, ChIP assays demonstrate that MYC upregulation in the everolimus resistant lines is mediated by increased association of the BRD4 transcription factor with the MYC gene. Finally, JQ1, a BRD4 inhibitor combined with everolimus exhibited increased tumor growth inhibition in 3D Matrigel models and an in vivo xenograft model. These data suggest that MYC plays an important role in mediating resistance to everolimus in ER+ and ER+/LTED models. Furthermore, given the regulation ofMYCby BRD4 in this setting, these data have implications for increased therapeutic potential of combining epigenetic agents with mTOR inhibitors to effectively downregulate otherwise difficult to target transcription factors such as MYC.

The hedgehog pathway has been implicated in the tumorigenesis, tumor progression, and metastasis of numerous human cancers. We generated the first fully human hedgehog antibody MEDI-5304 and characterized its antitumor activity and preclinical toxicology. MEDI-5304 bound sonic hedgehog (SHH) and Indian hedgehog (IHH) with low picomolar affinity and neutralized SHH and IHH activity in cellular mGLI1 reporter assays. The antibody inhibited transcription of hedgehog target genes and osteoblast differentiation of C3H10T1/2 cells. We evaluated the activity of MEDI-5304 in vivo in model systems that allowed us to evaluate two primary hypotheses of hedgehog function in human cancer, paracrine signaling between tumor and stromal cells and cancer stem cell (CSC) self-renewal. MEDI-5304 displayed robust pharmacodynamic effects in stromal cells that translated to antitumor efficacy as a single agent in an HT-29/MEF coimplantation model of paracrine hedgehog signaling. MEDI-5304 also improved responses to carboplatin in the HT-29/MEF model. The antibody, however, had no effect as a single agent or in combination with gemcitabine on the CSC frequency or growth of several primary pancreatic cancer explant models. These findings support the conclusion that hedgehog contributes to tumor biology via paracrine tumor-stromal signaling but not via CSC maintenance or propagation. Finally, the only safety study finding associated with MEDI-5304 was ondontodysplasia in rats. Thus, MEDI-5304 represents a potent dual hedgehog inhibitor suitable for continued development to evaluate efficacy and safety in human patients with tumors harboring elevated levels of SHH or IHH.

Strains within the genus Salinospora have been shown to produce complex natural products having antibiotic and antiproliferative activities. The biochemical basis for the cytotoxic effects of salinosporamide A has been linked to its ability to inhibit the proteasome. Synthetically accessible salinosporamide A (ML858) was used to determine its biochemical and biological activities and to compare its effects with those of bortezomib. ML858 and bortezomib show time- and concentration-dependent inhibition of the proteasome in vitro. However, unlike bortezomib, which is a reversible inhibitor, ML858 covalently binds to the proteasome, resulting in the irreversible inhibition of 20S proteasome activity. ML858 was equipotent to bortezomib in cell-based reporter stabilization assays, but due to intramolecular instability is less potent in long-term assays. ML858 failed to maintain levels of proteasome inhibition necessary to achieve efficacy in tumor models responsive to bortezomib. Our results show that ML858 and bortezomib exhibit different kinetic and pharmacologic profiles and suggest that additional characterization of ML858 is warranted before its therapeutic potential can be fully appreciated.