A phase I, first-in-human study of TAK-164, an antibody-drug conjugate, in patients with advanced gastrointestinal cancers expressing guanylyl cyclase C.

PURPOSE: Guanylyl cyclase C (GCC) is highly expressed in several gastrointestinal malignancies and preclinical studies suggest that it is a promising target for antibody-based therapeutics. This phase I trial assessed the safety and tolerability of TAK-164, an investigational, anti-GCC antibody-drug conjugate (NCT03449030). METHODS: Thirty-one patients with GCC-positive, advanced gastrointestinal cancers received intravenous TAK-164 on day 1 of 21-day cycles. Dose escalation proceeded based on cycle 1 safety data via a Bayesian model. RESULTS: Median age was 58 years (range 32-72), 25 patients (80.6%) had colorectal carcinoma, and median number of prior therapies was four. No dose-limiting toxicities (DLTs) were reported during cycle 1 DLT evaluation period. After cycle 2 dosing, 3 patients reported dose-limiting treatment-emergent adverse events (TEAEs): grade 3 pyrexia and grade 5 hepatic failure (0.19 mg/kg), grade 4 hepatic failure and platelet count decreased (0.25 mg/kg), grade 3 nausea, grade 4 platelet and neutrophil count decreased (0.25 mg/kg). The recommended phase II dose (RP2D) was 0.064 mg/kg. Common TAK-164-related TEAEs included platelet count decreased (58.1%), fatigue (38.7%), and anemia (32.3%). There was a dose-dependent increase in TAK-164 exposure over the range, 0.032-0.25 mg/kg. TAK-164 half-life ranged from 63.5 to 159 h. One patient (0.008 mg/kg) with high baseline GCC expression had an unconfirmed partial response. CONCLUSIONS: TAK-164 appeared to have a manageable safety profile at 0.064 mg/kg. Hepatic toxicity was identified as a potential risk. The RP2D of 0.064 mg/kg was considered insufficient to derive clinical benefit; there are no plans for further clinical development. CLINICAL TRIAL REGISTRATION: NCT03449030.

Receptor Guanylyl Cyclase C and Cyclic GMP in Health and Disease: Perspectives and Therapeutic Opportunities.

Receptor Guanylyl Cyclase C (GC-C) was initially characterized as an important regulator of intestinal fluid and ion homeostasis. Recent findings demonstrate that GC-C is also causally linked to intestinal inflammation, dysbiosis, and tumorigenesis. These advances have been fueled in part by identifying mutations or changes in gene expression in GC-C or its ligands, that disrupt the delicate balance of intracellular cGMP levels and are associated with a wide range of clinical phenotypes. In this review, we highlight aspects of the current knowledge of the GC-C signaling pathway in homeostasis and disease, emphasizing recent advances in the field. The review summarizes extra gastrointestinal functions for GC-C signaling, such as appetite control, energy expenditure, visceral nociception, and behavioral processes. Recent research has expanded the homeostatic role of GC-C and implicated it in regulating the ion-microbiome-immune axis, which acts as a mechanistic driver in inflammatory bowel disease. The development of transgenic and knockout mouse models allowed for in-depth studies of GC-C and its relationship to whole-animal physiology. A deeper understanding of the various aspects of GC-C biology and their relationships with pathologies such as inflammatory bowel disease, colorectal cancer, and obesity can be leveraged to devise novel therapeutics.

A ß-Catenin-TCF-Sensitive Locus Control Region Mediates GUCY2C Ligand Loss in Colorectal Cancer.

BACKGROUND & AIMS: Sporadic colorectal cancers arise from initiating mutations in APC, producing oncogenic ß-catenin/TCF-dependent transcriptional reprogramming. Similarly, the tumor suppressor axis regulated by the intestinal epithelial receptor GUCY2C is among the earliest pathways silenced in tumorigenesis. Retention of the receptor, but loss of its paracrine ligands, guanylin and uroguanylin, is an evolutionarily conserved feature of colorectal tumors, arising in the earliest dysplastic lesions. Here, we examined a mechanism of GUCY2C ligand transcriptional silencing by ß-catenin/TCF signaling. METHODS: We performed RNA sequencing analysis of 4 unique conditional human colon cancer cell models of ß-catenin/TCF signaling to map the core Wnt-transcriptional program. We then performed a comparative analysis of orthogonal approaches, including luciferase reporters, chromatin immunoprecipitation sequencing, CRISPR/Cas9 (clustered regularly interspaced short palindromic repeats) knockout, and CRISPR epigenome editing, which were cross-validated with human tissue chromatin immunoprecipitation sequencing datasets, to identify functional gene enhancers mediating GUCY2C ligand loss. RESULTS: RNA sequencing analyses reveal the GUCY2C hormones as 2 of the most sensitive targets of ß-catenin/TCF signaling, reflecting transcriptional repression. The GUCY2C hormones share an insulated genomic locus containing a novel locus control region upstream of the guanylin promoter that mediates the coordinated silencing of both genes. Targeting this region with CRISPR epigenome editing reconstituted GUCY2C ligand expression, overcoming gene inactivation by mutant ß-catenin/TCF signaling. CONCLUSIONS: These studies reveal DNA elements regulating corepression of GUCY2C ligand transcription by ß-catenin/TCF signaling, reflecting a novel pathophysiological step in tumorigenesis. They offer unique genomic strategies that could reestablish hormone expression in the context of canonical oncogenic mutations to reconstitute the GUCY2C axis and oppose transformation.

Diarrheal pathogens trigger rapid evolution of the guanylate cyclase-C signaling axis in bats.

The pathogenesis of infectious diarrheal diseases is largely attributed to enterotoxins that cause dehydration by disrupting intestinal water absorption. We investigated patterns of genetic variation in mammalian guanylate cyclase-C (GC-C), an intestinal receptor targeted by bacterially encoded heat-stable enterotoxins (STa), to determine how host species adapt in response to diarrheal infections. Our phylogenetic and functional analysis of GC-C supports long-standing evolutionary conflict with diarrheal bacteria in primates and bats, with highly variable susceptibility to STa across species. In bats, we further show that GC-C diversification has sparked compensatory mutations in the endogenous uroguanylin ligand, suggesting an unusual scenario of pathogen-driven evolution of an entire signaling axis. Together, these findings suggest that conflicts with diarrheal pathogens have had far-reaching impacts on the evolution of mammalian gut physiology.

Guanylyl cyclase 2C (GUCY2C) in gastrointestinal cancers: recent innovations and therapeutic potential.

INTRODUCTION: Gastrointestinal (GI) cancers account for the second leading cause of cancer-related deaths in the United States. Guanylyl cyclase C (GUCY2C) is an intestinal signaling system that regulates intestinal fluid and electrolyte secretion as well as intestinal homeostasis. In recent years, it has emerged as a promising target for chemoprevention and therapy for GI malignancies. AREAS COVERED: The loss of GUCY2C signaling early in colorectal tumorigenesis suggests it could have a significant impact on tumor initiation. Recent studies highlight the importance of GUCY2C signaling in preventing colorectal tumorigenesis using agents such as linaclotide, plecanatide, and sildenafil. Furthermore, GUCY2C is a novel target for immunotherapy and a diagnostic marker for primary and metastatic diseases. EXPERT OPINION: There is an unmet need for prevention and therapy in GI cancers. In that context, GUCY2C is a promising target for prevention, although the precise mechanisms by which GUCY2C signaling affects tumorigenesis remain to be defined. Furthermore, clinical trials are exploring its role as an immunotherapeutic target for vaccines to prevent metastatic disease. Indeed, GUCY2C is an emerging target across the disease continuum from chemoprevention, to diagnostic management, through the treatment and prevention of metastatic diseases.

Peripheral Guanylate Cyclase-C modulation of corticolimbic activation and corticotropin-releasing factor signaling in a rat model of stress-induced colonic hypersensitivity.

BACKGROUND: Psychological stress is a risk factor for irritable bowel syndrome, a functional gastrointestinal pain disorder featuring abnormal brain-gut connectivity. The guanylate cyclase-C (GC-C) agonist linaclotide has been shown to relieve abdominal pain in IBS-C and exhibits antinociceptive effects in rodent models of post-inflammatory visceral hypersensitivity. However, the role GC-C signaling plays in psychological stress-induced visceral hypersensitivity is unknown. Here, we test the hypothesis that GC-C agonism reverses stress-induced colonic hypersensitivity via inhibition of nociceptive afferent signaling resulting in normalization of stress-altered corticotropin-releasing factor (CRF) expression in brain regions involved in pain perception and modulation. METHODS: Adult female rats were exposed to water avoidance stress or sham stress for 10 days, and the effects of linaclotide on stress-induced changes in colonic sensitivity, corticolimbic phospho-extracellular signal-regulated kinase (pERK), and CRF expression were measured using a combination of behavioral assessments, immunohistochemistry, and qRT-PCR. KEY RESULTS: Stressed rats exhibited colonic hypersensitivity and elevated corticolimbic pERK on day 11, which was inhibited by linaclotide. qRT-PCR analysis revealed dysregulated CRF expression in the medial prefrontal cortex, paraventricular nucleus of the hypothalamus, and central nucleus of the amygdala on day 28. Dysregulated CRF expression was not affected by linaclotide treatment. CONCLUSIONS AND INFERENCES: Our results demonstrate that exposure to repeated stress induces chronic colonic hypersensitivity in conjunction with altered corticolimbic activation and CRF expression. GC-C agonism attenuated stress-induced colonic hypersensitivity and ERK phosphorylation, but had no effect on CRF expression, suggesting the analgesic effects of linaclotide occur independent of stress-driven CRF gene expression in corticolimbic circuitry.

An update on guanylyl cyclase C in the diagnosis, chemoprevention, and treatment of colorectal cancer.

Introduction: Colorectal cancer remains the second leading cause of cancer death in the United States, underscoring the need for novel therapies. Despite the successes of new targeted agents for other cancers, colorectal cancer suffers from a relative scarcity of actionable biomarkers. In this context, the intestinal receptor, guanylyl cyclase C (GUCY2C), has emerged as a promising target.Areas covered: GUCY2C regulates a tumor-suppressive signaling axis that is silenced through loss of its endogenous ligands at the earliest stages of tumorigenesis. A body of literature supports a cancer chemoprevention strategy involving reactivation of GUCY2C through FDA-approved cGMP-elevating agents such as linaclotide, plecanatide, and sildenafil. Its limited expression in extra-intestinal tissues, and retention on the surface of cancer cells, also positions GUCY2C as a target for immunotherapies to treat metastatic disease, including vaccines, chimeric antigen receptor T-cells, and antibody-drug conjugates. Likewise, GUCY2C mRNA identifies metastatic cells, enhancing colorectal cancer detection, and staging. Pre-clinical and clinical programs exploring these GUCY2C-targeting strategies will be reviewed.Expert opinion: Recent mechanistic insights characterizing GUCY2C ligand loss early in tumorigenesis, coupled with results from the first clinical trials testing GUCY2C-targeting strategies, continue to elevate GUCY2C as an ideal target for prevention, detection, and therapy.

Pharmacokinetics and Catabolism of [3H]TAK-164, a Guanylyl Cyclase C Targeted Antibody-Drug Conjugate.

TAK-164 is an antibody-drug conjugate (ADC) comprising human anti-guanylyl cyclase C (GCC) monoclonal antibody conjugated to indolinobenzodiazepine DNA alkylator IGN-P1 through a cleavable alanine-alanine dipeptide linker. TAK-164 is currently being evaluated for the treatment of gastrointestinal cancers expressing GCC. The catabolism of TAK-164 was studied using 3H-labeled ADC using GCC-expressing HEK-293 (GCC-HEK-293) cells, rat tritosomes, cathepsin B, and tumor-bearing mice. Time- and target-dependent uptake of [3H]TAK-164 was observed in GCC-HEK-293 cells with approximately 12% of radioactivity associated with DNA after 24 hours of incubation. Rat liver tritosomes and cathepsin B yielded IGN-P1 aniline, sulfonated IGN-P1 (s-IGN-P1) aniline, and a lysine conjugate of IGN-P1 (IGN-P1-Lys) aniline as catabolites. In tumor-bearing mice, [3H]TAK-164 exhibited a terminal half-life of approximately 41 and 51 hours in plasma and blood, respectively, with low plasma clearance (0.75 ml/h per kilogram). The extractable radioactivity in plasma and tumor samples revealed the presence of s-IGN-P1 aniline and IGN-P1 aniline as payload-related components. The use of a radiolabeled payload in the ADC in tumor uptake investigations provided direct and quantitative evidence for tumor uptake, DNA binding, and proof of mechanism of action of the payload. SIGNIFICANCE STATEMENT: Since payload-related species are potent cytotoxins, a thorough characterization of released products of ADCs, metabolites, and their drug interaction potential is necessary prior to clinical investigations. This study characterized in vitro and in vivo DNA binding mechanisms and released products of TAK-164. The methodologies described here will be highly useful for characterization of payload-related products of ADCs in general.

Chimeric Ad5.F35 vector evades anti-adenovirus serotype 5 neutralization opposing GUCY2C-targeted antitumor immunity.

BACKGROUND: Adenovirus serotype 5 (Ad5) is a commonly used viral vector for transient delivery of transgenes, primarily for vaccination against pathogen and tumor antigens. However, endemic infections with Ad5 produce virus-specific neutralizing antibodies (NAbs) that limit transgene delivery and constrain target-directed immunity following exposure to Ad5-based vaccines. Indeed, clinical trials have revealed the limitations that virus-specific NAbs impose on the efficacy of Ad5-based vaccines. In that context, the emerging focus on immunological approaches targeting cancer self-antigens or neoepitopes underscores the unmet therapeutic need for more efficacious vaccine vectors. METHODS: Here, we evaluated the ability of a chimeric adenoviral vector (Ad5.F35) derived from the capsid of Ad5 and fiber of the rare adenovirus serotype 35 (Ad35) to induce immune responses to the tumor-associated antigen guanylyl cyclase C (GUCY2C). RESULTS: In the absence of pre-existing immunity to Ad5, GUCY2C-specific T-cell responses and antitumor efficacy induced by Ad5.F35 were comparable to Ad5 in a mouse model of metastatic colorectal cancer. Furthermore, like Ad5, Ad5.F35 vector expressing GUCY2C was safe and produced no toxicity in tissues with, or without, GUCY2C expression. Importantly, this chimeric vector resisted neutralization in Ad5-immunized mice and by sera collected from patients with colorectal cancer naturally exposed to Ad5. CONCLUSIONS: These data suggest that Ad5.F35-based vaccines targeting GUCY2C, or other tumor or pathogen antigens, may produce clinically relevant immune responses in more (≥90%) patients compared with Ad5-based vaccines (~50%).

APC-ß-catenin-TCF signaling silences the intestinal guanylin-GUCY2C tumor suppressor axis.

Sporadic colorectal cancer initiates with mutations in APC or its degradation target ß-catenin, producing TCF-dependent nuclear transcription driving tumorigenesis. The intestinal epithelial receptor, GUCY2C, with its canonical paracrine hormone guanylin, regulates homeostatic signaling along the crypt-surface axis opposing tumorigenesis. Here, we reveal that expression of the guanylin hormone, but not the GUCY2C receptor, is lost at the earliest stages of transformation in APC-dependent tumors in humans and mice. Hormone loss, which silences GUCY2C signaling, reflects transcriptional repression mediated by mutant APC-ß-catenin-TCF programs in the nucleus. These studies support a pathophysiological model of intestinal tumorigenesis in which mutant APC-ß-catenin-TCF transcriptional regulation eliminates guanylin expression at tumor initiation, silencing GUCY2C signaling which, in turn, dysregulates intestinal homeostatic mechanisms contributing to tumor progression. They expand the mechanistic paradigm for colorectal cancer from a disease of irreversible mutations in APC and ß-catenin to one of guanylin hormone loss whose replacement, and reconstitution of GUCY2C signaling, could prevent tumorigenesis.

Inflammation in cancer and depression: a starring role for the kynurenine pathway.

Depression is a common comorbidity in cancer cases, but this is not only due to the emotional distress of having a life-threatening disease. A common biological mechanism, involving a dysregulated immune system, seems to underpin this comorbidity. In particular, the activation of the kynurenine pathway of tryptophan degradation due to inflammation may play a key role in the development and persistence of both diseases. As a consequence, targeting enzymes involved in this pathway offers a unique opportunity to develop new strategies to treat cancer and depression at once. In this work, we provide a systematic review of the evidence up to date on the kynurenine pathway role in linking depression and cancer and on clinical implications of this evidence. In particular, complications due to chemotherapy are discussed, as well as the potential antidepressant efficacy of novel immunotherapies for cancer.

Synthesis of phenylpyrimidinones as guanylyl cyclase C inhibitors.

Diarrhea is one of the most important causes of mortality in the developing world, being responsible for 2.5 million deaths each year. Many of these deaths are caused by enterotoxigenic strains of bacteria, like Escherichia coli, that produce enterotoxins that cause acute watery diarrhea, commonly defined as secretory diarrhea. Studies on symptomatic patients indicate a high prevalence of enterotoxigenic E. coli strains producing the heat-stable toxin, STa. STa is a small, cysteine-rich peptide that binds to the extracellular receptor domain of guanylyl cyclase C (GCC), located at the luminal membrane of intestinal epithelial cells. GCC and its endogenous peptide ligands, guanylin and uroguanylin, play a key role in balancing water absorption and hydration of the intestinal lumen, as exemplified by the finding that loss of GCC function causes severe dehydration of the intestinal lumen, culminating in intestinal obstruction. From a mechanistic viewpoint, reduction of GCC activity offers an efficient approach to limit enterotoxigenic E. coli- provoked secretory diarrhea. Inhibition of GCC-mediated cGMP production would not only reduce anion secretion, but would also restore NHE3 activity, resulting in a comprehensive antidiarrheal action. In the present study, two novel phenylpyrimidinone derivatives were simultaneously synthesized and tested for their ability to block STa-induced CFTR activity in T84 cells.

Targeting the paracrine hormone-dependent guanylate cyclase/cGMP/phosphodiesterases signaling pathway for colorectal cancer prevention.

Colorectal cancer (CRC) is one of the leading causes of cancer related-deaths. The risk of development of CRC is complex and multifactorial, and includes disruption of homeostasis of the intestinal epithelial layer mediated though dysregulations of tumor suppressing/promoting signaling pathways. Guanylate cyclase 2C (GUCY2C), a membrane-bound guanylate cyclase receptor, is present in the apical membranes of intestinal epithelial cells and maintains homeostasis. GUCY2C is activated upon binding of paracrine hormones (guanylin and uroguanylin) that lead to formation of cyclic GMP from GTP and activation of downstream signaling pathways that are associated with normal homeostasis. Dysregulation/suppression of the GUCY2C-mediated signaling promotes CRC tumorigenesis. High-calorie diet-induced obesity is associated with deficiency of guanylin expression and silencing of GUCY2C-signaling in colon epithelial cells, leading to tumorigenesis. Thus, GUCY2C agonists, such as linaclotide, exhibit considerable role in preventing CRC tumorigenesis. However, phosphodiesterases (PDEs) are elevated in intestinal epithelial cells during CRC tumorigenesis and block GUCY2C-mediated signaling by degrading cyclic GMP to 5`-GMP. PDE5-specific inhibitors, such as sildenafil, show considerable anti-tumorigenic potential against CRC by amplifying the GUCY2C/cGMP signaling pathway, but cannot achieve complete anti-tumorigenic effects. Hence, dual targeting the elevation of cGMP by providing paracrine hormone stimuli to GUCY2C and by inhibition of PDEs may be a better strategy for CRC prevention than alone. This review delineates the involvement of the GUCY2C/cGMP/PDEs signaling pathway in the homeostasis of intestinal epithelial cells. Further, the events are associated with dysregulation of this pathway during CRC tumorigenesis are also discussed. In addition, current updates on targeting the GUCY2C/cGMP/PDEs pathway with GUCY2C agonists and PDEs inhibitors for CRC prevention and treatment are described in detail.

Optimizing the radiosynthesis of [68Ga]DOTA-MLN6907 peptide containing three disulfide cyclization bonds - a GCC specific chelate for clinical radiopharmaceuticals.

In the present study, the effect of radiolabeling conditions on radiolabeling efficiency and achievable specific activity of a DOTA-conjugated highly-lipophilic peptide containing three disulfide cyclization bonds was examined. The peptide is designed to bind specifically (with high affinity) to cell-surface receptor guanylyl cyclase C (GCC), which is universally expressed by colorectal cancer cells. The effect of systematic variation of chemical parameters pH, mass of peptide, acetate buffer concentration (ionic strength), and inclusion of ethanol in the radiolabeling reaction vessel on achievable specific activity and labeling efficiency was examined. In addition, a unique approach to acetone-based elution of 68Ga from an initial cation-exchange pre-concentration column is introduced, which improved radiochemical yield and radiochemical purity. For the evaluation of the acetone-based method, two different post-radiolabeling reverse-phase (C18) approaches to purify the final radiolabeled peptide were tested. These results revealed the potential for peptide degradation via the cleavage of disulfide cyclization bonds to form free thiols when using one of these C18 cartridges. The final optimized procedure enabled radiolabeling efficiency of greater than 99% and specific activity greater than 35 MBq/nmole in less than 30 min. The optimized parameters were amenable to the use of an automated 68Ge/68Ga generator and fluid-handling system for clinical production of the GCC receptor-specific [68Ga]DOTA-MLN6907 peptide. The chemical characteristics of individual peptides govern the most appropriate radiolabeling conditions for the preparation of radiopharmaceuticals.

Oral treatment with plecanatide or dolcanatide attenuates visceral hypersensitivity via activation of guanylate cyclase-C in rat models.

AIM: To investigate the effects of plecanatide and dolcanatide on maintenance of paracellular permeability, integrity of tight junctions and on suppression of visceral hypersensitivity. METHODS: Transport of fluorescein isothiocyanate (FITC)-dextran was measured to assess permeability across cell monolayers and rat colon tissues. Effects of plecanatide and dolcanatide on the integrity of tight junctions in Caco-2 and T84 monolayers and on the expression and localization of occludin and zonula occludens-1 (ZO-1) were examined by immunofluorescence microscopy. Anti-nociceptive activity of these agonists was evaluated in trinitrobenzene sulfonic acid (TNBS)-induced inflammatory as well as in non-inflammatory partial restraint stress (PRS) rat models. Statistical significance between the treatment groups in the permeability studies were evaluated using unpaired t-tests. RESULTS: Treatment of T84 and Caco-2 monolayers with lipopolysaccharide (LPS) rapidly increased permeability, which was effectively suppressed when monolayers were also treated with plecanatide or dolcanatide. Similarly, when T84 and Caco-2 monolayers were treated with LPS, cell surface localization of tight junction proteins occludin and ZO-1 was severely disrupted. When cell monolayers were treated with LPS in the presence of plecanatide or dolcanatide, occludin and ZO-1 were localized at the cell surface of adjoining cells, similar to that observed for vehicle treated cells. Treatment of cell monolayers with plecanatide or dolcanatide without LPS did not alter permeability, integrity of tight junctions and cell surface localization of either of the tight junction proteins. In rat visceral hypersensitivity models, both agonists suppressed the TNBS-induced increase in abdominal contractions in response to colorectal distension without affecting the colonic wall elasticity, and both agonists also reduced colonic hypersensitivity in the PRS model. CONCLUSION: Our results suggest that activation of GC-C signaling might be involved in maintenance of barrier function, possibly through regulating normal localization of tight junction proteins. Consistent with these findings, plecanatide and dolcanatide showed potent anti-nociceptive activity in rat visceral hypersensitivity models. These results imply that activation of GC-C signaling may be an attractive therapeutic approach to treat functional constipation disorders and inflammatory gastrointestinal conditions.

A monomethyl auristatin E-conjugated antibody to guanylyl cyclase C is cytotoxic to target-expressing cells in vitro and in vivo.

Guanylyl cyclase C (GCC) is a cell-surface protein that is expressed by normal intestinal epithelial cells, more than 95% of metastatic colorectal cancers (mCRC), and the majority of gastric and pancreatic cancers. Due to strict apical localization, systemically delivered GCC-targeting agents should not reach GCC in normal intestinal tissue, while accessing antigen in tumor. We generated an investigational antibody-drug conjugate (TAK-264, formerly MLN0264) comprising a fully human anti-GCC monoclonal antibody conjugated to monomethyl auristatin E via a protease-cleavable peptide linker. TAK-264 specifically bound, was internalized by, and killed GCC-expressing cells in vitro in an antigen-density-dependent manner. In GCC-expressing xenograft models with similar GCC expression levels/patterns observed in human mCRC samples, TAK-264 induced cell death, leading to tumor regressions and long-term tumor growth inhibition. TAK-264 antitumor activity was generally antigen-density-dependent, although some GCC-expressing tumors were refractory to TAK-264-targeted high local concentrations of payload. These data support further evaluation of TAK-264 in the treatment of GCC-expressing tumors.

TAK-264 (MLN0264) in Previously Treated Asian Patients with Advanced Gastrointestinal Carcinoma Expressing Guanylyl Cyclase C: Results from an Open-Label, Non-randomized Phase 1 Study.

PURPOSE: This phase 1 dose-escalation portion of the study evaluated the safety, pharmacokinetics (PK), and antitumor activity of TAK-264 in Asian patients with advanced gastrointestinal (GI) carcinoma or metastatic or recurrent gastric or gastroesophageal junction adenocarcinoma expressing guanylyl cyclase C (GCC). MATERIALS AND METHODS: Adult patients with advanced GI malignancies expressing GCC (H-score ≥ 10) received TAK-264 on day 1 of 3-week cycles as 30-minute intravenous infusions for up to 1 year or until disease progression or unacceptable toxicity. The primary objectives were to evaluate the safety profile including dose-limiting toxicities (DLTs) during cycle 1, determine the maximum tolerated dose (MTD), and characterize the PK profile of TAK-264. RESULTS: Twelve patients were enrolled and treated with 1.2 mg/kg (n=3), 1.5 mg/kg (n=3), or 1.8 mg/kg TAK-264 (n=6). Median number of treatment cycles received was two (range, 1 to 10). None of the patients experienced a DLT and the MTD was not determined. Ten patients (83%) experienced adverse events (AEs). The most common were neutropenia, anorexia, and nausea (each reported by four patients). Five patients (42%) experienced grade ≥ 3 AEs consisting of tumor hemorrhage and hypertension, ascites, adrenal insufficiency, neutropenia and asthenia. Serum exposure to TAK-264 increased proportionally with the dose and the median half-life was approximately 5.5-6.6 days. No patients experienced an objective response. CONCLUSION: TAK-264 demonstrated a manageable safety profile with limited antitumor activity consistent with studies conducted in Western patients with advanced GI malignancies. TAK-264 exposure increased proportionally with the dose.

Consistent expression of guanylyl cyclase-C in primary and metastatic gastrointestinal cancers.

BACKGROUND: The transmembrane receptor guanylate cyclase-C (GCC) has been found to be expressed in colorectal cancers. However, limited data are available on GCC protein expression in non-colorectal gastrointestinal tumors and few studies have reported whether GCC protein expression was consistently preserved in synchronous primary and metastatic cancer tissues. METHODS: GCC protein status was assessed by immunohistochemistry in tumor specimens from individuals (n = 627) with gastrointestinal tumors, including esophageal (n = 130), gastric (n = 276), pancreatic (n = 136), and colorectal (n = 85) primary and metastatic tumors. Tissue specimens consisted of tissue microarrays containing esophageal, gastric, pancreatic tumors, and whole-slide tissue sections from colorectal cancer patients with matching primary and metastatic tumors. RESULT: Among the evaluated esophageal, gastric, and pancreatic tumors, the frequency of GCC positivity at the protein level ranged from 59% to 68%. GCC was consistently expressed in primary and matched/synchronous metastatic lesions of colorectal cancer tissues derived from the same patients. CONCLUSION: This observational study demonstrated the protein expression of GCC across various gastrointestinal malignancies. In all cancer histotypes, GCC protein localization was observed predominantly in the cytoplasm compared to the membrane region of tumor cells. Consistent immunohistochemistry detection of GCC protein expression in primary colorectal cancers and in their matched liver metastases suggests that the expression of GCC is maintained throughout the process of tumor progression and formation of metastatic disease.

ST-Producing E. coli Oppose Carcinogen-Induced Colorectal Tumorigenesis in Mice.

There is a geographic inequality in the incidence of colorectal cancer, lowest in developing countries, and greatest in developed countries. This disparity suggests an environmental contribution to cancer resistance in endemic populations. Enterotoxigenic bacteria associated with diarrheal disease are prevalent in developing countries, including enterotoxigenic E. coli (ETEC) producing heat-stable enterotoxins (STs). STs are peptides that are structurally homologous to paracrine hormones that regulate the intestinal guanylyl cyclase C (GUCY2C) receptor. Beyond secretion, GUCY2C is a tumor suppressor universally silenced by loss of expression of its paracrine hormone during carcinogenesis. Thus, the geographic imbalance in colorectal cancer, in part, may reflect chronic exposure to ST-producing organisms that restore GUCY2C signaling silenced by hormone loss during transformation. Here, mice colonized for 18 weeks with control E. coli or those engineered to secrete ST exhibited normal growth, with comparable weight gain and normal stool water content, without evidence of secretory diarrhea. Enterotoxin-producing, but not control, E. coli, generated ST that activated colonic GUCY2C signaling, cyclic guanosine monophosphate (cGMP) production, and cGMP-dependent protein phosphorylation in colonized mice. Moreover, mice colonized with ST-producing E. coli exhibited a 50% reduction in carcinogen-induced colorectal tumor burden. Thus, chronic colonization with ETEC producing ST could contribute to endemic cancer resistance in developing countries, reinforcing a novel paradigm of colorectal cancer chemoprevention with oral GUCY2C-targeted agents.

A phase II study of antibody-drug conjugate, TAK-264 (MLN0264) in previously treated patients with advanced or metastatic pancreatic adenocarcinoma expressing guanylyl cyclase C.

Background This phase II open-label, multicenter study evaluated the efficacy, safety, and tolerability of TAK-264 in previously treated patients with advanced or metastatic pancreatic adenocarcinoma expressing guanylyl cyclase C (GCC). Methods Patients with advanced or metastatic pancreatic adenocarcinoma expressing GCC (H-score ≥ 10) received TAK-264 1.8 mg/kg on day 1 of a 21-day cycle as a 30-min intravenous infusion for up to 1 year or until disease progression or unacceptable toxicity. The primary objective was overall response rate (ORR [complete response + partial response (PR)]). Secondary objectives included evaluations of the safety and pharmacokinetic profile of TAK-264 (NCT02202785). Results 43 patients were enrolled and treated with 1.8 mg/kg TAK-264: 11, 15, and 17 patients with low, intermediate, and high GCC expression, respectively. Median number of treatment cycles received was two (range 1-10). The ORR was 3%, including one patient with intermediate GCC expression who achieved a PR. All patients experienced ≥1 adverse events (AE). The majority of patients experienced grade 1/2 AEs affecting the gastrointestinal tract. Fifteen (35%) patients experienced ≥grade 3 drug-related AEs; five (12%) patients had a serious AE. The most common (≥10% of patients) all-grade drug-related AEs were nausea (33%), fatigue (28%), neutropenia (23%), decreased appetite (23%), vomiting (16%), asthenia (16%), and alopecia (14%). Conclusions TAK-264 demonstrated a manageable safety profile; however, the low efficacy of TAK-264 observed in this study did not support further clinical investigation.