A phase IB trial of 24-hour intravenous PX-12, a thioredoxin-1 inhibitor, in patients with advanced gastrointestinal cancers.

We investigated the safety, pharmacokinetics, and pharmacodynamics of PX-12, a thioredoxin-1 (Trx-1) inhibitor, administered as a 24-hour infusion every 7 or 14 days in patients with gastrointestinal malignancies. PX-12 is the first Trx-1 inhibitor to undergo clinical development. The first Phase 1 study of PX-12 demonstrated promising clinical activity, but the 1 and 3 hour-infusion schedules investigated were associated with a strong and irritating odor due to exhalation of one of its metabolites, 2-butanethiol. In an effort to achieve tolerability and achieve a drug exposure level necessary for biological activity, the current study was undertaken. While the maximally tolerated dose was estimated to be 300 mg/m(2) /24 h once a week as the 2-butanethiol expirate was tolerable at that dose level, no evidence of clinical activity was observed. Pharmacokinetic studies of the parent compound PX-12 demonstrated rapid, irreversible binding to plasma components, resulting in low (ng/ml) peak plasma concentrations of non-bound PX-12 during infusion. DCE-MRI was performed pre-and post-infusion in three patients. There were no significant trends observed in changes in plasma Trx-1, vascular endothelial growth factor (VEGF), or beta fibroblast growth factor (FGF-2) pre- or post-treatment. However, there was a trend for a decrease in circulating Trx-1 during the first four PX-12 treatment cycles in patients that had a Trx-1 baseline level >18 ng/mL. Aggregate clinical trial results suggest that further clinical development of PX-12, as an intravenous infusion, is not feasible. However, the Trx-1 pathway remains a target of interest in patients with gastrointestinal malignancies.

Tgf-beta1, Tgf-beta2, Tgf-beta3 and Msx2 expression is elevated during frontonasal suture morphogenesis and during active postnatal facial growth.

OBJECTIVES: It is hypothesized that regulation of facial suture morphogenesis is similar to that of cranial sutures, with expression of similar regulatory molecules, governing suture formation and patency. The present study was designed to characterize the morphology of the frontonasal (FN) suture of the rat at different developmental stages and to investigate the presence and temporal-spatial expression of transforming growth factor-beta 1 (Tgf-beta1), Tgf-beta2, Tgf-beta3 and Msx2 mRNA within these structures. SETTING AND SAMPLE POPULATION: The Department of Biomedical Sciences at Texas A&M University System Health Science Center, Baylor College of Dentistry, Dallas, TX USA. Histological sections and RNA isolated from FN suture tissues of Sprague-Dawley rats, aged embryonic day 16 through postnatal day 20. METHOD: Sections were examined after immunohistochemical staining. Gene expression was determined by densitometric analysis of RT-PCR products run on agarose gels. RESULTS: FN sutures develop slightly later than cranial sutures and show increased complexity over time when compared to cranial sutures. FN sutures were closely associated with the nasal capsular cartilage, with intervening layers of perichondrium and periosteum. The pattern of expression of Tgf-betas within the FN suture tissues was similar to that seen in the cranial sutures. However, mRNA and protein of the Tgf-betas were differentially expressed over time compared to cranial sutures. In FN sutures, Tgf-beta mRNA levels were elevated both during the period of suture morphogenesis and during active bone growth from the suture in the early postnatal period. Msx2 mRNA expression was elevated in both the prenatal and postnatal periods, similar to Tgf-beta mRNA expression. CONCLUSION: Tgf-beta and Msx2 are present in facial sutures similar to cranial sutures, but are differentially expressed over time, perhaps reflecting different bone growth rates from these sutures.

Transforming growth factor-beta3 (Tgf-beta3) down-regulates Tgf-beta3 receptor type I (Tbetar-I) during rescue of cranial sutures from osseous obliteration.

Appropriate biochemical regulation of intramembranous bone growth from sutures is necessary to achieve correct craniofacial morphology. Failure to form sutures (agenesis) or to maintain sutures in their unossified state (craniosynostosis) can result in severe facial dysmorphology. Several factors such as Twist, Msx2, fibroblast growth factors (Fgfs), bone morphogenetic proteins (Bmps) and transforming growth factors-beta (Tgf-betas) regulate suture patency, likely by interacting with one another. Tgf-beta2 and Tgf-beta3 use the same cell surface receptors, yet have opposite effects on suture patency, cellular proliferation and apoptosis within the suture. One possible mechanism by which Tgf-beta3 rescues sutures from obliteration is by regulating the ability of suture cells to respond to Tgf-beta2. As Tgf-beta3 does not regulate protein levels of Tgf-beta2 in sutures, Tgf-beta3 could regulate tissue responsiveness to Tgf-beta2 by regulating Tgf-beta2 access to receptors. Tgf-beta3 is a more potent competitor than Tgf-beta2 for cell surface receptors, so it is proposed that Tgf-beta3 binds to and down-regulates Tgf-beta receptor type I (Tbetar-I) expression by suture cells. This down-regulation would limit the ability of cells to respond to all Tgf-betas, including Tgf-beta2. To test this hypothesis, an in vitro culture model was used in which fetal rat sutures either remain patent or are induced to fuse when cultured in the presence or absence of dura mater, respectively. Tgf-beta3 was added to cultured calvaria and changes in the number of receptor positive cells within the suture were established. Data were compared with that seen in control sutures and in normal sutures in vivo. It was found that the numbers of cells expressing Tbetar-I within the suture matrix increased over time in sutures remaining patent. Osteoblastic cells lining the bone fronts on either side of sutures were Tbetar-I positive during early morphogenesis, but these numbers declined as sutures fused, both in vivo and in vitro. Addition of Tgf-beta3 to calvaria in culture decreased the number of Tbetar-I expressing cells in both fusing and non-fusing sutures, with dramatic decreases in the numbers of osteoblasts expressing Tbetar-I.

Transforming growth factor-beta 2 and TGF-beta 3 regulate fetal rat cranial suture morphogenesis by regulating rates of cell proliferation and apoptosis.

Cranial vault sutures are the major intramembranous bone growth sites during rapid expansion of the neurocranium. To function as bone growth sites, sutures need to remain patent, while allowing rapid bone formation at the edges of the bone fronts. Premature osseous obliteration of sutures (craniosynostosis) by fusion of bone fronts across the suture site prevents further bone formation at this site, often leading to severe facial dysmorphology. Although several growth factor receptor and transcription factor mutations have been implicated in craniosynostosis, the underlying mechanisms leading to sutural obliteration remain unclear. Previous studies have shown that dura secreted soluble factors responsible for maintaining suture patency and that suture fusion observed in the absence of dura was preceded by elevated levels of DNA synthesis and collagen production in the suture region. The use of neutralizing antibodies in a fetal calvarial culture model further demonstrated that removal of transforming growth factor (TGF) -beta 3 activity induced premature sutural obliteration, whereas removal of TGF-beta 2 activity prevented sutural obliteration. Data presented here demonstrate that suture obliteration induced by removal of TGF-beta 3 activity was preceded by elevated levels of DNA synthesis, similar to that seen upon removal of the dura. Addition of exogenous TGF-beta 3 to calvaria cultured without dura both prevented suture obliteration and reduced DNA synthesis to levels comparable to those seen with intact dura. Addition of exogenous TGF-beta 2 to calvarial cultures induced sutural fusion accompanied by elevated levels of cell proliferation. However, sutures rescued from obliteration by removal of TGF-beta 2 activity did not have decreased levels of cell proliferation, but rather appeared to be due to inhibited differentiation. In all calvaria in which sutures remained patent in culture, numbers of apoptotic cells were high within the suture, whereas in sutures destined to fuse, numbers of apoptotic cells were low. Results indicate that one of the critical regulators of suture patency is cell number. Alterations in cell number can trigger premature differentiation of cells, resulting in sutural obliteration. Furthermore, a complex interplay between closely related molecules is required to maintain cranial vault sutures in an unossified state, while allowing new bone to be formed at the edges of the bone fronts.