Phase I study of CC-90010, a reversible, oral BET inhibitor in patients with advanced solid tumors and relapsed/refractory non-Hodgkin's lymphoma.

BACKGROUND: Bromodomain and extra-terminal (BET) proteins are epigenetic readers that regulate expression of genes involved in oncogenesis. CC-90010 is a novel, oral, reversible, small-molecule BET inhibitor. PATIENTS AND METHODS: CC-90010-ST-001 (NCT03220347; 2015-004371-79) is a phase I dose-escalation and expansion study of CC-90010 in patients with advanced or unresectable solid tumors and relapsed/refractory (R/R) non-Hodgkin's lymphoma (NHL). We report results from the dose escalation phase, which explored 11 dose levels and four dosing schedules, two weekly (2 days on/5 days off; 3 days on/4 days off), one biweekly (3 days on/11 days off), and one monthly (4 days on/24 days off). The primary objectives were to determine the safety, maximum tolerated dose (MTD) and/or recommended phase II dose (RP2D) and schedule. Secondary objectives were to evaluate signals of early antitumor activity, pharmacokinetics, and pharmacodynamics. RESULTS: This study enrolled 69 patients, 67 with solid tumors and two with diffuse large B-cell lymphoma (DLBCL). The median age was 57 years (range, 21-80) and the median number of prior regimens was four (range, 1-9). Treatment-related adverse events (TRAEs) were mostly mild and manageable; grade 3/4 TRAEs reported in more than two patients were thrombocytopenia (13%), anemia, and fatigue (4% each). Six patients had dose-limiting toxicities. MTDs were 15 mg (2 days on/5 days off), 30 mg (3 days on/11 days off), and 45 mg (4 days on/24 days off). The RP2D and schedule selected for expansion was 45 mg (4 days on/24 days off). As of 8 October 2019, one patient with grade 2 astrocytoma achieved a complete response, one patient with endometrial carcinoma had a partial response, and six patients had prolonged stable disease ≥11 months. CONCLUSIONS: CC-90010 is well tolerated, with single-agent activity in patients with heavily pretreated, advanced solid tumors.

Expression of a truncated, kinase-defective TGF-beta type II receptor in mouse skeletal tissue promotes terminal chondrocyte differentiation and osteoarthritis.

Members of the TGF-beta superfamily are important regulators of skeletal development. TGF-betas signal through heteromeric type I and type II receptor serine/threonine kinases. When over-expressed, a cytoplasmically truncated type II receptor can compete with the endogenous receptors for complex formation, thereby acting as a dominant-negative mutant (DNIIR). To determine the role of TGF-betas in the development and maintenance of the skeleton, we have generated transgenic mice (MT-DNIIR-4 and -27) that express the DNIIR in skeletal tissue. DNIIR mRNA expression was localized to the periosteum/perichondrium, syno-vium, and articular cartilage. Lower levels of DNIIR mRNA were detected in growth plate cartilage. Transgenic mice frequently showed bifurcation of the xiphoid process and sternum. They also developed progressive skeletal degeneration, resulting by 4 to 8 mo of age in kyphoscoliosis and stiff and torqued joints. The histology of affected joints strongly resembled human osteo-arthritis. The articular surface was replaced by bone or hypertrophic cartilage as judged by the expression of type X collagen, a marker of hypertrophic cartilage normally absent from articular cartilage. The synovium was hyperplastic, and cartilaginous metaplasia was observed in the joint space. We then tested the hypothesis that TGF-beta is required for normal differentiation of cartilage in vivo. By 4 and 8 wk of age, the level of type X collagen was increased in growth plate cartilage of transgenic mice relative to wild-type controls. Less proteoglycan staining was detected in the growth plate and articular cartilage matrix of transgenic mice. Mice that express DNIIR in skeletal tissue also demonstrated increased Indian hedgehog (IHH) expression. IHH is a secreted protein that is expressed in chondrocytes that are committed to becoming hypertrophic. It is thought to be involved in a feedback loop that signals through the periosteum/ perichondrium to inhibit cartilage differentiation. The data suggest that TGF-beta may be critical for multifaceted maintenance of synovial joints. Loss of responsiveness to TGF-beta promotes chondrocyte terminal differentiation and results in development of degenerative joint disease resembling osteoarthritis in humans.

Bone remodelling: a signalling system for osteoclast regulation.

Two physiological regulators of osteoclast maturation have recently been identified: the secreted protein osteoprotegerin and the cell-surface ligand to which it binds. These proteins are likely to play an important part in the control of bone resorption, but are also likely to have important roles in other tissues.

Specific changes of Ras GTPase-activating protein (GAP) and a GAP-associated p62 protein during calcium-induced keratinocyte differentiation.

Induction of tyrosine phosphorylation occurs as an early and specific event in keratinocyte differentiation. A set of tyrosine-phosphorylated substrates which transduce mitogenic signals by tyrosine kinases has previously been identified. We show here that of these substrates, the Ras GTPase-activating protein, GAP, is specifically affected during calcium-induced keratinocyte differentiation. As early as 10 min after calcium addition to cultured primary mouse keratinocytes, GAP associates with tyrosine-phosphorylated proteins and translocates to the membrane. In addition, a GAP-associated protein of approximately 62 kDa (p62) becomes rapidly and heavily tyrosine phosphorylated in both membrane and cytosolic fractions. This protein corresponds to the major tyrosine-phosphorylated protein that is induced in differentiating keratinocytes as early as 5 min after calcium addition. p62 phosphorylation was not observed after exposure of these cells to epidermal growth factor, phorbol ester, or transforming growth factor beta. In contrast, PLC gamma and P13K were tyrosine phosphorylated after epidermal growth factor, but not calcium, stimulation. Thus, changes of Ras GAP and an associated p62 protein occur as early and specific events in keratinocyte differentiation and appear to involve a calcium-induced tyrosine kinase.

Tyrosine phosphorylation is an early and specific event involved in primary keratinocyte differentiation.

Very little is known about early molecular events triggering epithelial cell differentiation. We have examined the possible role of tyrosine phosphorylation in this process, as observed in cultures of primary mouse keratinocytes after exposure to calcium or 12-O-tetradecanoylphorbol-13-acetate (TPA). Immunoblotting with phosphotyrosine-specific antibodies as well as direct phosphoamino acid analysis revealed that induction of tyrosine phosphorylation occurs as a very early and specific event in keratinocyte differentiation. Very little or no induction of tyrosine phosphorylation was observed in a keratinocyte cell line resistant to the differentiating effects of calcium. Treatment of cells with tyrosine kinase inhibitors prevented induction of tyrosine phosphorylation by calcium and TPA and interfered with the differentiative effects of these agents. These results suggest that specific activation of tyrosine kinase(s) may play an important regulatory role in keratinocyte differentiation.

Osteoblastic responses to TGF-beta during bone remodeling.

Bone remodeling depends on the spatial and temporal coupling of bone formation by osteoblasts and bone resorption by osteoclasts; however, the molecular basis of these inductive interactions is unknown. We have previously shown that osteoblastic overexpression of TGF-beta2 in transgenic mice deregulates bone remodeling and leads to an age-dependent loss of bone mass that resembles high-turnover osteoporosis in humans. This phenotype implicates TGF-beta2 as a physiological regulator of bone remodeling and raises the question of how this single secreted factor regulates the functions of osteoblasts and osteoclasts and coordinates their opposing activities in vivo. To gain insight into the physiological role of TGF-beta in bone remodeling, we have now characterized the responses of osteoblasts to TGF-beta in these transgenic mice. We took advantage of the ability of alendronate to specifically inhibit bone resorption, the lack of osteoclast activity in c-fos-/- mice, and a new transgenic mouse line that expresses a dominant-negative form of the type II TGF-beta receptor in osteoblasts. Our results show that TGF-beta directly increases the steady-state rate of osteoblastic differentiation from osteoprogenitor cell to terminally differentiated osteocyte and thereby increases the final density of osteocytes embedded within bone matrix. Mice overexpressing TGF-beta2 also have increased rates of bone matrix formation; however, this activity does not result from a direct effect of TGF-beta on osteoblasts, but is more likely a homeostatic response to the increase in bone resorption caused by TGF-beta. Lastly, we find that osteoclastic activity contributes to the TGF-beta-induced increase in osteoblast differentiation at sites of bone resorption. These results suggest that TGF-beta is a physiological regulator of osteoblast differentiation and acts as a central component of the coupling of bone formation to resorption during bone remodeling.

Binding protein-3-selective insulin-like growth factor I variants: engineering, biodistributions, and clearance.

Insulin-like growth factor I (IGF-I) is a potent anabolic peptide that mediates most of its pleiotropic effects through association with the IGF type I receptor. Biological availability and plasma half-life of IGF-I are modulated by soluble binding proteins (IGFBPs), which sequester free IGF-I into high affinity complexes. Elevated levels of specific IGFBPs have been observed in several pathological conditions, resulting in inhibition of IGF-I activity. Administration of IGF-I variants that are unable to bind to the up-regulated IGFBP species could potentially counteract this effect. We engineered two IGFBP-selective variants that demonstrated 700- and 80,000-fold apparent reductions in affinity for IGFBP-1 while preserving low nanomolar affinity for IGFBP-3, the major carrier of IGF-I in plasma. Both variants displayed wild-type-like potency in cellular receptor kinase assays, stimulated human cartilage matrix synthesis, and retained their ability to associate with the acid-labile subunit in complex with IGFBP-3. Furthermore, pharmacokinetic parameters and tissue distribution of the IGF-I variants in rats differed from those of wild-type IGF-I as a function of their IGFBP affinities. These IGF-I variants may potentially be useful for treating disease conditions associated with up-regulated IGFBP-1 levels, such as chronic or acute renal and hepatic failure or uncontrolled diabetes. More generally, these results suggest that the complex biology of IGF-I may be clarified through in vivo studies of IGFBP-selective variants.

Soluble form of an HLA-B7 class I antigen specifically suppresses humoral alloimmunization.

A soluble HLA-B7 molecule, designated sB7 and generated by genetically engineering the B7 gene to remove the transmembrane and cytoplasmic domains, was tested as a tolerogen. Supernatants from cultures of C1R cells transfected with the gene for sB7 were harvested and concentrated, as were control supernatants. From days -17 to -1, C57Bl/6 mice were pretreated with a total of 11 intraperitoneal doses of 1.0 microgram each of sB7 or appropriate control supernatant, and then were challenged intraperitoneally on each of days 0, 7, and 14 with 10(6) C1R-B7 cells (expressing surface HLA-B7). Antibody kinetics revealed (1) anti-B7 was not induced after sB7 pretreatment; (2) the anti-B7 response of sB7-pretreated mice was marginal and of apparent low avidity compared with the brisk anti-B7 response of control mice; (3) none of the mice made antibody to a control HLA antigen, A24; (4) all mice made strong antibody responses to the non-B7 surface antigens of C1R; (5) free sB7 did not appear in the blood of the treated mice; and (6) all mice appeared to be generally healthy. These data show soluble B7 antigen is not immunogenic and appears to specifically block humoral immune response to cell membrane-bound HLA-B7 in a nontoxic manner.

Bioengineered soluble HLA-B7. Genesis, characterization, and occurrence of dimerization.

A soluble, secreted form of HLA-B7 was engineered by replacing the exons encoding the transmembrane and cytoplasmic domains of the B7 gene with a CI. The modified gene, gsB7, transfected into J27.2 or C1R cell lines, produced a secreted protein, sB7, serologically recognized as B7. Size fractionation showed one species of sB7 at the approximately 55 kD expected for an sB7 alpha-chain-beta 2m heteroduplex, and another at approximately 120 kD which had the same constituent chains and was a dimer of the 55-kD species. Dimer formation appeared to be related to protein concentration but not to disulfide bridging. The sB7 heavy chain on SDS-PAGE showed a doublet at approximately 39 and approximately 42 kD; enzyme analysis indicated that the two bands differed only by a carboxyl terminal polypeptide. Analysis of gsB7 transfectants' mRNA by Northern blots and PCR revealed message fully spliced or with retained CI, accounting for the 39- and 42-kD bands, respectively, and apparently untranslated message with I3 retained. sB7 was not detectable on the surface of gsB7 transfectants by CTLs, nor did it inhibit those CTLs. Production of the sB7 protein provides a ready, consistent source of soluble class I antigen for further study, including test materials for tolerogenicity studies in animal models.

Functional inactivation of the IGF-I receptor delays differentiation of skeletal muscle cells.

Skeletal myoblasts are inherently programmed to leave the cell cycle and begin the differentiation process following removal of exogenous growth factors. Serum withdrawal results in a marked induction of IGF production which is essential for skeletal muscle differentiation in vitro. However, the potential role of the tyrosine kinase IGF-I receptor (thought to be the principal mediator of both IGF-I and II signaling in skeletal muscle) in the decision of myoblasts to begin differentiation following serum withdrawal is unknown. To explore the role of the IGF-I receptor in this decision by skeletal myoblasts, we functionally inactivated endogenous IGF-I receptors in mouse C2C12 cells using a dominant negative, kinase-inactive IGF-I receptor in which the ATP-binding site lysine (K) at residue 1003 has been mutated to alanine (A). Cell lines with the greatest degree of mutant IGF-I receptor expression (A/K cells) demonstrated functional inactivation of endogenous IGF-I receptors as determined by their impaired ability to phosphorylate the principal substrate of the IGF-I receptor, IRS-1, in response to treatment with IGF-I. In addition, the proliferative response of myoblasts to IGF-I was completely abolished in A/K cells. Following withdrawal of exogenous growth factors, A/K cells demonstrated a marked delay in the induction of the gene expression of myogenin, a skeletal muscle-specific transcription factor essential for differentiation, and a subsequent delay in the induction of muscle creatine kinase activity. Delayed differentiation in A/K cells was associated with prolonged phosphorylation of the cell cycle regulatory retinoblastoma (Rb) protein; it is the un- (or hypo-) phosphorylated form of Rb which is known to promote differentiation in skeletal myoblasts. Thus, the IGF-I receptor regulates the timing of myoblast differentiation induced by serum withdrawal. The delayed differentiation of skeletal myoblasts with functionally inactive IGF-I receptors may result, at least in part, from delayed induction of myogenin gene expression and prolonged phosphorylation of the Rb protein.

Induction of myogenesis in mesenchymal cells by MyoD depends on their degree of differentiation.

Expression of a transfected MyoD gene induces myogenic differentiation of most cell types. In this study, we evaluated the ability of an exogenous MyoD gene to induce myogenic conversion in two pairs of matched cell lines with different degrees of differentiation within either the osteoblastic or chondrocytic lineage. We show that osteoblasts and chondrocytes are resistant to the myogenic effects of MyoD alone. However, in their less-differentiated cell line counterparts, MyoD induces expression of muscle-cell-specific markers. Less-differentiated osteoblasts can be made resistant to MyoD-induced myogenic conversion by induction of adipogenic differentiation using dexamethasone. Finally, a dominant positive form of MyoD, one which is tethered to a partner, E47, activates muscle-specific gene expression in osteoblasts. Our results suggest that the response of a cell to MyoD depends on its lineage and its degree of differentiation. Furthermore, commitment of cells to the osteoblastic or chondrocytic lineage may involve inhibition of alternative pathways, such as those leading to myoblastic differentiation. Finally, osteoblasts may express a protein(s) which interferes with the activity of MyoD by inhibiting its association with E proteins. This interference can be overcome by expression of the MyoD-E47 hybrid, suggesting that osteoblasts are otherwise competent to undergo myogenic conversion.

Functional evidence for an extracellular calcium receptor mechanism triggering tyrosine kinase activation associated with mouse keratinocyte differentiation.

Calcium-induced keratinocyte differentiation is associated with tyrosine phosphorylation of a p62 protein which associates with the ras-GTPase activating protein (GAP). We have examined the nature of the calcium signal triggering p62 phosphorylation. EGTA, a specific chelator of calcium, was able to completely block calcium-induced p62 phosphorylation, even after using conditioned medium from calcium-treated keratinocytes. Preventing calcium-induced cell-cell contacts by anti-cadherin antibodies did not inhibit tyrosine phosphorylation. Slight increases in extracellular calcium concentrations (0.15 or 0.30 mM) were already sufficient to induce p62 phosphorylation. Other divalent cations, such as magnesium, zinc, nickel, and cobalt, but not the trivalent cation lanthanum, induced p62 phosphorylation to a similar extent as calcium. There was no close correlation between the ability of the various ions to induce p62 phosphorylation and increase free intracellular calcium. Similarly, treatment of primary keratinocytes with the calcium ionophores A23187 or X537A did not induce p62 phosphorylation, although it increased free intracellular calcium levels. Finally, blockers of potassium uptake, which is induced by calcium, did not inhibit p62 phosphorylation. Thus, in keratinocyte differentiation, calcium is likely to provide the primary signal for p62 tyrosine phosphorylation and may act directly at the cell membrane through a "cationic receptor mechanism" analogous to that described in other cell types.

Induction of transforming growth factor beta 1 resistance by the E1A oncogene requires binding to a specific set of cellular proteins.

Transforming growth factors beta (TGF-beta s) are potent inhibitors of epithelial cell growth in culture and might play a similar role in vivo. Several studies have suggested that acquisition of TGF-beta resistance is an important step in epithelial tumor development. Here, we show that resistance to TGF-beta 1 growth inhibition can be induced by transformation of keratinocytes with the E1A, but not the ras, oncogene. Mutational analysis revealed that these effects closely correlate with the ability of E1A proteins to bind to the retinoblastoma gene product (p105) as well as to three other cellular proteins (p60, p107, and p300). Only partial resistance to TGF-beta 1 growth inhibition was elicited by E1A mutants that bind to a subset of proteins, whereas complete resistance was induced by E1A mutants that bind to all four proteins together. Total protection against TGF-beta growth inhibition was also induced by concomitant introduction into cells of an E1A mutant binding to the p60/p105/p107 proteins and one binding to p300. In parallel with these effects, epidermal transglutaminase, a marker of keratinocyte differentiation, was induced by TGF-beta in control but not in E1A-transformed cells. TGF-beta 1 receptor levels were only partially down-modulated by an intact E1A gene and not significantly affected by the various truncated mutants. Thus, the ability of E1A to induce TGF-beta resistance depends on its ability to bind, and presumably inactivate, several cellular proteins that may be involved in transmission of the TGF-beta signal and seem to act downstream from its receptor(s).

Inhibition of myogenic differentiation in myoblasts expressing a truncated type II TGF-beta receptor.

Transforming growth factor-beta (TGF-beta) is thought to play a role in mesenchymal cell development and, specifically, in muscle differentiation, yet its precise role in the latter process remains unclear. TGF-beta has been shown to both inhibit and induce myoblast maturation in vitro, depending on the culture conditions. Whether the type I or type II TGF-beta receptor mediates the various TGF-beta effects on myogenesis is not known. In the present study, C2C12 myoblasts were transfected with an expression vector for a truncated type II TGF-beta receptor, which has been shown to act as a dominant negative inhibitor of type II receptor signaling. In contrast to the parental cells, the transfected clones did not efficiently form myotubes or induce expression of MyoD, myogenin and several other differentiation markers following incubation in low serum media. However, some muscle differentiation markers continued to be expressed in the transfected cells suggesting that at least two pathways are involved in muscle cell differentiation. These cells could still growth arrest in low serum media, showing that decreased proliferation can be dissociated from differentiation. Unlike several oncogenes known to block myogenic differentiation, expression of the truncated TGF-beta receptor did not result in myoblast transformation. Injection of the parental or the transfected C2C12 cells into the limb muscle of nude mice revealed quantitative and qualitative differences in their behavior, and suggested that myoblasts expressing the truncated TGF-beta receptor cannot fuse in vivo. Finally, retrovirus-mediated expression of MyoD in the transfected cells restored their ability to form myotubes in vitro, indicating that inhibition of myoblast differentiation by the truncated TGF-beta receptor may depend on decreased MyoD expression. We propose that TGF-beta signaling through the type II receptor is required for several distinct aspects of myogenic differentiation and that TGF-beta acts as a competence factor in this multistep process.

Transforming growth factor-beta (TGF-beta)-induced down-regulation of cyclin A expression requires a functional TGF-beta receptor complex. Characterization of chimeric and truncated type I and type II receptors.

Transforming growth factor-beta (TGF-beta) inhibits the proliferation of epithelial cells by altering the expression or function of various components of the cell cycle machinery. Expression of one of these components, cyclin A, is inhibited by TGF-beta treatment. We have identified a 760-base pair fragment of the human cyclin A gene promoter that is sufficient to confer TGF-beta responsiveness. Using this promoter fragment, we have developed a cyclin A-based luciferase reporter assay that quantitates the growth inhibitory effect of TGF-beta in transient transfection assays. This assay was used to determine which domains of the type I (RI) and type II (RII) receptors were required for the antiproliferative effect of TGF-beta. In parallel, the functionality of chimeric receptors, between RI and RII (RI-RII or RII-RI), was tested for TGF-beta effect on gene expression using a reporter assay based on the plasminogen activator inhibitor type 1 (PAI-1) promoter. We found that TGF-beta-induced inhibition of cyclin A expression was absent in RI or RII-deficient Mv1Lu cells and that this response was restored by expression of wild-type type I or type II receptors in these cells. Furthermore, expression of a single chimeric receptor, either RI-RII or RII-RI, did not confer cyclin A regulation by TGF-beta. However, expression of two reciprocal chimeras (RI-RII and RII-RI) resulted in growth inhibition, similarly to wild-type receptors. In addition, chimeric receptors as well as mutant receptors with a deleted cytoplasmic domain and kinase-negative receptors inhibited TGF-beta responsiveness in the cyclin A reporter assay in a dominant negative fashion. Finally, in both receptor types, the juxtamembrane domain preceding the kinase domain was essential for receptor function but the cytoplasmic tail was dispensable. Our results suggest that a functional TGF-beta receptor complex is required for TGF-beta-dependent down-regulation of cyclin A gene expression and illustrate the identical receptor requirements for TGF-beta-induced growth inhibition and gene expression.

Induction of different morphologic features of malignant melanoma and pigmented lesions after transformation of murine melanocytes with bFGF-cDNA and H-ras, myc, neu, and E1a oncogenes.

Malignant melanomas show a remarkable degree of heterogeneity because of different morphologic features, biologic behavior, and prognosis. In this communication, the authors attempted to correlate morphologic heterogeneity of melanomas with transformation by different activated oncogenes; they studied the histologic features of melanocytic lesions induced by murine melanocytes transformed by basic fibroblast growth factor (b-FGF-cDNA) or H-ras, neu, myc, and E1a oncogenes, and the lesions were compared with those observed in human pathology. Tumors formed after grafting onto syngenic mice or subcutaneous injections in nude mice were studied. In syngenic mice, benign melanocytic lesions reminiscent of intradermal nevus were observed with melanocytes transformed with b-FGF-cDNA, and myc and E1a oncogenes. Benign lesions were also formed by neu-transformed melanocytes when they were grafted concomitantly with keratinocytes, whereas malignant tumors were formed by the same cells when grafted alone or together with fibroblasts. In contrast, H-ras melanocytes always formed malignant tumors. In nude mice, b-FGF-transformed melanocytes induced benign lesions, whereas transformed melanocytes by the other oncogenes formed malignant tumors with distinctive and homogeneous morphologic features that depended on the transforming oncogene. Melanomas with either epithelioid cell, spindle cell, small round cell, and anaplastic cell growth patterns could be distinguished after transformation with H-ras, neu, E1a, and myc oncogenes, respectively. These various histologic types are analogous to those that may be observed in human melanomas, even within the same tumor. These studies suggest a possible molecular mechanism for tumor heterogeneity in which distinct oncogenes or oncogenelike activities can be activated in different tumors or discrete parts of the same tumor.

An emerging complexity of receptors for transforming growth factor-beta.

Transforming growth factor-beta (TGF-beta) is a multifunctional protein that modulates cell proliferation and interaction with the extracellular matrix. Three common TGF-beta receptors are found on the cell surface. The type III receptor is a transmembrane proteoglycan with a short cytoplasmic domain and is thought not to be involved in TGF-beta induced signalling. In contrast, the type II and type I receptors are transmembrane serine/threonine kinases. The type II receptor determines the ligand specificity, whereas the type I receptor interacts with the type II receptor and may not have a ligand binding specificity by itself. Both type II and type I receptors are involved in TGF-beta induced signalling. The type II receptor, likely in conjunction with the type I receptor, is required for the antiproliferative effect of TGF-beta, whereas the type I receptor is the likely mediator of the effects of TGF-beta on the expression of several genes including some extracellular matrix proteins. To address the role of TGF-beta signalling in myoblast differentiation, we transfected a dominant negative mutant of the type II receptor in myoblasts, thus inhibiting type II receptor mediated signalling. These cells not longer had the ability to differentiate in vitro or in vivo, suggesting that TGF-beta signalling through the type II receptor provides competence for myoblastic differentiation. These studies also indicate that there are several signalling pathways involved in myoblastic differentiation, one of which is modulated by the TGF-beta signalling.

The effect of TGF-beta 2 on dentin apposition and hardness in transgenic mice.

Transforming growth factor beta, TGF-beta, is expressed during tooth formation and can induce pre-odontoblast differentiation and formation of functional odontoblast-like cells in vitro. In addition, exogenous TGF-beta can increase reparative dentin formation, presumably by acting on odontoblasts. In this study, we examined the tooth phenotype of transgenic mice, in which TGF-beta 2 expression is directed by the osteocalcin promoter. Previous studies have shown that these mice have a bone phenotype that resembles that of human osteoporosis, including the existence of spontaneous fractures. Microhardness testing of the enamel and dentin showed no differences in the molars of these transgenic mice as compared with those of their wild-type littermates. Consistent with the increase in bone mineral apposition rate previously reported in these mice, the dentin apposition rate appeared to be increased in the TGF-beta 2-overexpressing mice. Thus, in teeth, as in bone, TGF-beta 2 appears to stimulate the synthesis and deposition of matrix. Further studies are needed to understand the effect of TGF-beta 2 on distinct mineralized tissues (bone, dentin, and cementum) and to determine whether exogenous TGF-beta 2 may be useful for tooth repair.