Emerging antibacterial resistance is a consequence of the continued use of our current antibacterial therapies, and it is limiting their utility, especially for infections caused by multidrug-resistant isolates. β-Lactams have enjoyed extensive clinical success, but their broad usage is linked to perhaps the most extensive and progressive example of resistance development for any antibacterial scaffold. In Gram-negative pathogens, this largely involves constant evolution of new β-lactamases able to degrade successive generations of this scaffold. In addition, more recently, alterations in the targets of these compounds, penicillin-binding proteins (PBPs), are being described in clinical isolates, which often also have multiple β-lactamases. This study underscores the multifactorial nature of β-lactam resistance by uncovering alterations of PBP2 that reduce susceptibility to carbapenems in E. coli clinical isolates that also have alterations of PBP3 and express the NDM-1 β-lactamase. The changes in PBP2 also reduced susceptibility to the intrinsic antibacterial activity of some diazabicyclooctane (DBO) compounds that can target PBP2. This may have implications for the development and use of the members of this relatively newer scaffold that are inhibitors of PBP2 in addition to their inhibition of serine-β-lactamases.

A partial cDNA clone for the B-26 region of apolipoprotein B was isolated from an adult human liver DNA library by screening with an oligonucleotide probe derived from amino-terminal protein sequence obtained from purified B-26 peptide. Antisera against a synthetic 17-residue peptide whose amino acid sequence was encoded by the clone cross. reacts with apolipoproteins B 26, B -100, and B -48, but not with B-74. The nucleotide sequence immediately upstream from the amino terminus of B-26 codes for an apparent signal sequence, implying that the B-26 moiety is in an amino-terminal locus in the B- 100 protein. That this' sequence represents a S' end region is further supported by primer extension analysis using a fragment of the cDNA clone and by S1 nuclease protection experiments using the corresponding region in a genomic clone.

Transforming growth factor-beta (TGF-beta) is a proinvasive and immunosuppressive cytokine that plays a major role in the malignant phenotype of gliomas. One novel strategy of disabling TGF-beta activity in gliomas is to disrupt the signaling cascade at the level of the TGF-beta receptor I (TGF-betaRI) kinase, thus abrogating TGF-beta-mediated invasiveness and immune suppression. SX-007, an orally active, small-molecule TGF-betaRI kinase inhibitor, was evaluated for its therapeutic potential in cell culture and in an in vivo glioma model. The syngeneic, orthotopic glioma model SMA-560 was used to evaluate the efficacy of SX-007. Cells were implanted into the striatum of VM/Dk mice. Dosing began three days after implantation and continued until the end of the study. Efficacy was established by assessing survival benefit. SX-007 dosed at 20 mg/kg p.o. once daily (q.d.) modulated TGF-beta signaling in the tumor and improved the median survival. Strikingly, approximately 25% of the treated animals were disease-free at the end of the study. Increasing the dose to 40 mg/kg q.d. or 20 mg/kg twice daily did not further improve efficacy. The data suggest that SX-007 can exert a therapeutic effect by reducing TGF-beta-mediated invasion and reversing immune suppression. SX-007 modulates the TGF-beta signaling pathway and is associated with improved survival in this glioma model. Survival benefit is due to reduced tumor invasion and reversal of TGF-beta-mediated immune suppression, allowing for rejection of the tumor. Together, these results suggest that treatment with a TGF-betaRI inhibitor may be useful in the treatment of glioblastoma.

6033 Many cancers overexpress transforming growth factor beta (TGF-β), a protein that plays a major role in tumor progression by regulating cell proliferation, angiogenesis, metastasis and immunosupression. Glioblastoma multiformae (GBM) is one such cancer. TGF-β secretion may play a role in the highly invasive nature of the disease and appears to be responsible for the suppressed immune system in GBM patients. The therapeutic potential of SX-007, an orally bioavailable small molecule TGF-β RI kinase inhibitor (IC50=33 nM), was investigated in the SMA560 model, a syngeneic murine model of glioma. SMA560 cells were implanted into the right striatum of male VM/Dk mice. Dosing began three days after implanting tumor cells and continued until the end of the study. The efficacy of SX-007 was assessed at 5, 20 and 40 mg/kg po qd in the SMA560 model. Maximal efficacy was observed at 20 mg/kg (median survival=24 days). Increasing dosing from 20 mg/kg once to twice daily did not result in improved survival benefit. From each study, approximately 20% of animals from the groups dosed with 20 mg/kg SX-007 remained alive at the end of the study (Day 35). These long-term survivors appear to be disease-free. In a separate study, the animals were sacrificed at Day 15 and brain tissue removed to verify that SX-007 reached the site of action. The levels of phosphorylated Smad 2/3 and transcription of TGF-β related genes (most notable, PAI-1) were reduced in tumors from compound-treated animals confirming an inhibition of TGF-β signaling. In addition, there was increased CD3+ T-cell infiltration into the tumor and a concomitant increase in caspase-3 staining in the brain tissue of compound-treated animals as measured by immunohistochemistry demonstrating a functional consequence of inhibiting TGF-β signaling. SX-007 can modulate TGF-β signaling pathway in vivo and treatment with SX-007 is associated with an improvement in animal survival. Survival benefit is due, at least in part, to a reversal of the immune suppressed state allowing for rejection of the tumor. Thus SX-007 may be useful in the treatment of glioblastoma.