A phase I trial of vertical inhibition of IGF signalling using cixutumumab, an anti-IGF-1R antibody, and selumetinib, an MEK 1/2 inhibitor, in advanced solid tumours.

BACKGROUND: We completed a phase I clinical trial to test the safety and toxicity of combined treatment with cixutumumab (anti-IGF-1R antibody) and selumetinib (MEK 1/2 inhibitor). METHODS: Patients with advanced solid tumours, refractory to standard therapy received selumetinib hydrogen sulphate capsules orally twice daily, and cixutumumab intravenously on days 1 and 15 of each 28-day cycle. The study used a 3+3 design, with a dose-finding cohort followed by an expansion cohort at the maximally tolerated dose that included pharmacokinetic and pharmacodynamic correlative studies. RESULTS: Thirty patients were enrolled, with 16 in the dose-finding cohort and 14 in the expansion cohort. Grade 3 or greater toxicities included nausea and vomiting, anaemia, CVA, hypertension, hyperglycaemia, and ophthalmic symptoms. The maximally tolerated combination dose was 50 mg twice daily of selumetinib and 12 mg kg(-1) every 2 weeks of cixutumumab. Two patients achieved a partial response (one unconfirmed), including a patient with BRAF wild-type thyroid carcinoma, and a patient with squamous cell carcinoma of the tongue, and six patients achieved time to progression of >6 months, including patients with thyroid carcinoma, colorectal carcinoma, and basal cell carcinoma. Comparison of pre- and on-treatment biopsies showed significant suppression of pERK and pS6 activity with treatment. CONCLUSIONS: Our study of anti-IGF-1R antibody cixutumumab and MEK 1/2 inhibitor selumetinib showed that the combination is safe and well-tolerated at these doses, with preliminary evidence of clinical benefit and pharmacodynamic evidence of target inhibition.

Potential role of tumor vaccines in GI malignancies.

Although surgery remains the only curative option for patients with gastrointestinal (GI) malignancies, the use of adjuvant chemotherapy and/or localized radiation is considered standard therapy for patients who present with locoregional disease. Even with adjuvant therapy, however, the 5-year survival rate for such patients ranges from 2% to 50%, depending on the specific cancer type and stage. As a result, more effective interventions are necessary for all but the earliest stages of GI malignancies. Colon cancer represents the paradigm for the management of GI malignancies, not only because it is, by far, the most common cancer in this group, but also because the biological progression to disease is well characterized. Immunotherapy is an alternative approach for treating GI malignancies that can either: (1) activate tumor-specific T-cells; or (2) use monoclonal antibodies derived from tumor-specific antigens. Monoclonal antibodies act by a mechanism that is distinct from that of chemotherapy and, thus, represent a non-cross-resistant treatment with an entirely different spectrum of toxicities. Thanks to an improved understanding of tumor immunology, as well as the events needed to generate an optimal immune response, the possibility of designing an effective colon cancer vaccine approach that induces both humoral and cellular responses has become even more realistic.

A fourth human MEF2 transcription factor, hMEF2D, is an early marker of the myogenic lineage.

The transition from multipotent mesodermal precursor to committed myoblast and its differentiation into a mature myocyte involve molecular events that enable the cell to activate muscle-specific genes. Among the participants in this process is the myocyte-specific enhancer factor 2 (MEF2) family of tissue-restricted transcription factors. These factors, which share a highly conserved DNA-binding domain including a MADS box, are essential for the expression of multiple muscle genes with cognate target MEF2 sites in cis. We report here a new human MEF2 factor, hMEF2D, which is unique among the members of this family in that it is present not only in myotubes but also in undifferentiated myoblasts, even before the appearance of myogenin. hMEF2D comprises several alternatively spliced products of a single gene, one of which is the human homolog of the Xenopus SRF-related factor SL-1. Like its relatives, cloned hMEF2D is capable of activating transcription via sequence-specific binding to the MEF2 site, recapitulating endogenous tissue-specific MEF2 activity. Indeed, while MEF2D mRNAs are ubiquitous, the protein is highly restricted to those cell types that contain this activity, implicating posttranscriptional mechanisms in the regulation of MEF2D expression. Alternative splicing may be important in this process: two alternative MEF2D domains, at least one of which is specifically included during myogenic differentiation, also correlate precisely with endogenous MEF2 activity. These findings provide compelling evidence that MEF2D is an integral link in the regulatory network for muscle gene expression. Its presence in undifferentiated myoblasts further suggests that it may be a mediator of commitment in the myogenic lineage.

Probing the determinants of disulfide stability in native pancreatic trypsin inhibitor.

The effects of single amino acid replacements on the stability of the 14-38 disulfide bond in the native form of bovine pancreatic trypsin inhibitor (BPTI) were measured. A total of 17 mutant proteins, with substitutions at one of 7 residues located 5-15 A from the disulfide in the native wild-type protein, were examined. The replacements were found to decrease the thermodynamic stability of the disulfide, as measured by exchange with thiol-disulfide reagents, by 0.6-5 kcal/mol, corresponding to a range of nearly 100 mV in redox potentials. The effects of the substitutions on disulfide stability were roughly correlated with the changes in side-chain volume, suggesting that optimal packing is a major factor in determining the stability of the disulfide in the wild-type protein. With only one exception, the substitutions also led to increases, as large as 50-fold, in the rates of disulfide reduction by dithiothreitol. The increased rates of reduction suggest that at least a fraction of the mutational destabilization of the disulfide is due to strain in the native protein that is relieved in the transition state for reduction. The stability of the disulfide in a peptide corresponding to the segments that are linked together by the 14-38 disulfide in native BPTI was found to be about 5 kcal/mol less than that of the disulfide in the intact wild-type protein. Together, the results with the mutant proteins and the peptide indicate that the stability of the disulfide in the native protein depends on both the local environment of the disulfide and on the ability of the rest of the protein to favor a conformation that promotes disulfide formation.

Genetic dissection of pancreatic trypsin inhibitor.

In a previous study, a genetic screening procedure was used to identify variants of bovine pancreatic trypsin inhibitor that can fold to an active conformation but that are inactivated much more rapidly than the wild-type protein in the presence of dithiothreitol (DTT). The mechanisms by which 30 of these DTT-sensitive variants are inactivated have now been investigated. Some of the amino acid replacements cause rapid inactivation in the presence of DTT because the three disulfides of the native protein are reduced up to 300-fold faster than for the wild-type protein, leading to complete unfolding. Other substitutions, however, do not greatly increase the rate of complete reduction and unfolding but lead to accumulation of an inactive two-disulfide species. There is a striking correlation between the locations of the DTT-sensitive amino acid replacements in the three-dimensional structure of the protein and the mechanisms by which the variants are inactivated. All of the substitutions that cause rapid unfolding are clustered at one end of the folded protein, in the vicinity of the two disulfides that are reduced most slowly during unfolding of the wild-type protein, while substitutions of the other class are all located at the other end of the protein, near the trypsin binding site. These results indicate that the kinetic stability of native bovine pancreatic trypsin inhibitor and its ability to function as a protease inhibitor are largely influenced by residues in two distinguishable regions of the folded protein.