Targeting a c-Myc inhibitory polypeptide to specific intracellular compartments using cell penetrating peptides
ABSTRACT The therapeutic index of current anti-cancer chemotherapeutics can be improved by two major mechanisms: 1) developing drugs which are specifically toxic to the cancer cells and 2) developing methods to deliver drugs to the tumor site. In an attempt to combine these approaches, we developed a thermally responsive polypeptide inhibitor of c-Myc. This polypeptide is based on the thermally responsive Elastin-like polypeptide (ELP). When injected systemically, ELP-fused drugs will aggregate and accumulate at the tumor site where local hyperthermia is applied. ELP was fused to a peptide which blocks c-Myc/Max dimerization (H1), thereby inhibiting transcription activation by c-Myc (ELP-H1). In this study, the cellular uptake, intracellular distribution, and potency of the Pen, Tat and Bac cell penetrating peptides fused to ELP-H1 were evaluated. While Pen-ELP-H1 and Tat-ELP-H1 were localized in the cytoplasm, Bac-ELP-H1 localized to the nucleus in a subset of the cells and was the most potent inhibitor of MCF-7 cell proliferation. This data demonstrates that ELP can be targeted to the desired cellular compartment simply by choice of the CPP used, resulting in a more potent nuclear targeted c-Myc inhibitory polypeptide which may be beneficial in cancer therapy.
- SourceAvailable from: Gene L Bidwell
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- "The peptide has been used extensively in many disease models including AngII-induced hypertension, renovascular hypertension, and arterial balloon injury (reviewed in Cifuentes-Pagano et al., 2012). For application in PE, we fused the Nox2ds peptide to the CPP-ELP carrier (Bidwell and Raucher, 2009). The CPP-ELP carrier will increase the plasma half-life relative to the free peptide, and the use of the CPP will mediate uptake of the polypeptide into target cells in the placenta or systemic vasculature . "
ABSTRACT: The last several decades have seen intensive research into the molecular mechanisms underlying the symptoms of preeclampsia. While the underlying cause of preeclampsia is believed to be defective placental development and resulting placental ischemia, it is only recently that the links between the ischemic placenta and maternal symptomatic manifestation have been elucidated. Several different pathways have been implicated in the development of the disorder; most notably production of the anti-angiogenic protein sFlt-1, induction of auto-immunity and inflammation, and production of reactive oxygen species. While the molecular mechanisms are becoming clearer, translating that knowledge into effective therapeutics has proven elusive. Here we describe a number of peptide based therapies we have developed to target theses pathways, and which are currently being tested in preclinical models. These therapeutics are based on a synthetic polymeric carrier elastin-like polypeptide (ELP), which can be synthesized in various sequences and sizes to stabilize the therapeutic peptide and avoid crossing the placental interface. This prevents fetal exposure and potential developmental effects. The therapeutics designed will target known pathogenic pathways, and the ELP carrier could prove to be a versatile delivery system for administration of a variety of therapeutics during pregnancy.Frontiers in Pharmacology 09/2014; 5:201. DOI:10.3389/fphar.2014.00201 · 3.80 Impact Factor
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- "Because it is genetically encoded, the sequence of ELP is easily manipulated to change the repeat length and resulting molecule size, and the sequence can be modified to incorporate targeting peptides, reactive sites for drug attachment, or therapeutic peptides and proteins (Bidwell and Raucher, 2009; Raucher et al., 2009). ELP has previously been used for delivery of small molecule chemotherapeutics (Bidwell et al., 2007; Dreher et al., 2003; MacKay et al., 2009; Moktan et al., 2012, 2010), peptidebased therapeutics (Bidwell and Raucher, 2010, 2005; Bidwell et al., 2013, 2012, 2010; Massodi et al., 2010, 2009b, 2005), and even protein cargo (Shamji et al., 2008a, 2008b, 2007). A second advantage of ELP derives from the fact that it exhibits reversible temperature-induced aggregation. "
ABSTRACT: Abstract Background: Pregnant females are largely overlooked in drug development due to concerns for fetal health. Additionally, pregnancy is often an exclusion criterion in clinical trials, so the safety of many drugs during pregnancy is unknown. Purpose: The goal of this study was to evaluate Elastin-like Polypeptide (ELP), a synthetic protein derived from human elastin, for maternally sequestered drug delivery. ELP is a versatile drug carrier with a long plasma half-life, low immunogenicity, and the ability to be fused to nearly any small molecule or protein-based therapeutic. Methods: We determined the pharmacokinetics, biodistribution, and fetal exposure to the ELP drug carrier using quantitative fluorescence techniques in a rat pregnancy model. Results: After either bolus IV administration or continuous infusion over five days, ELPs accumulated strongly in the kidneys, liver, and placenta, but importantly, little to no ELPs were detectable in the fetus. Within the placenta, ELPs were localized to the chorionic plate and broadly distributed within the labyrinth, but were excluded from the fetal portion of the chorionic villi. Conclusion: These data indicate that ELP does not cross the placenta, and they suggest that this adaptable drug delivery system is a promising platform for prevention of fetal drug exposure.Journal of Drug Targeting 08/2014; 22(10):1-13. DOI:10.3109/1061186X.2014.950666 · 2.72 Impact Factor
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- "This target temperature range was chosen because it is sufficient to induce aggregation of Bac-ELP1-H1 in plasma at the dose used ( and unpublished data). Due to the sharp temperature transition curve, Bac-ELP1-H1 transitions from completely soluble at 37°C to nearly fully aggregated at 40°C . The rats’ core body temperature was maintained using a homeothermic blanket system, and was not elevated above 37°C during the hyperthermia treatment. "
ABSTRACT: Treatment of glioblastoma is complicated by the tumors' high resistance to chemotherapy, poor penetration of drugs across the blood brain barrier, and damaging effects of chemotherapy and radiation to normal neural tissue. To overcome these limitations, a thermally responsive polypeptide was developed for targeted delivery of therapeutic peptides to brain tumors using focused hyperthermia. The peptide carrier is based on elastin-like polypeptide (ELP), which is a thermally responsive biopolymer that forms aggregates above a characteristic transition temperature. ELP was modified with cell penetrating peptides (CPPs) to enhance delivery to brain tumors and mediate uptake across the tumor cells' plasma membranes and with a peptide inhibitor of c-Myc (H1). In rats with intracerebral gliomas, brain tumor targeting of ELP following systemic administration was enhanced up to 5-fold by the use of CPPs. When the lead CPP-ELP-fused c-Myc inhibitor was combined with focused hyperthermia of the tumors, an additional 3 fold increase in tumor polypeptide levels was observed, and 80% reduction in tumor volume, delayed onset of tumor-associated neurological deficits, and at least doubled median survival time including complete regression in 80% of animals was achieved. This work demonstrates that a c-Myc inhibitory peptide can be effectively delivered to brain tumors.PLoS ONE 01/2013; 8(1):e55104. DOI:10.1371/journal.pone.0055104 · 3.23 Impact Factor