Biomedical subjects

Joseph A Reddy

Publications and source records attributed to Joseph A Reddy.

10 recordsLinked to original sources

PubMed2006

Preclinical pharmacokinetics, tissue distribution, and antitumor activity of a folate-hapten conjugate-targeted immunotherapy in hapten-immunized mice.

Folic acid (pteroylglutamic acid) represents a useful ligand for targeted cancer therapies because it binds to a common epithelial tumor antigen known as the folate receptor. We previously devised an immunotherapy strategy that uses a bispecific ligand, a folate-hapten (FITC) conjugate, to redirect endogenously induced anti-FITC antibodies to folate receptor-positive tumor cells following parenteral administration. Here, we present results from preclinical pharmacokinetic and tissue biodistribution studies using a radioactive folate-FITC conjugate and results from dose optimization studies done in tumor-bearing animals. Folate-FITC was found to be rapidly eliminated in non-immunized mice; however, in immunized hosts, folate-FITC was shown to form immune complexes with FITC-specific antibodies, the consequence of which was a approximately 173-fold increase in drug exposure (i.e., area under the curve). Using a newly developed ELISA assay, the extent of circulating anti-FITC antibodies occupied by parenterally given folate-FITC was determined to be proportional to the given dose. Furthermore, high doses of folate-FITC were found to promote the cosaturation of tumor cell surface folate receptors and circulating FITC-specific antibodies, blocking the immune recognition of tumor cells and thereby reducing antitumor activity. Nonetheless, by extending the duration of treatment and administering subsaturating doses of folate-FITC, enhanced antitumor response was observed in mice bearing established folate receptor-positive M109 tumors. Overall, results from the present study may help to guide clinicians through on-going clinical investigations of folate-targeted immunotherapy.

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PubMed2005

Folate receptor specific anti-tumor activity of folate-mitomycin conjugates.

PURPOSE: Folate receptor (FR) targeted drug conjugates were prepared by covalently attaching the vitamin folate, to the potent anticancer drug, mitomycin C (MMC). One such conjugate, called EC72, was synthesized with an intramolecular disulfide bond, and it was found to exhibit efficacious anti-tumor activity against FR-expressing M109 tumors in a manner that yielded no gross or microscopic toxicity, even to FR-positive kidneys. METHODS: EC72's specificity was demonstrated by two methods: (1) blocking EC72's activity with an excess of co-administered folic acid (FA) in M109 tumor bearing mice and (2) the absence of therapeutic activity in mice bearing FR-negative tumors. The importance of having a cleavable bond in the conjugate was also exemplified, since EC110 (a folate-MMC conjugate constructed with a more resilient amide bond) failed to produce anti-M109 tumor activity. EC72's therapeutic potential was found to decrease with respect to the increasing size of subcutaneous tumor. However, a combination therapy with paclitaxel reproducibly improved the anti-tumor efficacy relative to either agent alone at well tolerated dose levels and with no apparent increase in toxicity. A more advanced folate-MMC conjugate was also synthesized in an effort to improve activity. Thus, EC118, a molecule constructed with both a reducible disulfide bond and an acid-labile hydrazone bond in the linker region, was tested and found to produce a significantly greater number of tumor regressions of more established M109 tumors than that achieved with EC72. CONCLUSION: Overall, these data indicate that folate-targeted drug therapy alone, or in combination with paclitaxel, may be a novel and effective clinical approach towards treating FR-positive cancers.

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PubMed2004

Ligand binding and kinetics of folate receptor recycling in vivo: impact on receptor-mediated drug delivery.

Folate receptor-targeted cancer therapies constitute a promising treatment for the approximately one third of human cancers that overexpress the folate receptor (FR). However, the potencies of all folate-receptor targeted therapies depend on 1) the rate of folate-linked drug conjugate binding to the cancer cell surface, 2) the dose of folate conjugate that will saturate tumor cell surface FR in vivo, 3) the rate of FR internalization, unloading, and recycling back to the tumor cell surface for another round of conjugate uptake, and 4) the residence time of the folate conjugate before its metabolism or release from the cell. Because little information exists on any of these processes, we have undertaken to characterize them on both cancer cells in culture and solid tumors in live mice. We quantitate here the properties of FR saturation, internalization, recycling, and unloading in several cultured cancer cell lines and murine tumor models, and we describe the conditions that should maximize both the potencies and specificities of folate receptor-targeted therapies in vivo.

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PubMed2004

Folate-targeted chemotherapy.

The folate receptor (FR) is a confirmed tumor-associated antigen that binds folate and folate-drug conjugates with very high affinity and shuttles these bound molecules inside cells via an endocytic mechanism. Using folate (or an analog thereof) as the ligand, a wide variety of drug payloads can be delivered to FR-positive cells, ranging from small radioactive imaging agents up to large DNA-containing formulations. For therapeutic purposes, attachment of small molecular weight, highly potent agents to folate is a novel approach. As such, this review will outline the manner in which folate-drug conjugates are screened preclinically, it will summarize published activity data of various conjugates, and it will also highlight some newly emerging animal data from novel compounds that are currently under preclinical investigation.

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PubMed2004

Preclinical evaluation of (99m)Tc-EC20 for imaging folate receptor-positive tumors.

UNLABELLED: Our group previously reported on the synthesis and characterization of a novel (99m)Tc-based folate-peptide chelator called EC20. This agent was found to bind folate receptor (FR)-positive cells and tissues with high affinity and was deemed useful for radiodiagnostic applications. In this study, we investigated the effect of D-amino acid substitution within EC20 on its tissue biodistribution. Expanded in vivo studies were also performed with unmodified (99m)Tc-EC20 to determine the effect of tumor FR expression, tumor size, tumor location, route of dose administration, and rodent diet on the agent's tissue biodistribution pattern. METHODS: EC20 and EC53, the all-D-isomer of EC20, were synthesized and radiolabeled with (99m)Tc. The relative affinity of EC53 to the FR with respect to EC20 was then determined in cultured tumor cells. The ability of (99m)Tc-EC20 and (99m)Tc-EC53 to target tumors in vivo was examined using BALB/c mice with subcutaneously inoculated M109 or 4T1 cells, yielding 0.1- to 0.5-g tumors in 20 d. RESULTS: The D-amino acid substitutions of EC20 were found to reduce the uptake of the agent into tumor and major organs. Subsequent studies using the original (99m)Tc-EC20 agent confirmed that its net tumor uptake was specific and proportional to FR expression levels in tumor cells as well as linear with respect to the overall tumor size. Further, (99m)Tc-EC20 uptake was found to be independent of both solid tumor location (intraperitoneal vs. subcutaneous) and the route of administration (intraperitoneal vs. intravenous). Interestingly, leucovorin supplementation of a commonly used folate-deficient laboratory chow had no effect on the agent's overall tissue biodistribution pattern. But, tumor-to-nontumor ratios could be increased up to 2.7-fold when 1 equivalent of free folic acid was coinjected with (99m)Tc-EC20. CONCLUSION: Taken together, these results confirm that (99m)Tc-EC20 has the potential to be a clinically useful noninvasive radiodiagnostic agent for detecting the locus of FR-positive cancers.

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PubMed2002

Characterization of a novel pH-sensitive peptide that enhances drug release from folate-targeted liposomes at endosomal pHs.

Although liposomes have proven useful for the delivery of drugs and gene therapy vectors, their potencies are often compromised by poor unloading following uptake into their target cells. We have consequently explored the properties of a novel 29-residue amphipathic peptide that was designed by arrangement of hydrophobic and hydrophilic residues to disrupt liposomes at lower peptide concentrations than previously tested peptides. The peptide was indeed found to promote pH-dependent liposome unloading with improved efficiency. A peptide of the same sequence, but half the length, however, promoted pH-dependent permeabilization only at much higher concentrations. Further characterization of the longer peptide revealed that release of liposome contents (i) occurred at a pH of approximately 6, (ii) became less efficient as the size of the encapsulated cargo increased, and (iii) was moderately suppressed in cholesterol-containing liposomes. Use of this peptide to enhance the cytotoxicity of cytosine arabinoside encapsulated in folate-targeted liposomes demonstrated an increase in drug potency of approximately 30-fold. Gene expression by a serum-stable folate-targeted liposomal vector was also measurably enhanced by inclusion of the peptide. We conclude that intracellular unloading of liposomal contents can be significantly improved by co-encapsulation of an optimally designed, pH-sensitive peptide.

Amino Acid Sequence↗

PubMed2002

Folate-targeted gene transfer in vivo.

Nonviral systemic delivery is one of the most attractive approaches for cancer gene therapy. To achieve this goal, various laboratories have developed cationic liposomes. However, when injected intravenously, cationic lipid-DNA complexes accumulate mostly into and transfect lung tissue. Here, we describe a method by which these complexes can be targeted to tumors using folic acid. Adding polyethylene glycol (PEG)-lipids to the complexes dramatically reduced both lung accumulation and gene transfer to lungs and tumors after intravenous administration. The presence of folic acid at the distal end of the PEG-lipid did not modify tumor accumulation of the complexes. However, with folate-targeted complexes, gene transfer activity was restored in tumors while the activity in lungs was reduced by 50- to 100-fold compared with nontargeted lipid-DNA complexes. This approach provides a first in vivo proof of concept to achieve targeted tumor gene delivery.

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PubMedDate not supplied

Synthesis and biological evaluation of EC20: a new folate-derived, (99m)Tc-based radiopharmaceutical.

A new peptide derivative of folic acid was designed to efficiently coordinate (99m)Tc. This new chelate, referred to as EC20, was found to bind cultured folate receptor (FR)-positive tumor cells in both a time- and concentration-dependent manner with very high affinity (K(D) approximately 3 nM). Using an in vitro relative affinity assay, EC20 was also found to effectively compete with (3)H-folic acid for cell binding when presented either alone or as a formulated metal chelate. Following intravenous injection into Balb/c mice, (99m)Tc-EC20 was rapidly removed from circulation (plasma t(1/2) approximately 4 min) and excreted into the urine in a nonmetabolized form. Data from gamma scintigraphic and quantitative biodistribution studies performed in M109 tumor-bearing Balb/c mice confirmed that (99m)Tc-EC20 predominantly accumulates in FR-positive tumor and kidney tissues. These results suggest that (99m)Tc-EC20 may be clinically useful as a noninvasive radiodiagnostic imaging agent for the detection of FR-positive human cancers.

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PubMedDate not supplied

Synthesis and biological evaluation of EC72: a new folate-targeted chemotherapeutic.

A novel folate conjugate of mitomycin C, herein referred to as EC72, was designed and evaluated for biological activity against FR-positive cells and tumors. EC72 was produced by coupling folic acid-gamma-cysteine to 7-N-modified MMC via a disulfide bond. This water soluble conjugate was found to retain high affinity for FR-positive cells, and it produced dose responsive activity in vitro against a panel of folate receptor (FR)-positive cell lines. EC72's activity was considered to be targeted and specific for the FR since (i) excess folic acid blocked biological activity, and (ii) FR-negative cell lines were unresponsive to this drug. Initial in vivo tests confirmed EC72's activity in both syngeneic and xenograft models, and this activity occurred in the apparent absence of gross or pathological toxicity. These results are significant, since daily dosing of EC72 for more than 30 consecutive days yielded no evidence of myelosuppression or toxicity to major organs, including the FR-positive kidneys. The latter observation supports published data, indicating that the apically oriented kidney proximal tubule FRs function to salvage folates prior to their excretion and to return these molecules back into systemic circulation. Overall, EC72's performance in vitro and in vivo warrants further preclinical study before this novel targeted chemotherapeutic is considered for clinical investigation.

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PubMedDate not supplied

Synthesis and biological evaluation of EC140: a novel folate-targeted vinca alkaloid conjugate.

A novel folate conjugate of desacetylvinblastine monohydrazide (DAVLBH), herein referred to as EC140, was designed and evaluated for biological activity against folate receptor (FR)-positive cells and tumors. EC140 was produced by coupling a peptidic analogue of the vitamin folic acid to DAVLBH via an acylhydrazone bond. This water-soluble conjugate was found to retain high affinity for FR-positive cells, and it produced specific, dose-responsive activity in vitro. Initial in vivo tests confirmed EC140's activity in both syngeneic and xenograft models. Hence, enduring complete responses were observed in animals bearing established, subcutaneous tumors prior to therapy using regimens that produced minor toxicity. In contrast, treatment with the unconjugated DAVLBH drug produced nominal efficacy when dosed at its MTD. Overall, EC140's performance in vitro and in vivo warrants further preclinical study before this novel targeted chemotherapeutic is considered for clinical investigation.

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