Roger Zabinski - Academia.edu (original) (raw)

Papers by Roger Zabinski

ABSTRACT: Kinetic and product analyses of the reactions of dialkylglycine decarboxylase with seve... more ABSTRACT: Kinetic and product analyses of the reactions of dialkylglycine decarboxylase with several alternative substrates are presented. Rate constants for the reactions of amino and keto acids of several substrates decrease logarithmically with increasing side-chain size. Conversely, kcat for L-amino acid decarboxylation increases with side-chain size. These and other data confirm a proposed model for three binding subsites in the active site. In this model, bond making and breaking in both the decarboxylation and transamination half-reactions occurs at the “A ” subsite, which maintains the scissile bond aligned with the p orbitals of the conjugated aldimine and thus maximizes stereoelectronic effects. This strongly supports the proposal by Dunathan (Proc. Natl. Acad. Sci. U.S.A. 55, 712-716) that PLP-dependent enzymes can largely control reaction specificity by specific orientation about CR in the external aldimine intermediate. The “B ” subsite can accept either an alkyl or a c...

Cancer Research, 2007

AACR Annual Meeting-- Apr 14-18, 2007; Los Angeles, CA 4086 Antibody-drug conjugates (ADCs) incor... more AACR Annual Meeting-- Apr 14-18, 2007; Los Angeles, CA 4086 Antibody-drug conjugates (ADCs) incorporating protease sensitive linkages release drugs through the action of proteases after being taken into cells. In target cells ADC uptake is antigen mediated, and drug release occurs in lysosomes. To help define the time and location of drug release a FRET reagent was designed and synthesized. The FRET reagent comprises a fluorophore (Alexa Fluor 488) and a quencher (DABMI) connected through the same linkage by which drugs are attached to antibodies in ADCs. When the antibody-FRET conjugate (AFC) encounters proteolytic activity capable of releasing drug from an ADC, the linkage between fluorophore and quencher is broken revealing a fluorescent signal. Characterization of AFCs prepared with the FRET reagent indicates that the fluorescence is very efficiently quenched in the native form, as the fluorescent signal increases approximately 250 fold when the attachment to the quencher is rup...

Journal of Biological …, 2006

The chimeric anti-CD30 monoclonal antibody cAC10, linked to the antimitotic agents monomethyl aur... more The chimeric anti-CD30 monoclonal antibody cAC10, linked to the antimitotic agents monomethyl auristatin E (MMAE) or F (MMAF), produces potent and highly CD30-selective anti-tumor activity in vitro and in vivo. These drugs are appended via a valine-citrulline (vc) dipeptide ...

Autoantibodies that react with double-stranded DNA (dsDNA) are a hallmark for diagnosis of system... more Autoantibodies that react with double-stranded DNA (dsDNA) are a hallmark for diagnosis of systemic lupus erythematosus (SLE) and are also considered the pathogenic subset that is most associated with lupus nephritis. As an agent to remove the pathogenic dsDNA antibodies from the circulation of SLE patients, we are developing an antigen-based heteropolymer (AHP). The AHP consists of a monoclonal antibody to the complement receptor (CR1) cross-linked to salmon testis dsDNA to effect clearance of anti-DNA antibodies by binding them to erythrocyte CR1. Utilizing a cynomolgus monkey model for SLE in which we infused plasma from SLE patients containing a high titer of high-avidity anti-dsDNA antibody, we have evaluated the safety and efficacy of AHP infusion. The results demonstrate that AHP rapidly (within 2 min of infusion) binds to monkey erythrocytes without causing any toxicological effects. We also demonstrate that human Ig (G؉M) antibodies are rapidly bound to the AHP-erythrocyte complex. These events are mirrored in their kinetics by a substantial drop in the level of high-avidity dsDNA antibody in the plasma.

Tetrahedron Letters, 1993

... As for 7, the 1Sconfiguration obtained for 10 was established by difference nOe experiments .... more ... As for 7, the 1Sconfiguration obtained for 10 was established by difference nOe experiments ... BAH thanks the American Cancer Society for Postdoctoral Fellowship PF3298, and the Albert Einstein College of Medicine for a Milder and Emil Holland Postdoctoral Scholarship Award ...

Molecular Cancer Therapeutics, 2007

B-cell maturation antigen (BCMA) is expressed on normal and malignant plasma cells and represents... more B-cell maturation antigen (BCMA) is expressed on normal and malignant plasma cells and represents a potential target for therapeutic intervention. BCMA binds to two ligands that promote tumor cell survival, a proliferation inducing ligand (APRIL) and B-cell activating factor. To selectively target BCMA for plasma cell malignancies, we developed antibodies with ligand blocking activity that could promote cytotoxicity of multiple myeloma (MM) cell lines as naked antibodies or as antibody-drug conjugates. We show that SG1, an inhibitory BCMA antibody, blocks APRIL-dependent activation of nuclear factor-KB in a dose-dependent manner in vitro. Cytotoxicity of SG1 was assessed as a naked antibody after chimerization with and without Fc mutations that enhance Fc;RIIIA binding. The Fc mutations increased the antibody-dependent cellmediated cytotoxicity potency of BCMA antibodies against MM lines by f100-fold with a z2-fold increase in maximal lysis. As an alternative therapeutic strategy, anti-BCMA antibodies were endowed with direct cytotoxic activity by conjugation to the cytotoxic drug, monomethyl auristatin F. The most potent BCMA antibody-drug conjugate displayed IC 50 values of V130 pmol/L for three different MM lines. Hence, BCMA antibodies show cytotoxic activity both as naked IgG and as drug conjugates and warrant further evaluation as therapeutic candidates for plasma cell malignancies.

Journal of the American Chemical Society, 1997

Journal of the American Chemical Society, 2001

Nonenzymatic pyridoxal phosphate (PLP) catalyzed decarboxylations and transaminations have been r... more Nonenzymatic pyridoxal phosphate (PLP) catalyzed decarboxylations and transaminations have been revisited experimentally. Metal ions are known to catalyze a variety of PLP-dependent reactions in solution, including transamination. It is demonstrated here that the rate accelerations previously observed are due solely to enhancement of Schiff base formation under subsaturating conditions. A variety of metal ions were tested for their effects on the reactivity of the 2-methyl-2-aminomalonate Schiff bases. All were found to have either no effect or a small inhibitory one. The effects of Al(3+) were studied in detail with the Schiff bases of 2-methyl-2-aminomalonate, 2-aminoisobutyrate, alanine, and ethylamine. The decarboxylation of 2-methyl-2-aminomalonate is unaffected by metalation with Al(3+), while the decarboxylation of 2-aminoisobutyrate is inhibited 125-fold. The transamination reaction of ethylamine is 75-fold slower than that of alanine. Ethylamine transamination is inhibited 4-fold by Al(3+) metalation, while alanine transamination is inhibited only 1.3-fold. Metal ion inhibition of Schiff base reactivity suggests a simple explanation for the lack of known PLP dependent enzymes that make direct mechanistic use of metal ions. A comparison of enzyme catalyzed, PLP catalyzed, and uncatalyzed reactions shows that PLP dependent decarboxylases are among the best known biological rate enhancers: decarboxylation occurs 10(18)-fold faster on the enzyme surface than it does free in solution. PLP itself provides the lion's share of the catalytic efficiency of the holoenzyme: at pH 8, free PLP catalyzes 2-aminoisobutyrate decarboxylation by approximately 10(10)-fold, with the enzyme contributing an additional approximately 10(8)-fold.

Clinical Cancer Research, 2004

Purpose: An antibody-drug conjugate consisting of monomethyl auristatin E (MMAE) conjugated to th... more Purpose: An antibody-drug conjugate consisting of monomethyl auristatin E (MMAE) conjugated to the anti-CD30 monoclonal antibody (mAb) cAC10, with eight drug moieties per mAb, was previously shown to have potent cytotoxic activity against CD30 ؉ malignant cells. To determine the effect of drug loading on antibody-drug conjugate therapeutic potential, we assessed cAC10 antibody-drug conjugates containing different drug-mAb ratios in vitro and in vivo. Experimental Design: Coupling MMAE to the cysteines that comprise the interchain disulfides of cAC10 created an antibody-drug conjugate population, which was purified using hydrophobic interaction chromatography to yield antibody-drug conjugates with two, four, and eight drugs per antibody (E2, E4, and E8, respectively). Antibody-drug conjugate potency was tested in vitro against CD30 ؉ lines followed by in vivo xenograft models. The maximum-tolerated dose and pharmacokinetic profiles of the antibody-drug conjugates were investigated in mice. Results: Although antibody-drug conjugate potency in vitro was directly dependent on drug loading (IC 50 values E8<E4<E2), the in vivo antitumor activity of E4 was comparable with E8 at equal mAb doses, although the E4 contained half the amount of MMAE per mAb. E2 was also an active antitumor agent but required higher doses. The maximum-tolerated dose of E2 in mice was at least double that of E4, which in turn was twice that of E8. MMAE loading affected plasma clearance, as E8 cleared 3-fold faster than E4 and 5-fold faster than E2. Conclusions: By decreasing drug loading per antibody, the therapeutic index was increased demonstrating that drug loading is a key design parameter for antibody-drug conjugates.

Biochemistry, 1998

Kinetic and product analyses of the reactions of dialkylglycine decarboxylase with several altern... more Kinetic and product analyses of the reactions of dialkylglycine decarboxylase with several alternative substrates are presented. Rate constants for the reactions of amino and keto acids of several substrates decrease logarithmically with increasing side-chain size. Conversely, k cat for L-amino acid decarboxylation increases with side-chain size. These and other data confirm a proposed model for three binding subsites in the active site. In this model, bond making and breaking in both the decarboxylation and transamination half-reactions occurs at the "A" subsite, which maintains the scissile bond aligned with the p orbitals of the conjugated aldimine and thus maximizes stereoelectronic effects. This strongly supports the proposal by Dunathan (Proc. Natl. Acad. Sci. U.S.A. 55, 712-716) that PLP-dependent enzymes can largely control reaction specificity by specific orientation about C R in the external aldimine intermediate. The "B" subsite can accept either an alkyl or a carboxylate group, while the "C" subsite accepts only small alkyl groups. This model predicts the existence of nonproductive binding modes for amino acids, which is proposed to be the ultimate origin of the k cat increase with side-chain size for L-amino acid decarboxylation. The specificity of the 2-aminoisobutyrate decarboxylation half-reaction toward oxidative decarboxylation is very high (<1 in 10 5 turnovers yields nonoxidative decarboxylation). The origin of this specificity is explored with the reactions of amino-and methylaminomalonate. These substrates exhibit high yields of nonoxidative decarboxylation, providing support for a model in which the interaction between a carboxylate group in the B subsite and Arg406 is a prerequisite to proton donation to and removal from C R .

British Journal of Cancer, 2008

CD133/prominin-1 is a pentaspan transmembrane glycoprotein overexpressed in various solid tumours... more CD133/prominin-1 is a pentaspan transmembrane glycoprotein overexpressed in various solid tumours including colorectal and glioblastomas. CD133 was found here to be highly expressed in X50% of pancreatic, gastric and intrahepatic cholangiocarcinomas. Quantitative flow cytometric analysis showed that a panel of established hepatocellular, pancreatic and gastric cancer cell lines expressed CD133 at levels higher than normal epithelial cells or bone marrow progenitor cells. A murine anti-human CD133 antibody (AC133) conjugated to a potent cytotoxic drug, monomethyl auristatin F (MMAF), effectively inhibited the growth of Hep3B hepatocellular and KATO III gastric cancer cells in vitro with IC 50 values of 2-7 ng ml À1. MMAF induced apoptosis in the cancer cells as measured by caspase activation. The anti-CD133-drug conjugate (AC133-vcMMAF) was shown to internalise and colocalised with the lysosomal marker CD107a in the sensitive cell lines. In contrast, in the resistant cell line Su.86.86, the conjugate internalised and colocalised with the caveolae marker, Cav-1. Addition of ammonium chloride, an inhibitor of lysosomal trafficking and processing, suppressed the cytotoxic effect of AC133-vcMMAF in both Hep3B and KATO III. Anti-CD133-drug conjugate treatment resulted in significant delay of Hep3B tumour growth in SCID mice. Anti-CD133 antibody-drug conjugates warrant further evaluation as a therapeutic strategy to eradicate CD133 þ tumours.

Induction of apoptosis in endothelial cells is considered an attractive strategy to therapeutical... more Induction of apoptosis in endothelial cells is considered an attractive strategy to therapeutically interfere with a solid tumor&amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;#39;s blood supply. In the present paper, we constructed cytotoxic conjugates that specifically target angiogenic endothelial cells, thus preventing typical side effects of apoptosis-inducing drugs. For this purpose, we conjugated the potent antimitotic agent monomethyl-auristatin-E (MMAE) via a lysosomal cleavable linker to human serum albumin (HSA) and further equipped this drug-albumin conjugate with cyclic c(RGDfK) peptides for multivalent interaction with alphavbeta3-integrin. The RGD-peptides were conjugated via either an extended poly(ethylene glycol) linker or a short alkyl linker. The resulting drug-targeting conjugates RGDPEG-MMAE-HSA and RGD-MMAE-HSA demonstrated high binding affinity and specificity for alphavbeta3-integrin expressing human umbilical vein endothelial cells (HUVEC). Both types of conjugates were internalized by endothelial cells and killed the target cells at low nM concentrations. Furthermore, we observed RGD-dependent binding of the conjugates to C26 carcinoma. Upon i.v. administration to C26-tumor bearing mice, both drug-targeting conjugates displayed excellent tumor homing properties. Our results demonstrate that RGD-modified albumins are suitable carriers for cell selective intracellular delivery of cytotoxic compounds, and further studies will be conducted to assess the antivascular and tumor inhibitory potential of RGDPEG-MMAE-HSA and RGD-MMAE-HSA.

Molecular Pharmaceutics, 2007