RESUMO
CD33 (Siglec-3) is a cell surface receptor expressed in approximately 90% of acute myeloid leukemia (AML) blasts, making it an attractive target for therapy of AML. Although previous CD33-targeting antibody-drug conjugates (ADC) like gemtuzumab ozogamicin (GO, Mylotarg) have shown efficacy in AML treatment, they have suffered from toxicity and narrow therapeutic window. This study aimed to develop a novelADCwith improved tolerability and a wider therapeutic window. GLK-33 consists of the anti-CD33 antibody lintuzumab and eight mavg-MMAU auristatin linkerpayloads per antibody. The experimental methods included testing in cell cultures, patient-derived samples, mouse xenograft models, and rat toxicology studies. GLK-33 exhibited remarkable efficacy in reducing cell viability within CD33-positive leukemia cell lines and primary AML samples. Notably, GLK-33 demonstrated antitumor activity at single dose as low as 300 mg/kg in mice, while maintaining tolerability at single dose of 20 to 30 mg/kg in rats. In contrast with both GO and lintuzumab vedotin, GLK-33 exhibited a wide therapeutic window and activity against multidrug-resistant cells. The development of GLK-33 addresses the limitations of previous ADCs, offering a wider therapeutic window, improved tolerability, and activity against drug-resistant leukemia cells. These findings encourage further exploration of GLK-33 in AML through clinical trials.
Assuntos
Anticorpos Monoclonais Humanizados , Imunoconjugados , Leucemia Mieloide Aguda , Oligopeptídeos , Lectina 3 Semelhante a Ig de Ligação ao Ácido Siálico , Humanos , Animais , Leucemia Mieloide Aguda/tratamento farmacológico , Leucemia Mieloide Aguda/patologia , Camundongos , Lectina 3 Semelhante a Ig de Ligação ao Ácido Siálico/antagonistas & inibidores , Lectina 3 Semelhante a Ig de Ligação ao Ácido Siálico/metabolismo , Ratos , Anticorpos Monoclonais Humanizados/farmacologia , Oligopeptídeos/farmacologia , Imunoconjugados/farmacologia , Imunoconjugados/uso terapêutico , Aminobenzoatos/farmacologia , Ensaios Antitumorais Modelo de Xenoenxerto , Linhagem Celular Tumoral , FemininoRESUMO
PURPOSE: Antibody-drug conjugates (ADCs) have shown impressive clinical activity with approval of many agents in hematological and solid tumors. However, challenges remain with both efficacy and safety of ADCs. This study describes novel trastuzumab-auristatin conjugates with the hydrophilic MMAE prodrug MMAU, and optimization of a glycopeptide linker leading to a wider therapeutic window. EXPERIMENTAL DESIGN: Trastuzumab was conjugated with auristatin payloads via a series of linkers using a stabilized maleimide handle. The ADCs were characterized in vitro and their relative in vivo anti-tumor efficacies were assessed in HER2+ xenograft models. Relative linker stabilities and the mechanism of linker cleavage were studied using in vitro assays. Toxicity and toxicokinetics of the best performing ADC were evaluated in cynomolgus monkey (cyno). RESULTS: The trastuzumab-MMAU ADC with stabilized glycopeptide linker showed maleimide stabilization and higher resistance to cleavage by serum and lysosomal enzymes compared to a valine-citrulline conjugated trastuzumab ADC (trastuzumab-vc-MMAE). A single dose of 1 or 2 mg/kg of trastuzumab-MMAU at drug-to-antibody ratios (DAR) of 8 and 4 respectively resulted in xenograft tumor growth inhibition, with superior efficacy to trastuzumab-vc-MMAE. Trastuzumab-MMAU DAR4 was tolerated at doses up to 12 mg/kg in cyno, which represents 2- to 4-fold higher dose than that observed with vedotin ADCs, and had increased terminal half-life and exposure. CONCLUSIONS: The optimized trastuzumab-MMAU ADC showed potent antitumor activity and was well tolerated with excellent pharmacokinetics in non-human primates, leading to a superior preclinical therapeutic window. The data supports potential utility of trastuzumab-MMAU for treatment of HER2+ tumors.
RESUMO
Commensal gut microbiota and probiotics have numerous effects on the host's metabolic and protective systems, which occur primarily through the intestinal epithelial cell interface. Prebiotics, like galacto-oligosaccharides (GOS) are widely used to modulate their function and abundance. However, important structure-function relations may exist, requiring a detailed structural characterization. Here, we detailed the structural characterization of bovine whey derived oligosaccharide preparations enriched with GOS or not, dubbed GOS-enriched milk oligosaccharides (GMOS) or MOS, respectively. We explore GMOS's and MOS's potential to improve intestinal epithelial barrier function, assessed in a model based on barrier disruptive effects of the Clostridioides difficile toxin A. GMOS and MOS contain mainly GOS species composed of ß1-6- and ß1-3-linked galactoses, and 3'- and 6'-sialyllactose. Both GMOS and MOS, combined with lactobacilli, like Lactobacillus rhamnosus (LPR, NCC4007), gave synergistic epithelial barrier protection, while no such effect was observed with Bifidobacterium longum (BL NCC3001), Escherichia coli (Nissle) or fructo-oligosaccharides. Mechanistically, for barrier protection with MOS, (i) viable LPR was required, (ii) acidification of growth medium was not enough, (iii) LPR did not directly neutralize toxin A, and (iv) physical proximity of LPR with the intestinal epithelial cells was necessary. This is the first study, highlighting the importance of structure-function specificity and the necessity of the simultaneous presence of prebiotic, probiotic and host cell interactions required for a biological effect.
Assuntos
Microbioma Gastrointestinal , Mucosa Intestinal , Oligossacarídeos , Simbióticos , Soro do Leite , Animais , Toxinas Bacterianas/efeitos adversos , Bovinos , Linhagem Celular Tumoral , Enterotoxinas/efeitos adversos , Galactose/química , Galactose/metabolismo , Microbioma Gastrointestinal/efeitos dos fármacos , Microbioma Gastrointestinal/fisiologia , Humanos , Mucosa Intestinal/efeitos dos fármacos , Mucosa Intestinal/metabolismo , Lactobacillus/metabolismo , Oligossacarídeos/química , Oligossacarídeos/metabolismo , Oligossacarídeos/farmacologia , Prebióticos , Probióticos/farmacologia , Substâncias Protetoras/química , Substâncias Protetoras/metabolismo , Substâncias Protetoras/farmacologiaRESUMO
The GalNAcbeta1-4GlcNAc determinant (LdN) occurs in some human and bovine glycoconjugates and also in lower vertebrates and invertebrates. It has been found in unsubstituted as well as terminally substituted forms at the distal end of conjugated glycans, but it has not been reported previously at truly internal positions of polylactosamine chains. Here, we describe enzyme-assisted conversion of LdNbeta1-OR oligosaccharides into GlcNAcbeta1-3GalNAcbeta1-4GlcNAcbeta1-OR. The extension reactions, catalyzed by human serum, were modeled after analogous beta3-GlcNAc transfer processes that generate GlcNAcbeta1-3Galbeta1-4GlcNAcbeta1-OR. The newly synthesized GlcNAcbeta1-3GalNAc linkages were unambiguously identified by nuclear magnetic resonance data, including the appropriate long-range correlations in heteronuclear multiple bond correlation spectra. The novel GlcNAcbeta1-3'LdN determinant proved to be a functional acceptor for several mammalian glycosyltransferases, suggesting that human polylactosamines may contain internal LdN units in many distinct forms. The GlcNAcbeta1-3'LdN determinant was unusually resistant toward jackbean beta-N-acetylhexosaminidase; the slow degradation should lead to a convenient method for the search of putative internal LdN determinants in natural polylactosamine chains.