Publications by authors named "Kedan Lin"

21 Publications

  • Page 1 of 1

Biodistribution and efficacy of an anti-TENB2 antibody-drug conjugate in a patient-derived model of prostate cancer.

Oncotarget 2019 Oct 22;10(58):6234-6244. Epub 2019 Oct 22.

Genentech Research and Early Development, South San Francisco, 94080 CA, USA.

TENB2, a transmembrane proteoglycan protein, is a promising target for antibody drug conjugate (ADC) therapy due to overexpression in human prostate tumors and rapid internalization. We previously characterized how predosing with parental anti-TENB2 monoclonal antibody (mAb) at 1 mg/kg in a patient-derived LuCap77 explant model with high (3+) TENB2 expression could (i) block target-mediated intestinal uptake of tracer (& 0.1 mg/kg) levels of radiolabeled anti-TENB2-monomethyl auristatin E ADC while preserving tumor uptake, and (ii) maintain efficacy relative to ADC alone. Here, we systematically revisit this strategy to evaluate the effects of predosing on tumor uptake and efficacy in LuCap96.1, a low TENB2-expressing (1+) patient-derived model that is more responsive to ADC therapy than LuCap77. Importantly, rather than using tracer (& 0.1 mg/kg) levels, radiolabeled ADC tumor uptake was assessed at 1 mg/kg - one of the doses evaluated in the tumor growth inhibition study - in an effort to bridge tissue distribution (PK) with efficacy (PD). Predosing with mAb up to 1 mg/kg had no effect on efficacy. These findings warrant further investigations to determine whether predosing prior to ADC therapy might improve therapeutic index by preventing ADC disposition and possible toxicological liabilities in antigen-expressing healthy tissues.
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http://dx.doi.org/10.18632/oncotarget.27263DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6817444PMC
October 2019

Phase Ia Study of Anti-NaPi2b Antibody-Drug Conjugate Lifastuzumab Vedotin DNIB0600A in Patients with Non-Small Cell Lung Cancer and Platinum-Resistant Ovarian Cancer.

Clin Cancer Res 2020 01 20;26(2):364-372. Epub 2019 Sep 20.

Sarah Cannon Research Institute, Nashville, Tennessee.

Purpose: This phase I trial assessed the safety, tolerability, and preliminary antitumor activity of lifastuzumab vedotin (LIFA), an antibody-drug conjugate of anti-NaPi2b mAb (MNIB2126A) and a potent antimitotic agent (monomethyl auristatin E).

Patients And Methods: LIFA was administered to patients with non-small cell lung cancer (NSCLC) and platinum-resistant ovarian cancer (PROC), once every 3 weeks, by intravenous infusion. The starting dose was 0.2 mg/kg in this 3+3 dose-escalation design, followed by cohort expansion at the recommended phase II dose (RP2D).

Results: Overall, 87 patients were treated at doses between 0.2 and 2.8 mg/kg. The MTD was not reached; 2.4 mg/kg once every 3 weeks was selected as the RP2D based on overall tolerability profile. The most common adverse events of any grade and regardless of relationship to study drug were fatigue (59%), nausea (49%), decreased appetite (37%), vomiting (32%), and peripheral sensory neuropathy (29%). Most common treatment-related grade ≥3 toxicities among patients treated at the RP2D ( = 63) were neutropenia (10%), anemia (3%), and pneumonia (3%). The pharmacokinetic profile was dose proportional. At active doses ≥1.8 mg/kg, partial responses were observed in four of 51 (8%) patients with NSCLC and 11 of 24 (46%) patients with PROC per RECIST. All RECIST responses occurred in patients with NaPi2b-high by IHC. The CA-125 biomarker assessed for patients with PROC dosed at ≥1.8 mg/kg showed 13 of 24 (54%) had responses (≥50% decline from baseline).

Conclusions: LIFA exhibited dose-proportional pharmacokinetics and an acceptable safety profile, with encouraging activity in patients with PROC at the single-agent RP2D of 2.4 mg/kg.
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http://dx.doi.org/10.1158/1078-0432.CCR-18-3965DOI Listing
January 2020

Single cell-produced and in vitro-assembled anti-FcRH5/CD3 T-cell dependent bispecific antibodies have similar in vitro and in vivo properties.

MAbs 2019 Feb/Mar;11(2):422-433. Epub 2018 Dec 17.

a Genentech Research and Early Development, Genentech, Inc ., South San Francisco , CA , USA.

Bispecific antibody production using single host cells has been a new advancement in the antibody engineering field. We previously showed comparable in vitro biological activity and in vivo mouse pharmacokinetics (PK) for two novel single cell variants (v10 and v11) and one traditional dual cell in vitro-assembled anti-human epidermal growth factor receptor 2/CD3 T-cell dependent bispecific (TDB) antibodies. Here, we extended our previous work to assess single cell-produced bispecific variants of a novel TDB against FcRH5, a B-cell lineage marker expressed on multiple myeloma (MM) tumor cells. An in vitro-assembled anti- FcRH5/CD3 TDB antibody was previously developed as a potential treatment option for MM. Two bispecific antibody variants (designs v10 and v11) for manufacturing anti-FcRH5/CD3 TDB in single cells were compared to in vitro-assembled TDB in a dual-cell process to understand whether differences in antibody design and production led to any major differences in their in vitro biological activity, in vivo mouse PK, and PK/pharmacodynamics (PD) or immunogenicity in cynomolgus monkeys (cynos). The binding, in vitro potencies, in vitro pharmacological activities and in vivo PK in mice and cynos of these single cell TDBs were comparable to those of the in vitro-assembled TDB. In addition, the single cell and in vitro-assembled TDBs exhibited robust PD activity and comparable immunogenicity in cynos. Overall, these studies demonstrate that single cell-produced and in vitro-assembled anti-FcRH5/CD3 T-cell dependent bispecific antibodies have similar in vitro and in vivo properties, and support further development of single-cell production method for anti-FcRH5/CD3 TDBs and other single-cell bispecifics.
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http://dx.doi.org/10.1080/19420862.2018.1551676DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6380433PMC
July 2019

Tutorial on Monoclonal Antibody Pharmacokinetics and Its Considerations in Early Development.

Clin Transl Sci 2018 11 7;11(6):540-552. Epub 2018 Aug 7.

Clinical Pharmacology, NGM Biopharmaceuticals, Inc., South San Francisco, California, USA.

The tutorial introduces the readers to the fundamentals of antibody pharmacokinetics (PK) in the context of drug development. Topics covered include an overview of antibody development, PK characteristics, and the application of antibody PK/pharmacodynamics (PD) in research and development decision-making. We also discuss the general considerations for planning a nonclinical PK program and describe the types of PK studies that should be performed during early development of monoclonal antibodies.
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http://dx.doi.org/10.1111/cts.12567DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6226118PMC
November 2018

Relative Target Affinities of T-Cell-Dependent Bispecific Antibodies Determine Biodistribution in a Solid Tumor Mouse Model.

Mol Cancer Ther 2018 04 16;17(4):776-785. Epub 2018 Jan 16.

Genentech, Inc., South San Francisco, California.

Anti-HER2/CD3, a T-cell-dependent bispecific antibody (TDB) construct, induces T-cell-mediated cell death in cancer cells expressing HER2 by cross-linking tumor HER2 with CD3 on cytotoxic T cells, thereby creating a functional cytolytic synapse. TDB design is a very challenging process that requires consideration of multiple parameters. Although therapeutic antibody design strategy is commonly driven by striving for the highest attainable antigen-binding affinity, little is known about how the affinity of each TDB arm can affect the targeting ability of the other arm and the consequent distribution and efficacy. To our knowledge, no distribution studies have been published using preclinical models wherein the T-cell-targeting arm of the TDB is actively bound to T cells. We used a combined approach involving radiochemistry, invasive biodistribution, and noninvasive single-photon emission tomographic (SPECT) imaging to measure TDB distribution and catabolism in transgenic mice with human CD3ε expression on T cells. Using CD3 affinity variants, we assessed the impact of CD3 affinity on short-term pharmacokinetics, tissue distribution, and cellular uptake. Our experimental approach determined the relative effects of (i) CD3 targeting to normal tissues, (ii) HER2 targeting to HER2-expressing tumors, and (iii) relative HER2/CD3 affinity, all as critical drivers for TDB distribution. We observed a strong correlation between CD3 affinity and distribution to T-cell-rich tissues, with higher CD3 affinity reducing systemic exposure and shifting TDB distribution away from tumor to T-cell-containing tissues. These observations have important implications for clinical translation of bispecific antibodies for cancer immunotherapy. .
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http://dx.doi.org/10.1158/1535-7163.MCT-17-0657DOI Listing
April 2018

Efficient production of bispecific IgG of different isotypes and species of origin in single mammalian cells.

MAbs 2017 Feb/Mar;9(2):213-230. Epub 2016 Dec 8.

a Department of Antibody Engineering , Genentech, Inc. , South San Francisco , CA , USA.

Bispecific IgG production in single host cells has been a much sought-after goal to support the clinical development of these complex molecules. Current routes to single cell production of bispecific IgG include engineering heavy chains for heterodimerization and redesign of Fab arms for selective pairing of cognate heavy and light chains. Here, we describe novel designs to facilitate selective Fab arm assembly in conjunction with previously described knobs-into-holes mutations for preferential heavy chain heterodimerization. The top Fab designs for selective pairing, namely variants v10 and v11, support near quantitative assembly of bispecific IgG in single cells for multiple different antibody pairs as judged by high-resolution mass spectrometry. Single-cell and in vitro-assembled bispecific IgG have comparable physical, in vitro biological and in vivo pharmacokinetics properties. Efficient single-cell production of bispecific IgG was demonstrated for human IgG, IgG and IgG thereby allowing the heavy chain isotype to be tailored for specific therapeutic applications. Additionally, a reverse chimeric bispecific IgG with humanized variable domains and mouse constant domains was generated for preclinical proof-of-concept studies in mice. Efficient production of a bispecific IgG in stably transfected mammalian (CHO) cells was shown. Individual clones with stable titer and bispecific IgG composition for >120 days were readily identified. Such long-term cell line stability is needed for commercial manufacture of bispecific IgG. The single-cell bispecific IgG designs developed here may be broadly applicable to biotechnology research, including screening bispecific IgG panels, and to support clinical development.
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http://dx.doi.org/10.1080/19420862.2016.1267089DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5297516PMC
November 2017

An anti-CD3/anti-CLL-1 bispecific antibody for the treatment of acute myeloid leukemia.

Blood 2017 02 1;129(5):609-618. Epub 2016 Dec 1.

Research and Early Development, Genentech, South San Francisco, CA.

Acute myeloid leukemia (AML) is a major unmet medical need. Most patients have poor long-term survival, and treatment has not significantly changed in 40 years. Recently, bispecific antibodies that redirect the cytotoxic activity of effector T cells by binding to CD3, the signaling component of the T-cell receptor, and a tumor target have shown clinical activity. Notably, blinatumomab is approved to treat relapsed/refractory acute lymphoid leukemia. Here we describe the design, discovery, pharmacologic activity, pharmacokinetics, and safety of a CD3 T cell-dependent bispecific (TDB) full-length human IgG1 therapeutic antibody targeting CLL-1 that could potentially be used in humans to treat AML. CLL-1 is prevalent in AML and, unlike other targets such as CD33 and CD123, is not expressed on hematopoietic stem cells providing potential hematopoietic recovery. We selected a high-affinity monkey cross-reactive anti-CLL-1 arm and tested several anti-CD3 arms that varied in affinity, and determined that the high-affinity CD3 arms were up to 100-fold more potent in vitro. However, in mouse models, the efficacy differences were less pronounced, probably because of prolonged exposure to TDB found with lower-affinity CD3 TDBs. In monkeys, assessment of safety and target cell depletion by the high- and low-affinity TDBs revealed that only the low-affinity CD3/CLL1 TDB was well tolerated and able to deplete target cells. Our data suggest that an appropriately engineered CLL-1 TDB could be effective in the treatment of AML.
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http://dx.doi.org/10.1182/blood-2016-08-735365DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5290988PMC
February 2017

Development and Translational Application of an Integrated, Mechanistic Model of Antibody-Drug Conjugate Pharmacokinetics.

AAPS J 2017 01 27;19(1):130-140. Epub 2016 Sep 27.

Genentech Research and Early Development, 1 DNA Way, South San Francisco, California, 94080, USA.

Antibody drug conjugates (ADC), in which small molecule cytotoxic agents are non-specifically linked to antibodies, can enable targeted delivery of chemotherapeutics to tumor cells. ADCs are often produced and administered as a mixture of conjugated antibodies with different drug to antibody ratios (DAR) resulting in complex and heterogeneous disposition kinetics. We developed a mechanism-based platform model that can describe and predict the complex pharmacokinetic (PK) behavior of ADCs with protease-cleavable valine-citrulline (VC) linker linked to Monomethylmonomethyl auristatin F/E by incorporating known mechanisms of ADC disposition. The model includes explicit representation of all DAR species; DAR-dependent sequential deconjugation of the drug, resulting in the conversion of higher DAR to lower DAR species; and DAR-dependent antibody/ADC clearance. PK profiles of multiple analytes (total antibody, drug-conjugated antibody, and/or antibody-conjugated drug) for different ADC molecules and targets in rodents and cynomolgus monkeys were used for model development. The integrated cross-species model was successful in capturing the multi-analyte PK profiles after administration of purified ADCs with defined DAR species and ADCs with mixtures of DAR. Human PK predictions for DSTP3086S (anti-STEAP1-vc-MMAE) with the platform model agreed well with PK (total antibody and antibody-conjugated drug concentrations) measurements in the dose-ranging phase I clinical study. The integrated model is applicable to various other ADCs with different formats, conjugated drugs, and linkers, and provides a valuable tool for the exploration of mechanisms governing disposition of ADCs and enables translational predictions.
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http://dx.doi.org/10.1208/s12248-016-9993-zDOI Listing
January 2017

Preclinical Development of an Anti-NaPi2b (SLC34A2) Antibody-Drug Conjugate as a Therapeutic for Non-Small Cell Lung and Ovarian Cancers.

Clin Cancer Res 2015 Nov 8;21(22):5139-50. Epub 2015 Jul 8.

Genentech Research and Early Development, South San Francisco, California.

Purpose: Antibody-drug conjugates (ADC) selectively deliver a cytotoxic drug to cells expressing an accessible antigenic target. Here, we have appended monomethyl auristatin E (MMAE) to an antibody recognizing the SLC34A2 gene product NaPi2b, the type II sodium-phosphate cotransporter, which is highly expressed on tumor surfaces of the lung, ovary, and thyroid as well as on normal lung pneumocytes. This study evaluated its efficacy and safety in preclinical studies.

Experimental Design: The efficacy of anti-NaPi2b ADC was evaluated in mouse ovarian and non-small cell lung cancer (NSCLC) tumor xenograft models, and its toxicity was assessed in rats and cynomolgus monkeys.

Results: We show here that an anti-NaPi2b ADC is effective in mouse ovarian and NSCLC tumor xenograft models and well-tolerated in rats and cynomolgus monkeys at levels in excess of therapeutic doses. Despite high levels of expression in normal lung of non-human primate, the cross-reactive ADC exhibited an acceptable safety profile with a dose-limiting toxicity unrelated to normal tissue target expression. The nonproliferative nature of normal pneumocytes, together with the antiproliferative mechanism of MMAE, likely mitigates the potential liability of this normal tissue expression.

Conclusions: Overall, our preclinical results suggest that the ADC targeting NaPi2b provides an effective new therapy for the treatment of NSCLC and ovarian cancer and is currently undergoing clinical developments.
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http://dx.doi.org/10.1158/1078-0432.CCR-14-3383DOI Listing
November 2015

Mechanism-Based Pharmacokinetic/Pharmacodynamic Model for THIOMAB™ Drug Conjugates.

Pharm Res 2015 Jun 2;32(6):1884-93. Epub 2014 Dec 2.

Genentech Research and Early Development, 1 DNA Way, South San Francisco, California, 94080, USA.

Purpose: THIOMAB™ drug conjugates (TDCs) with engineered cysteine residues allow site-specific drug conjugation and defined Drug-to-Antibody Ratios (DAR). In order to help elucidate the impact of drug-loading, conjugation site, and subsequent deconjugation on pharmacokinetics and efficacy, we have developed an integrated mathematical model to mechanistically characterize pharmacokinetic behavior and preclinical efficacy of MMAE conjugated TDCs with different DARs. General applicability of the model structure was evaluated with two different TDCs.

Method: Pharmacokinetics studies were conducted for unconjugated antibody and purified TDCs with DAR-1, 2 and 4 for trastuzumab TDC and Anti-STEAP1 TDC in mice. Total antibody concentrations and individual DAR fractions were measured. Efficacy studies were performed in tumor-bearing mice.

Results: An integrated model consisting of distinct DAR species (DAR0-4), each described by a two-compartment model was able to capture the experimental data well. Time series measurements of each Individual DAR species allowed for the incorporation of site-specific drug loss through deconjugation and the results suggest a higher deconjugation rate from heavy chain site HC-A114C than the light chain site LC-V205C. Total antibody concentrations showed multi-exponential decline, with a higher clearance associated with higher DAR species. The experimentally observed effects of TDC on tumor growth kinetics were successfully described by linking pharmacokinetic profiles to DAR-dependent killing of tumor cells.

Conclusion: Results from the integrated model evaluated with two different TDCs highlight the impact of DAR and site of conjugation on pharmacokinetics and efficacy. The model can be used to guide future drug optimization and in-vivo studies.
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http://dx.doi.org/10.1007/s11095-014-1582-1DOI Listing
June 2015

Conjugates of Small Molecule Drugs with Antibodies and Other Proteins.

Biomedicines 2014 Jan 24;2(1):1-13. Epub 2014 Jan 24.

Protein Interactions Group, Cancer and Inflammation Program, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Frederick, MD 21702, USA.

Conjugates of small molecule drugs with antibodies (ADCs) and with other proteins (protein-drug conjugates, PDC) are used as a new class of targeted therapeutics combining the specificity of monoclonal antibodies (mAbs) and other proteins with potent cytotoxic activity of small molecule drugs for the treatment of cancer and other diseases. A(P)DCs have three major components, antibody (targeting protein), linker and payload, the cytotoxic drug. Recently, advances in identifying targets, selecting highly specific mAbs of preferred isotypes, optimizing linker technology and improving chemical methods for conjugation have led to the approval of two ADCs by Food and Drug Administration (FDA) and more than 30 ADCs in advanced clinical development. However, the complex and heterogeneous nature of A(P)DCs often cause poor solubility, instability, aggregation and eventually unwanted toxicity. This article reviews the main components of A(P)DCs, and discusses the choices for drugs, linkers and conjugation methods currently used. Future work will need to focus on developments and strategies for overcoming such major problems associated with the A(P)DCs.
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http://dx.doi.org/10.3390/biomedicines2010001DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5423484PMC
January 2014

Pharmacokinetics and ADME characterizations of antibody-drug conjugates.

Methods Mol Biol 2013 ;1045:117-31

Early Development Pharmacokinetics and Pharmacodynamics, Genentech Inc., South San Francisco, CA, USA.

Pharmacokinetic and absorption, distribution, metabolism, and excretion (ADME) characterization of antibody-drug conjugates (ADCs) reflects the dynamic interactions between the biological system and ADC, and provides critical assessments in lead selection, optimization, and clinical development. Understanding the pharmacokinetics (PK), ADME properties and consequently the pharmacokinetic-pharmacodynamic properties of ADCs is critical for their successful development. This chapter discusses the PK properties of ADCs, types of PK and ADME studies in supporting different stages of development, general design of PK/ADME studies with a focus on ADC-specific characteristics, and interpretation of PK parameters.
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http://dx.doi.org/10.1007/978-1-62703-541-5_7DOI Listing
February 2014

Characterization of the drug-to-antibody ratio distribution for antibody-drug conjugates in plasma/serum.

Bioanalysis 2013 May;5(9):1057-71

Genentech, 1 DNA Way, South San Francisco, CA 94080, USA.

Background: Antibody-drug conjugates (ADCs) are a new class of cancer therapeutics that deliver potent cytotoxins specifically to tumors to minimize systemic toxicity. However, undesirable release of covalently linked drugs in circulation can affect safety and efficacy. The objective of this manuscript was to propose and assess the assays that allow for the characterization of the drug deconjugation in plasma/serum.

Results: ADCs of three main drug conjugation platforms, linked via lysine, site-specific engineered cysteine or reduced interchain disulfide cysteine residues, were analyzed using affinity capture for sample enrichment coupled with LC-MS or hydrophobic interaction chromatography-UV for detection. These novel approaches enabled measurement of the relative abundance of individual ADC species with different drug-to-antibody ratios, while maintaining their structural integrity.

Conclusion: The characterization data generated by affinity capture LC-MS or hydrophobic interaction chromatography-UV provided critical mechanistic insights into understanding the stability and bioactivity of ADCs in vivo, and also helped the development of appropriate quantitative ELISAs.
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http://dx.doi.org/10.4155/bio.13.66DOI Listing
May 2013

Total antibody quantification for MMAE-conjugated antibody-drug conjugates: impact of assay format and reagents.

Bioconjug Chem 2013 May 11;24(5):772-9. Epub 2013 Apr 11.

Department of Biochemical and Cellular Pharmacology, Genentech, Inc., South San Francisco, CA 94080, USA.

Antibody-drug conjugates (ADCs) are target-specific anticancer agents consisting of cytotoxic drugs covalently linked to a monoclonal antibody. The number of ADCs in the clinic is growing, and therefore thorough characterization of the quantitative assays used to measure ADC concentrations in support of pharmacokinetic, efficacy, and safety studies is of increasing importance. Cytotoxic drugs such as the tubulin polymerization inhibiting auristatin, monomethyl auristatin E, have been conjugated to antibodies via cleavable linkers (MC-vc-PAB) through internal cysteines. This results in a heterogeneous mixture of antibody species with drug-to-antibody ratios (DAR) ranging from 0 to 8. In order to characterize the assays used to quantitate total MC-vc-PAB-MMAE ADCs (conjugated and unconjugated antibody), we used purified fractions with defined DARs from 6 therapeutic antibodies to evaluate different assay formats and reagents. Our investigations revealed that for quantitation of total antibody, including all unconjugated and conjugated antibody species, sandwich ELISA formats did not always allow for recovery of all purified DAR fractions (DAR 0-8) to within ±20% of the expected values at the reagent concentrations tested. In evaluating alternative approaches, we found that the recovery of DAR fractions with semihomogeneous assay (SHA) formats, in which sample, capture, and detection reagents are preincubated in solution, were less affected by the antibody's MMAE drug load as compared to traditional stepwise sandwich ELISAs. Thus, choosing the optimal assay format and reagents for total antibody assays is valuable for developing accurate quantitative assays.
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http://dx.doi.org/10.1021/bc300491kDOI Listing
May 2013

Micro-volume wall-less immunoassays using patterned planar plates.

Lab Chip 2013 Apr;13(7):1342-50

Department of Biochemical and Cellular Pharmacology, Genentech, Inc., South San Francisco, CA, USA.

Miniaturization of immunoassays has numerous potential advantages over traditional ELISAs. Here we present a novel approach using patterned planar plates (PPPs). These 'wall-less' plates consist of a 16 × 24 array of 2 mm diameter hydrophilic regions surrounded by a hydrophobic polytetrafluoroethylene (PTFE) coating. Assays are performed by adding 2 μL droplets to the hydrophilic areas. These droplets are overlaid with an immiscible mixture of perfluorocarbon liquid (PFCL) that essentially eliminates evaporation. During wash steps, a thin film of PFCL covers the hydrophobic coating and prevents its wetting by wash buffer; as a result, the hydrophilic wells remain intact and inter-well cross-contamination is prevented. We compared the performance of three immunoassays using PPPs versus traditional 384-well ELISA plates. These included assays for soluble FcRH5 in human serum, SDF-1 in mouse serum, and human IgG in mouse plasma. The results show that the PPP assays were closely comparable to the ELISAs in terms of sensitivity, linearity of dilution, and sample quantitation. Moreover, the PPP assays were rapid to perform, easily adapted from ELISA protocols, and used 10- to 50-fold less sample and reagent volume as compared to 384- or 96-well plate ELISAs. As an additional advantage, PPPs conform to established microplate dimensional standards making them compatible with pre-existing equipment and workflows. PPPs therefore represent an attractive and broadly applicable approach to flexible miniaturization of plate-based immunochemical assays.
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http://dx.doi.org/10.1039/c3lc40973bDOI Listing
April 2013

Fourth World Antibody-Drug Conjugate Summit: February 29-March 1, 2012, Frankfurt, Germany.

MAbs 2012 Nov-Dec;4(6):637-47. Epub 2012 Aug 22.

Centre d'Immunologie Pierre Fabre, Saint-Julien-en-Genevois, France.

The 4th World Antibody Drug Conjugate (WADC) Summit, organized by Hanson Wade was held on February 29‑March 1, 2012 in Frankfurt, Germany, which was also the location for the Antibody Drug Conjugate Summit Europe held in February 2011. During the one year between these meetings, antibody drug conjugates (ADCs) have confirmed their technological maturity and their clinical efficacy in oncology. Brentuximab vedotin (ADCETRIS (TM) ) gained approval by the US Food and Drug Administration in August 2011 and trastuzumab emtansine (T-DM1) confirmed impressive clinical efficacy responses in a large cohort of breast cancer patients. During the 4th WADC meeting, antibody-maytansinoid conjugates were showcased by representatives of ImmunoGen (T-DM1, SAR3419, lorvotuzumab mertansine/IMGN801, IMGN529 and IMG853) and Biotest (BT-062). Data on antibody-auristatin conjugates were presented by scientists and clinicians from Seattle Genetics and Takeda (brentuximab vedotin), Pfizer (5T4-MMAF), Agensys/Astella (AGS-16M8F), Progenics (PSMA-ADC) and Genmab (anti-TF ADCs). Alternative payloads such as calicheamicins and duocarmycin used for preparation of ADCs were discussed by Pfizer and Synthon representatives, respectively. In addition, emerging technologies, including site-directed conjugation (Ambrx), a protein toxin as payload (Viventia), hapten-binding bispecific antibodies (Roche), and use of light activated drugs (Photobiotics), were also presented. Last but not least, progresses in solving Chemistry Manufacturing and Control, and pharmacokinetic issues were addressed by scientists from Genentech, Pfizer, Novartis and Pierre Fabre.
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http://dx.doi.org/10.4161/mabs.21697DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3502230PMC
January 2014

An integrated approach to identify normal tissue expression of targets for antibody-drug conjugates: case study of TENB2.

Br J Pharmacol 2013 Jan;168(2):445-57

Genentech Research and Early Development, South San Francisco, CA 94080, USA.

Background And Purpose: The success of antibody-drug conjugates (ADCs) depends on the therapeutic window rendered by the differential expression between normal and pathological tissues. The ability to identify and visualize target expression in normal tissues could reveal causes for target-mediated clearance observed in pharmacokinetic characterization. TENB2 is a prostate cancer target associated with the progression of poorly differentiated and androgen-independent tumour types, and ADCs specific for TENB2 are candidate therapeutics. The objective of this study was to locate antigen expression of TENB2 in normal tissues, thereby elucidating the underlying causes of target-mediated clearance.

Experimental Approach: A series of pharmacokinetics, tissue distribution and mass balance studies were conducted in mice using a radiolabelled anti-TENB2 ADC. These data were complemented by non-invasive single photon emission computed tomography - X-ray computed tomography imaging and immunohistochemistry.

Key Results: The intestines were identified as a saturable and specific antigen sink that contributes, at least in part, to the rapid target-mediated clearance of the anti-TENB2 antibody and its drug conjugate in rodents. As a proof of concept, we also demonstrated the selective disposition of the ADC in a tumoural environment in vivo using the LuCaP 77 transplant mouse model. High tumour uptake was observed despite the presence of the antigen sink, and antigen specificity was confirmed by antigen blockade.

Conclusions And Implications: Our findings provide the anatomical location and biological interpretation of target-mediated clearance of anti-TENB2 antibodies and corresponding drug conjugates. Further investigations may be beneficial in addressing the relative contributions to ADC disposition from antigen expression in both normal and pathological tissues.
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http://dx.doi.org/10.1111/j.1476-5381.2012.02138.xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3572570PMC
January 2013

Differential effects of predosing on tumor and tissue uptake of an 111In-labeled anti-TENB2 antibody-drug conjugate.

J Nucl Med 2012 Sep 7;53(9):1454-61. Epub 2012 Aug 7.

Genentech Research and Early Development, South San Francisco, California 94080, USA.

Unlabelled: TENB2, also known as tomoregulin or transmembrane protein with epidermal growth factor-like and 2 follistatin-like domains, is a transmembrane proteoglycan overexpressed in human prostate tumors. This protein is a promising target for antimitotic monomethyl auristatin E (MMAE)-based antibody-drug conjugate (ADC) therapy. Nonlinear pharmacokinetics in normal mice suggested that antigen expression in normal tissues may contribute to targeted mediated disposition. We evaluated a predosing strategy with unconjugated antibody to block ADC uptake in target-expressing tissues in a mouse model while striving to preserve tumor uptake and efficacy.

Methods: Unconjugated, unlabeled antibody was preadministered to mice bearing the TENB2-expressing human prostate explant model, LuCaP 77, followed by a single administration of (111)In-labeled anti-TENB2-MMAE for biodistribution and SPECT/CT studies. A tumor-growth-inhibition study was conducted to determine the pharmacodynamic consequences of predosing.

Results: Preadministration of anti-TENB2 at 1 mg/kg significantly increased blood exposure of the radiolabeled ADC and reduced intestinal, hepatic, and splenic uptake while not affecting tumor accretion. Similar tumor-to-heart ratios were measured by SPECT/CT at 24 h with and without the predose. Consistent with this, the preadministration of 0.75 mg/kg did not interfere with efficacy in a tumor-growth study dosed at 0.75 mg or 2.5 mg of ADC per kilogram.

Conclusion: Overall, the potential to mask peripheral, nontumor antigen uptake while preserving tumor uptake and efficacy could ameliorate toxicity and may significantly affect future dosing strategies for ADCs.
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http://dx.doi.org/10.2967/jnumed.112.103168DOI Listing
September 2012

Pharmacokinetic considerations for antibody drug conjugates.

Pharm Res 2012 Sep 28;29(9):2354-66. Epub 2012 Jun 28.

Genentech Research and Early Development, South San Francisco, California 94080, USA.

Antibody drug conjugates (ADCs) are a class of therapeutics that combine the target specificity of an antibody with the potency of a chemotherapeutic. This therapeutic strategy can significantly expand the therapeutic index of a chemotherapeutic by minimizing the systemic exposure and associated toxicity of the chemotherapeutic agent, while simultaneously maximizing the delivery of the chemotherapeutic to the target. The abundance of antibody targets, coupled with advances in antibody engineering, conjugation chemistry, and examples of early clinical success, have stimulated interest in developing ADCs. However, developing and optimizing the highly complex components of ADCs remain challenging. Understanding the pharmacokinetics (PK) and consequently the pharmacokinetic-pharmacodynamic (PKPD) properties of ADCs is critical for their successful development. This review discusses the PK properties of ADCs, with a focus on ADC-specific characteristics, including molecular heterogeneity, in vivo processing, and the implications of multiple analytes. The disposition of ADCs and the utility of PKPD modeling are discussed in the context of providing guidance to assist in the successful development of these complex molecules.
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http://dx.doi.org/10.1007/s11095-012-0800-yDOI Listing
September 2012

Impact of drug conjugation on pharmacokinetics and tissue distribution of anti-STEAP1 antibody-drug conjugates in rats.

Bioconjug Chem 2011 Oct 3;22(10):1994-2004. Epub 2011 Oct 3.

Department of Pharmacokinetic & Pharmacodynamic Sciences, Genentech Research & Early Development, South San Francisco, California 94080, United States.

Antibody-drug conjugates (ADCs) are designed to combine the exquisite specificity of antibodies to target tumor antigens with the cytotoxic potency of chemotherapeutic drugs. In addition to the general chemical stability of the linker, a thorough understanding of the relationship between ADC composition and biological disposition is necessary to ensure that the therapeutic window is not compromised by altered pharmacokinetics (PK), tissue distribution, and/or potential organ toxicity. The six-transmembrane epithelial antigen of prostate 1 (STEAP1) is being pursued as a tumor antigen target. To assess the role of ADC composition in PK, we evaluated plasma and tissue PK profiles in rats, following a single dose, of a humanized anti-STEAP1 IgG1 antibody, a thio-anti-STEAP1 (ThioMab) variant, and two corresponding thioether-linked monomethylauristatin E (MMAE) drug conjugates modified through interchain disulfide cysteine residues (ADC) and engineered cysteines (TDC), respectively. Plasma PK of total antibody measured by enzyme-linked immunosorbent assay (ELISA) revealed ∼45% faster clearance for the ADC relative to the parent antibody, but no apparent difference in clearance between the TDC and unconjugated parent ThioMab. Total antibody clearances of the two unconjugated antibodies were similar, suggesting minimal effects on PK from cysteine mutation. An ELISA specific for MMAE-conjugated antibody indicated that the ADC cleared more rapidly than the TDC, but total antibody ELISA showed comparable clearance for the two drug conjugates. Furthermore, consistent with relative drug load, the ADC had a greater magnitude of drug deconjugation than the TDC in terms of free plasma MMAE levels. Antibody conjugation had a noticeable, albeit minor, impact on tissue distribution with a general trend toward increased hepatic uptake and reduced levels in other highly vascularized organs. Liver uptakes of ADC and TDC at 5 days postinjection were 2-fold and 1.3-fold higher, respectively, relative to the unmodified antibodies. Taken together, these results indicate that the degree of overall structural modification in anti-STEAP1-MMAE conjugates has a corresponding level of impact on both PK and tissue distribution.
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http://dx.doi.org/10.1021/bc200212aDOI Listing
October 2011

Serum response factor activation by muscarinic receptors via RhoA. Novel pathway specific to M1 subtype involving calmodulin, calcineurin, and Pyk2.

J Biol Chem 2002 Oct 27;277(43):40789-98. Epub 2002 Aug 27.

Departments of Biopharmaceutical Sciences and Pharmaceutical Chemistry, University of California, San Francisco, CA 94143-0446, USA.

The muscarinic cholinergic receptor (mAChR) subtypes share high sequence similarity except in their third intracellular loop and COOH terminus, domains thought to be involved in signal transduction. Subtypes M1, M3, and M5 couple mainly through Galpha(q/11), and M2 and M4 couple mainly through Galpha(i/o). Whether subtypes within each of these two groups differ in their signaling pathways remains to be resolved. This study focused on nuclear signaling pathways leading to activation of the transcription factor, serum response factor (SRF). Genes encoding M1, M2, and M3 were co-expressed in Jurkat T lymphocytes with a reporter gene driven by a mutant serum response element, SRE.L, which responds to SRF activation. We show that only M1 mAChR activated SRF through a pathway involving the small GTPase RhoA, with no response observed for M2 and M3. Transfection of GTPase-deficient Galpha subunits (GalphaQL; constitutively active form) demonstrated that SRF was activated by Galpha(13)QL but only marginally by Galpha(q)QL and Galpha(12)QL in Jurkat cells. Yet transfection of regulator of G protein-signaling protein, RGS2 and RGS4, which inhibit Galpha(q/11) activity, indicated that Galpha(q/11) and Ca(2+) mobilization were required for SRF activation by M1. Calmodulin inhibitors suppressed the M1 and the Galpha(13)QL pathways, acting both upstream and downstream of RhoA. However, calcineurin inhibitors and the tyrosine kinase inhibitor genistein selectively suppressed SRF activation by M1, but not by Galpha(13)QL, indicating the presence of separate pathways. The calmodulin-dependent tyrosine kinase Pyk2 was also activated by M1 but not M3, and Pyk2 appears also to play a role in M1-SRF activation, as judged by experiments with two dominant-negative Pyk2 mutants. These results reveal a novel calmodulin-dependent RhoA-SRF signaling pathway unique to the M1 mAChR subtype.
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http://dx.doi.org/10.1074/jbc.M202745200DOI Listing
October 2002