Antibody-Drug Conjugates: Modes of Toxicity Nonclinical safety evaluation of immunoconjuates Melissa M. Schutten, DVM, PhD, DACVP Safety Assessment Pathology Melissa M Schutten, D NorCal Society of Toxicology Meeting September 27, 2012 Overview • Introduction to Antibody Drug Conjugates (ADCs) • Modes of ADC toxicity • Challenges associated with nonclinical safety evaluation of ADCs • Summary 2 Anatomy of an Antibody-Drug Conjugate (ADC) Linker stable in circulation Antibody targeted to tumor • Humanized monoclonal Ab (IgG1) • mAb with Fc modifications (modulate ADCC, CDC activity) • Other mAb fragments • Linker biochemistry • Acid labile (hydrazone) • Enzyme dipeptides (cleavable) • Thioether (uncleavable) • Hindered disulfide (uncleavable) • Site of conjugation • Fc, HC, LC Very potent chemotherapeutic drug • Tubulin polymerization inhibitors • Maytansines (DM1, DM4) • Auristatins (MMAE, MMAF) • DNA damaging agents • Calicheamicins • Duocarmycins • Anthracyclines (doxorubicin) Genentech Confidential—Internal Use Only 3 Improving the Therapeutic Window • ADCs can selectively deliver a potent cytotoxic drug to tumor cells via tumorspecific and/or over-expressed antigens • Increase drug delivery to tumor • Reduce normal tissue drug exposure Chemotherapy ADC DRUG DOSE TOXIC DOSE (MTD) TOXIC DOSE (MTD) Therapeutic Window Therapeutic Window EFFICACIOUS DOSE (MED) EFFICACIOUS DOSE (MED) MTD: Maximum tolerated dose; MED: Minimum Efficacious Dose 4 ADC More Efficacious than Free Cytotoxin in Mice MMTV-HER2 Fo5 mammary tumor (HER2-positive, trastuzumab-insensitive) 1500 Vehicle Mean Tumor Volume (mm3 ) +/- SEM DM1 Trastuzumab-mertansine 15 mg/kg, 817 µg/m 2 Trastuzumab 15 mg/kg Trastuzumab 15 mg/kg + Free DM1 817 µg/m2 1000 Free DM1 817 µg/m2 Free DM1 (near MTD) 1947 µg/m2 500 Free DM1 (cytotoxin) T-DM1 (ADC) 0 0 IV Dosing 5 10 15 20 25 30 Day Parsons et al, AACR (2007); Modified from S. Spencer ADC Better Tolerated than Free Cytotoxin in Rats Single IV dose; rats Body Weight (% change from baseline) T-DM1 (2040 µg DM1/m²) Free DM1 (2400 µg DM1/m²) Early mortality (100%) Time (Day) T-DM1: Trastuzumab emtansine 6 ADC Better Tolerated than Free Cytotoxin in Monkeys White Blood Cells 6 mg/kg ADC (~750 μg MMAE/m2) 0.063 mg/kg MMAE (~750 μg MMAE/m2) • No neutrophil decreases when cytotoxic drug delivered linked to an antibody • ~2-3 times more cytotoxic drug can be given as an ADC A. Kim, D. Danilenko, N. Dybdal, K. Flagella, K. Achilles-Poon Modes of Anti-tumor Activity of ADCs Tumor Cell Tumor cytotoxicity is target-directed ADC-Ag binding → internalization in lysosomes → ADC degradation → release of toxin intracellularly → tumor cell death Genentech Confidential—Internal Use Only Tumor Cells Tumor cytotoxicity is target-enhanced (bystander effect) ADC-Ag binding → extracellular cleavage of toxin → release of toxin in local tumor environment → diffusion of toxin intracellularly to neighboring tumor cells → tumor cell death 8 Tissue Antigen Characteristics Are Key in ADCs Careful selection of target antigens are an important criterion for both the safety and efficacy of an ADC •The ‘ideal’ tissue antigen should have: – – – – – High level of target expression in cancer cells Little to no expression in normal cells Expressed on the cell surface Readily internalized No shedding into the blood by cleavage of the antigen from cancer cell surface •The number of antigen molecules and antibody binding affinity for the antigen may affect the potency of the ADC 9 Modes of Toxicity of ADCs + Normal Cell Systemic release of toxin Unwanted ADC-mediated cytotoxicity • Instability of linker • Catabolism of ADC • Targeted binding to normal tissues expressing antigen • Off-target (cross reactive) binding to normal tissues • Non-antigen-mediated ADC uptake (e.g., Fcmediated uptake, pinocytosis) Genentech Confidential—Internal Use Only 10 Modes of Toxicity of ADCs + Normal Cell Systemic release of toxin Unwanted ADC-mediated cytotoxicity • Instability of linker • Catabolism of ADC • Targeted binding to normal tissues expressing antigen • Off-target (cross reactive) binding to normal tissues • Non-antigen-mediated ADC uptake (e.g., Fcmediated uptake, pinocytosis) Genentech Confidential—Internal Use Only 11 Slower Drug Deconjugation With Uncleavable Linker Concentration (µg/ml) Single IV dose 20 mg/kg ADC Total Ab Uncleavable linker Cleavable linker Days post dose Polson, et al., Cancer Res., 69(6), 2009 12 More Stable Linker Reduces Systemic Toxicity of ADC in Rats Change in bodyweight (grams) Single IV dose given on Day 1 : CD22-DM1 with cleavable linker Days post dose Polson, et al., Cancer Res., 69(6), 2009 13 More Stable Linker Reduces Systemic Toxicity of ADC in Rats Single IV dose given on Day 1 : Polson, et al., Cancer Res., 69(6), 2009 14 Modes of Toxicity of ADCs + Normal Cell Systemic release of toxin Unwanted ADC-mediated cytotoxicity • Instability of linker • Catabolism of ADC • DAR • Targeted binding to normal tissues expressing antigen • Off-target (cross reactive) binding to normal tissues • Non-antigen-mediated ADC uptake (e.g., Fcmediated uptake, pinocytosis) Genentech Confidential—Internal Use Only 15 Early Observation: Highly Drugged ADCs More Toxic DAR: Drug-to-Antibody Ratio 1500 1300 AST (U/L) 1100 900 DAR 6 Study Day 0 Study Day 3 Study Day 5 700 DAR 4 DAR 4 DAR 2 500 DAR 2 DAR 6 300 100 -100 Vehicle Control 840 ug/m2 840 ug/m2 840 ug/m2 Herceptin-MC-vc- Herceptin-MC-vc- Herceptin-MC-vcPAB-MMAF (2P) PAB-MMAF (4P) PAB-MMAF (6P) 16 ADC Proportion ThioMAb Technology: Controlling Heterogeneity TDC Proportion DAR DAR •Engineered ThioMAb backbone allows more homogenous drug load (MMAE) •Efficacy of TDC ADC (mg/kg basis) and 2 x ADC (ug MMAE/m2 basis) Junutula, et al., Nat. Biotech., 26(8), 2008 17 Catabolism and Deconjugation of TDC is Slower than ADC in Rats Single dose I.V. PK study: ADC or TDC with matched cytotoxin (MMAE) doses Catabolism of the Antibody Junutula, et al., Nat. Biotech., 26(8), 2008 Deconjugation of the Antibody MMAE TDC is Better Tolerated Than ADC in Monkeys Repeat IV doses of ADC or TDC, Days 1 and 23: • No neutrophil decreases with TDC compared to equivalent ug/m2 dose of ADC Junutula, et al., Nat. Biotech., 26(8), 2008 19 Modes of Toxicity of ADCs + Normal Cell Systemic release of toxin Unwanted ADC-mediated cytotoxicity • Instability of linker • Catabolism of ADC • DAR • Site of conjugation • Targeted binding to normal tissues expressing antigen • Off-target (cross reactive) binding to normal tissues • Non-antigen-mediated ADC uptake (e.g., Fcmediated uptake, pinocytosis) Genentech Confidential—Internal Use Only 20 Modes of Toxicity of ADCs + Normal Cell Systemic release of toxin Unwanted ADC-mediated cytotoxicity • Instability of linker • Catabolism of ADC • Targeted binding to normal tissues expressing antigen • Off-target (cross reactive) binding to normal tissues • Non-antigen-mediated ADC uptake (e.g., Fcmediated uptake, pinocytosis) 21 Target Antigen Binding Causes “On-Target” Lymphoid Depletion B-cell target depletion in splenic follicles: An example of “exaggerated pharmacology” Vehicle Anti-cyCD79b CD20 Ki-67 Polson, et. al, Mol Cancer Ther 8(10), 2009 Anti-cyCD79b MCC DM1 Target Toxicity to Normal Tissues Results: Seven patients received a total of 23 weekly doses of bivatuzumab mertansine. One patient at the 100 mg/m2 and one at the 120 mg/m2 level experienced stable disease during treatment phase but also developed grade 1 skin toxicity (desquamation). One of them received a second treatment course. At the highest dose level achieved in this study (140 mg/m2), one patient developed toxic epidermal necrolysis after two infusions and died. Massive apoptosis of skin keratinocytes had occurred, whereas only symptomatic therapy for skin toxicity was available. The risk-benefit assessment of all patients treated in the total phase I program (4 clinical trials, 70 patients) turned out to be negative after consideration of this case of a toxic epidermal necrolysis and the skin-related adverse events observed in the other trials. Therefore, development of the conjugate was Modes of Toxicity of ADCs + Normal Cell Systemic release of toxin Unwanted ADC-mediated cytotoxicity • Instability of linker • Catabolism of ADC • Targeted binding to normal tissues expressing antigen • Off-target (cross reactive) binding to normal tissues • Non-antigen-mediated ADC uptake (e.g., Fc-mediated uptake, pinocytosis) 24 Summary • An ADC is both a “large molecule” and a “small molecule”. • ADCs hold great promise for improving current oncology therapies. – Highly potent cytotoxic agents are delivered directly to cancer cells, sparing normal tissues. – ADCs tend to be better tolerated than standard chemotherapy. – Increased therapeutic window allows for better balance between safety/efficacy. • There is a fine balance between efficacy and toxicity. ─ Choice of linker, cytotoxic drug and mAb are all important determinants of safety, PK, and efficacy. ─ Toxicity is usually antigen-independent, ADC/drug-dependent. ─ Linker stability, DAR, and site of drug conjugation impacts toxicity. 25 Acknowledgements • • • • • • • • • • • • • • • Reina Fuji Kelly Flagella Willy Solis Kirsten Achilles-Poon Jacqueline Tarrant Rama Pai Ning Ma Joe Beyer Trung Nguyen Nghi La Fiona Zhong Michelle McDowell Noel Dybdal Donna Dambach Theresa Reynolds • • • • • • • • • • • Angela Hendricks Amy Oldendorp Surinder Kaur Ben Shen Jay Tibbitts Joo-Hee Yi Kedan Lin Doug Leipold Ola Saad Montserrat Carrasco-Triguero Keyang Xu • • • • Luna Liu Andy Boswell Helen Davis Margaret Kenrick • • • • • • • • • • • • • • Susan Spencer Paul Polakis Bonnee Rubinfeld Jagath Junutula Shang-Fan Yu David Kan Ivan Inigo Wai Lee Wong Kathy Kozak Elaine Mai Jeff Gorrell Michael Mamounas Andrew Polson Seattle Genetics 26