Discovery of STRO-002, a Novel Homogeneous ADC Targeting Folate Receptor Alpha, for the Treatment of Ovarian and Endometrial Cancers.

STRO-002 is a novel homogeneous folate receptor alpha (FolRα) targeting antibody-drug conjugate (ADC) currently being investigated in the clinic as a treatment for ovarian and endometrial cancers. Here, we describe the discovery, optimization, and antitumor properties of STRO-002. STRO-002 was generated by conjugation of a novel cleavable 3-aminophenyl hemiasterlin linker-warhead (SC239) to the nonnatural amino acid para-azidomethyl-L-phenylalanine incorporated at specific positions within a high affinity anti-FolRα antibody using Sutro's XpressCF+, which resulted in a homogeneous ADC with a drug-antibody ratio (DAR) of 4.

Enhanced detection of prion infectivity from blood by preanalytical enrichment with peptoid-conjugated beads.

Prions cause transmissible infectious diseases in humans and animals and have been found to be transmissible by blood transfusion even in the presymptomatic stage. However, the concentration of prions in body fluids such as blood and urine is extremely low; therefore, direct diagnostic tests on such specimens often yield false-negative results. Quantitative preanalytical prion enrichment may significantly improve the sensitivity of prion assays by concentrating trace amounts of prions from large volumes of body fluids.

Targeting CD74 in multiple myeloma with the novel, site-specific antibody-drug conjugate STRO-001.

STRO-001 is a site-specific, predominantly single-species, fully human, aglycosylated anti-CD74 antibody-drug conjugate incorporating a non-cleavable linker-maytansinoid warhead with a drug-antibody ratio of 2 which was produced by a novel cell-free antibody synthesis platform. We examined the potential pharmacodynamics and anti-tumor effects of STRO-001 in multiple myeloma (MM). CD74 expression was assessed in MM cell lines and primary bone marrow (BM) MM biopsies.

RF1 attenuation enables efficient non-natural amino acid incorporation for production of homogeneous antibody drug conjugates.

Amber codon suppression for the insertion of non-natural amino acids (nnAAs) is limited by competition with release factor 1 (RF1). Here we describe the genome engineering of a RF1 mutant strain that enhances suppression efficiency during cell-free protein synthesis, without significantly impacting cell growth during biomass production. Specifically, an out membrane protease (OmpT) cleavage site was engineered into the switch loop of RF1, which enables its conditional inactivation during cell lysis.

Engineering toward a bacterial "endoplasmic reticulum" for the rapid expression of immunoglobulin proteins.

Antibodies are well-established as therapeutics, and the preclinical and clinical pipeline of these important biologics is growing rapidly. Consequently, there is considerable interest in technologies to engineer and manufacture them. Mammalian cell culture is commonly used for production because eukaryotic expression systems have evolved complex and efficient chaperone systems for the folding of antibodies.

Production of site-specific antibody-drug conjugates using optimized non-natural amino acids in a cell-free expression system.

Antibody-drug conjugates (ADCs) are a targeted chemotherapeutic currently at the cutting edge of oncology medicine. These hybrid molecules consist of a tumor antigen-specific antibody coupled to a chemotherapeutic small molecule. Through targeted delivery of potent cytotoxins, ADCs exhibit improved therapeutic index and enhanced efficacy relative to traditional chemotherapies and monoclonal antibody therapies.

A universal method for detection of amyloidogenic misfolded proteins.

Diseases associated with the misfolding of endogenous proteins, such as Alzheimer's disease and type II diabetes, are becoming increasingly prevalent. The pathophysiology of these diseases is not totally understood, but mounting evidence suggests that the misfolded protein aggregates themselves may be toxic to cells and serve as key mediators of cell death. As such, an assay that can detect aggregates in a sensitive and selective fashion could provide the basis for early detection of disease, before cellular damage occurs.

Aβ40 oligomers identified as a potential biomarker for the diagnosis of Alzheimer's disease.

Alzheimer's Disease (AD) is the most prevalent form of dementia worldwide, yet the development of therapeutics has been hampered by the absence of suitable biomarkers to diagnose the disease in its early stages prior to the formation of amyloid plaques and the occurrence of irreversible neuronal damage. Since oligomeric Aβ species have been implicated in the pathophysiology of AD, we reasoned that they may correlate with the onset of disease. As such, we have developed a novel misfolded protein assay for the detection of soluble oligomers composed of Aβ x-40 and x-42 peptide (hereafter Aβ40 and Aβ42) from cerebrospinal fluid (CSF).

The octarepeat region of the prion protein is conformationally altered in PrP(Sc).

Background: Prion diseases are fatal neurodegenerative disorders characterized by misfolding and aggregation of the normal prion protein PrP(C). Little is known about the details of the structural rearrangement of physiological PrP(C) into a still-elusive disease-associated conformation termed PrP(Sc). Increasing evidence suggests that the amino-terminal octapeptide sequences of PrP (huPrP, residues 59-89), though not essential, play a role in modulating prion replication and disease presentation.

Defining the TRiC/CCT interactome links chaperonin function to stabilization of newly made proteins with complex topologies.

Folding within the crowded cellular milieu often requires assistance from molecular chaperones that prevent inappropriate interactions leading to aggregation and toxicity. The contribution of individual chaperones to folding the proteome remains elusive. Here we demonstrate that the eukaryotic chaperonin TRiC/CCT (TCP1-ring complex or chaperonin containing TCP1) has broad binding specificity in vitro, similar to the prokaryotic chaperonin GroEL.

Characterization of prion protein (PrP)-derived peptides that discriminate full-length PrPSc from PrPC.

On our initial discovery that prion protein (PrP)-derived peptides were capable of capturing the pathogenic prion protein (PrP(Sc)), we have been interested in how these peptides interact with PrP(Sc). After screening peptides from the entire human PrP sequence, we found two peptides (PrP(19-30) and PrP(100-111)) capable of binding full-length PrP(Sc) in plasma, a medium containing a complex mixture of other proteins including a vast excess of the normal prion protein (PrP(C)). The limit of detection for captured PrP(Sc) was calculated to be 8 amol from a approximately 10(5)-fold dilution of 10% (wt/vol) human variant Creutzfeldt-Jakob disease brain homogenate, with >3,800-fold binding specificity to PrP(Sc) over PrP(C).

Systems analyses reveal two chaperone networks with distinct functions in eukaryotic cells.

Molecular chaperones assist the folding of newly translated and stress-denatured proteins. In prokaryotes, overlapping sets of chaperones mediate both processes. In contrast, we find that eukaryotes evolved distinct chaperone networks to carry out these functions.

Hsp110 cooperates with different cytosolic HSP70 systems in a pathway for de novo folding.

Molecular chaperones such as Hsp70 use ATP binding and hydrolysis to prevent aggregation and ensure the efficient folding of newly translated and stress-denatured polypeptides. Eukaryotic cells contain several cytosolic Hsp70 subfamilies. In yeast, these include the Hsp70s SSB and SSA as well as the Hsp110-like Sse1/2p.

The cotranslational contacts between ribosome-bound nascent polypeptides and the subunits of the hetero-oligomeric chaperonin TRiC probed by photocross-linking.

The hetero-oligomeric eukaryotic chaperonin TRiC (TCP-1-ring complex, also called CCT) interacts cotranslationally with a diverse subset of newly synthesized proteins, including actin, tubulin, and luciferase, and facilitates their correct folding. A photocross-linking approach has been used to map the contacts between individual chaperonin subunits and ribosome-bound nascent chains of increasing length. Whereas a cryo-EM study suggests that chemically denatured actin interacts with only two TRiC subunits (delta and either beta or epsilon), actin and luciferase chains photocross-link to at least six TRiC subunits (alpha, beta, delta, epsilon, xi, and theta) at different stages of translation.