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<p>We have developed a broadly applicable platform that harnesses the power of protein
engineering and genetic screening to produce efficacious protein-drug combinations
for cancer therapy. For proof-of-concept, we implemented this strategy to engineer
targeted pro-apoptotic drug combinations that overcome cancer resistance to protein
agonists of death receptor 5 (DR5), a key upregulated marker in colorectal cancer
(CRC). Over the past decade, various DR5 agonists have shown poor clinical efficacy,
including both engineered antibodies and TRAIL, the natural ligand for this receptor.
Comprehensive studies suggest that there are three major obstacles to success of these
agents: 1) potency, 2) delivery, and 3) resistance. </p><p>We have systematically
addressed these challenges by engineering a sustained-release formulation of a highly
potent, hexavalent death receptor 5 agonist (DRA), and administering the agonist as
a sustained release depot, in combination with rationally nominated targeted drugs
that overcome intrinsic resistance to DRAs. To address the need for sustained delivery
of therapeutic proteins, we developed injectable depots of DRAs recombinantly fused
to thermally responsive elastin-like polypeptide (ELP) biopolymers. The bioactive
ELP-DRA fusions undergo temperature-driven phase transition upon subcutaneous injection
in vivo, resulting in the formation of a gel-like depot suitable for sustained drug
delivery. A single 30 mg/kg injection of the gel-like ELP-DRA depot induced significant
tumor regression in Colo205 mouse xenografts. To pinpoint the genetic drivers of CRC
resistance to the DRA, we used a gain-of-function ORF screen and a CRISPR/Cas9 knockout
screen. The screens identified genes that confer sensitivity to the DRA in resistant
CRC cell lines. Over twenty small molecule drugs targeting pathways and proteins
identified from the screens were then tested in combination with the DRA to identify
highly synergistic combinations using cytotoxicity assays. Clonogenic, time-to-progression,
and cell viability assays showed that pharmacological blockade of XIAP, Bcl-XL, and
CDK4/6 strongly enhances antitumor activity of DRA in established human CRC cell lines
and patient-derived CRC cells. In vivo tumor regression studies demonstrated the potent
anti-tumor efficacy of combining inhibitors of XIAP and Bcl-XL with the sustained
release formulation of ELP-DRA.</p><p>By addressing both delivery and resistance issues
with our protein engineering and genomics platform, we have overcome the key obstacles
to DRA translation as a successful drug in the clinic. Our rational approach elegantly
provides optimal protein-small molecule drug combinations that elicit a robust anticancer
response, exhibit minimal toxicity, and combat drug resistance.</p>
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