IFITM Protein Modulation Found to Impact Drug Potency

Traditional pharmaceuticals typically include small molecules with favorable chemical characteristics that cells can easily take up, but this isn’t the case for all drug targets. Certain kinase enzymes, for example, are notoriously difficult to target with traditional drugs due to specificity issues. To mitigate these issues and access new cellular targets thought to be “undruggable”, researchers from UCSF and the Arc Institute discovered a cellular uptake pathway that can accommodate larger drug molecules composed of linked subunits.

As newer classes of drugs became more complex, it became clear that there were limitations to getting these drugs to permeate through the cell membrane. Larger drug targets with multiple parts also have a hard time entering cells, requiring multiple ligands to be bound to receptor proteins before successful entry. “People have been reluctant to design, synthesize and test such molecules as they are so far beyond standard drug design rules that people just assumed they wouldn’t enter cells,” explains co-author Luke Gilbert. 

To learn how these large, complex drugs get into cells, they conducted a genome-wide functional screen to test the importance of individual genes on cellular processes. Their work, published in the journal Science, utilized CRISPRi and CRISPRa screens pioneered by Gilbert, where CRISPR machinery is used with a library of guide RNAs to decrease or increase expression levels of one gene at a time across the whole human genome. When applied to a pool of millions of cells, each cell receives a different modification at a different gene, and researchers can then determine which genetic manipulations lead to differences in functional outcomes of interest. For the study, the team was looking for gene expression manipulations that either made cells more sensitive or more resistant to a linked drug molecule.

Gilbert’s team and Kevan Shokat’s lab conducted paired CRISPRi and CRISPRa screens in human leukemia cell lines, followed by treatment with a large linked experimental anti-cancer drug, RapaLink-1. Their analyses revealed that three closely related genes seemed to promote RapaLink-1 activity but had no activity on unlinked drugs that were tested. These genes encode interferon-induced transmembrane (IFITM) proteins, which have been previously recognized for their roles in antiviral defense. However, no prior evidence has suggested that these proteins can impact drug efficacy. By modulating levels of IFITM proteins, the researchers could drastically change the potency of RapaLink-1 by about 30-fold.  

The group looked across 659 different cell types and saw a strong correlation between the level of IFITM expression and sensitivity to RapaLink-1, supporting a general role across different types of cells. After expanding to a larger set of 17 different linked and unlinked drug molecules, the team determined that the influence of IFITM gene expression was consistent across diverse types of linked drugs.

Based on this information, the Shokat lab next designed two new linked drugs, DasatiLink-1 and BisRoc-1, that they hypothesized might take advantage of this cellular entry pathway. Despite violating traditional drug design principles, the teams revealed that both drugs could enter cells, bind successfully with their intended targets, and work just as well as the unlinked versions. Additionally, these linked versions were uniquely dependent on IFITM protein expression in target cells, supporting a general role for the IFITM pathway across many linked molecules.

This discovery not only helps answer a long-standing question in biomedicine but also paves the way for better molecular linker design that can exploit this entry pathway for higher drug effectiveness and specificity. In the future, Gilbert suggests that it could be possible to “​​exploit pathways that mediate drug uptake, like IFITM, to boost drug uptake or even to target a drug to select cell types.”