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PAK4 Inhibition Increases Anti-PD-1 Response in Mouse Models Prompting Phase I Trial

NEW YORK – A new study has identified an apparent role for the PAK4 oncogene in anti-PD-1 checkpoint immunotherapy, prompting a Phase I clinical trial of a PAK4 inhibitor in combination with the PD-1 targeting drug nivolumab (Opdivo from Bristol-Myers Squibb) in around 175 individuals with advanced solid tumors or non-Hodgkin's lymphoma.

The Karyopharm Therapeutics-sponsored trial kicked off in the spring of 2016 and is expected to run until next spring at several sites in the US and Canada, according to available information on ClinicalTrials.gov. It involves a Karyopharm Therapeutics drug candidate known as KPT-9274 that targets both PAK4 and the nicotinamide phosphoribosyl-transferase (NAMPT) enzyme.

In the current study, published online today in Nature Cancer, researchers from the University of California at Los Angeles, Karyopharm Therapeutics, and elsewhere did RNA sequencing on dozens of individuals with melanoma, searching for gene expression features that coincided with enhanced or diminished immune T cell infiltration of tumors in samples collected before and during PD-1 blockage treatment.

In tumors with relatively little T cell infiltration or infiltration by antigen-presenting dendritic cells, the team reportedly saw an over-representation of the kinase enzyme-coding gene PAK4. On the other hand, T cell infiltration and anti-PD-1 treatment responses were improved in a B16 mouse model strain of melanoma missing the PAK4, or in B16 animals treated with both a PD-1 inhibitor and the PAK4/NAMPT inhibitor KPT-9274.

Likewise, the researchers reported, response to a similar checkpoint blockade treatment could be improved by targeting or knocking out PAK4 in a mouse model of  colon adenocarcinoma — a tumor type that is prone to having a high tumor mutational burden but does not always respond to anti-PD-1 monotherapy.

"Altogether," the study's authors wrote, "these data suggest that PAK4 inhibition synergizes with anti-PD-1 treatment."

Starting with 66 pre-treatment or on-treatment samples from 41 individuals with melanoma, the researchers successfully sequenced RNA in 27 samples collected prior to anti-PD-1 treatment, 13 samples from anti-PD-1 responders on treatment, and 14 samples from patients who did not respond to the checkpoint immunotherapy.

With the help of an established microenvironment cell population counter method, the team scrutinized gene expression patterns across immune cell populations in the responder and non-responder samples.

Along with differences in T cell, CD8+ T cell, myeloid dendritic cell, and natural killer cell infiltration in samples from patients who did respond to treatment, for example, the researchers saw higher-than-usual PAK4 expression in samples with diminished infiltration and treatment non-response. Those results were supported by their subsequent analyses of RNA sequence, immune cell infiltration, and other data for melanoma and other cancer types previously assessed for the Cancer Genome Atlas project and other published analyses.

"We found that biopsies of patients who did not respond to PD-1 blockade showed an overexpression of PAK4, so that led us to believe it played a role in suppressing the immunotherapy treatment," lead author Gabriel Abril-Rodriguez, a doctoral student in pharmacology and medicine at the UCLA David Geffen School of Medicine, said in a statement.

In their mouse models, meanwhile, the researchers relied on cytometry by time-of-flight-based immune profiling to define immune cell clusters and uncover immune cell infiltration differences in mice that had the PAK4 gene edited out. Results from those experiments, coupled with data from mice receiving both anti-PD-1 and anti-PAK4 treatments, suggested that PD-1 checkpoint blockade responses and immune cell infiltration are enhanced in the absence of PAK4.

"Genetic and pharmacological PAK4 inhibition altered WNT/[beta]-catenin signaling, increased intra-tumoral T cell infiltration, and improved the response to checkpoint blockade therapy in two mouse models," Abril-Rodriguez and his co-authors reported. "The negative correlation between PAK4 expression and T cell infiltration held true across several human cancers, including cancers notoriously resistant to PD-1 blockade, and hence expands the potential clinical applicability of the combined inhibition of PAK4 and PD-1."