Abstract 869P
Background
Despite remarkable clinical success with immune checkpoint inhibitor (ICI) therapy targeting the PD-1 pathway for the treatment of melanoma some patients still remain unresponsive to immunotherapy. Using an unbiased mass spectrometry (LC-MS) discovery-based approach we identified a panel of protein biomarkers that correlated with ICI therapy response, one of which was the p21-activated kinase 4 (PAK4) which we found to be significantly elevated in non-responding tumor biopsies. This finding is in line with a previously published study where PAK4 was reported to be enriched in non-responding tumor biopsies and its inhibition was shown to improve PD-1 blockade (Abril-Rodriguez et al. Nat Cancer 1, 46–58 (2020)). For an in-depth analysis of PAK4 in these same patient samples, we followed up with spatial tissue analysis using a multiplexed immunofluorescence (mIF) platform to gain information on PAK4 mechanisms in melanoma patients treated with ICI therapy.
Methods
FFPE tumor samples were firstly used for unbiased whole proteome profiling using TrueDiscoveryTM, a data-independent acquisition (DIA) LC-MS technology. Baseline patient samples were classified as responders (n = 9) or non-responders (n = 15) based on the response at 3 months post ICI-treatment. Subsequently, the same patient samples were analyzed by an 18-marker custom panel using MultiOmyx™, a multiplexed immunofluorescence (mIF) assay utilizing a pair of conjugated Cyanine dye-labeled antibodies per round of staining coupled with the deep-learning-based cell classification platform NeoLYTX.
Results
PAK4 was found to be elevated in tumor samples from non-responder patients by both unbiased proteomics analysis, as well as MultiOmyx mIF analysis. Furthermore, we observed a clear difference in the correlation to other immune cells between the two patient groups by MultiOmyx analysis, PAK4 density being negatively correlated to T cells and TAMs only in non-responders, while positively correlated to MDSCs only in responders.
Conclusions
Analysis of the 24 FFPE samples from metastatic melanoma patients treated with ICI therapy stratified into responders and non-responders lead to the identification of PAK4 as a potential marker for poor response to immunotherapy in melanoma patient samples.
Clinical trial identification
Editorial acknowledgement
Legal entity responsible for the study
Biognosys AG.
Funding
In kind funding from Biognosys AG and NeoGenomics.
Disclosure
J. Kuo: Financial Interests, Institutional, Full or part-time Employment: Neogenomics. H. Nunns: Financial Interests, Institutional, Full or part-time Employment: Neogenomics. T. Treiber: Financial Interests, Institutional, Full or part-time Employment, Options: Biognosys. K. Kakalacheva - Beeler: Financial Interests, Institutional, Full or part-time Employment, Options: Biognosys; Financial Interests, Institutional, Officer: Biognosys. J. Vowinckel: Financial Interests, Institutional, Full or part-time Employment, Options: Biognosys. A. Juncker-Jensen: Financial Interests, Institutional, Full or part-time Employment: Neogenomics; Financial Interests, Institutional, Stocks/Shares: Neogenomics. All other authors have declared no conflicts of interest.