Emerging data support a critical role for PD-L1/PD-1 signaling in tumor immunity, with approximately 30- 50% of clinical cancers responding to antibody-mediated immune checkpoint blockade. Based on immunohistochemistry staining of human tumor samples, some tumor cells displayed very little PD-L1 staining (as in Medulloblastoma tumor samples; Majzner RG et al. Cancer Res. 2015; 75(15 Suppl): Abstract nr 249. Doi:10.1158/1538-7445.AM2015-249). However, ongoing immunotherapy trials supports the notion that tumor- associated PD-L1 expression is highly dynamic, and one of the known positive regulator of PD-L1 expression on tumor cells is interferon-gamma (IFN?). Recently, we made a novel observation that cyclin-dependent kinase 5, a proline-directed serine-threonine kinase that is aberrantly active across many tumor types, plays an important immune-modulatory role in controlling PD-L1 up-regulation in response to IFN? signaling (Dorand et al. Science 2016; 353(6297): 399-403). Genetic and pharmacologic disruption of Cdk5 function in MM1, a Ptch+/-/p53-/- derived syngeneic tumor model, sensitizes the aggressive tumor to CD4+ T cell-dependent rejection. This phenotype was associated with a blunted PD-L1 up-regulated in response to IFN? stimulation, a process associated with the hyper-phosphorylation of Serine-439, Serine-440 and Serine-443 on IRF2BP2, a co-repressor in complex with IRF2 in suppressing PD-L1 gene transcription. The same phenomenon can also be observed in human and mouse rhabdomyosarcoma (RMS) cell lines. All these lines share similar genetic mutations in SHH and p53 pathways. In the proposal, we wish to provide mechanistic understanding of how Cdk5 controls PD-L1 expression via IRF2BP2. We hypothesize that hyper-phosphorylation of S439, S440 and S443 on IRF2BP2 in Cdk5-deficient tumors prolongs the half-lives of IRF2BP2 and its co-repressor IRF2, resulting in sustain PD-L1 suppression at the transcriptional level upon IFN? exposure, and reduced phosphorylation at these sites will result in IFN?-induced PD-L1 up-regulation in tumors that harbor p53 and SHH alteration.
In Aim 1, we propose to study the functional link betweenPD-l1 transcription and site-specific IRF2BP2 hyper-phosphorylation by creating tumor harboring S->A or S->D phosphomimetic mutants, or target the putative kinases (GSK3 and ERK1/2) and phosphatase (PP2A) potentially associated with this signaling cascade.
In Aim 2, we will extend our molecular interrogation to include other tumor types harboring p53 / SHH alterations, including RMS and pancreatic cancers that have been found to frequently harbor these genetic alterations, and patient-derived xenograft (PDX) lines available in our PDX repository at the Angie Fowler AYA Cancer Institute. These additional molecular investigations will provide a solid functional link between Cdk5 and PD-L1 regulation in tumors, and allows the future pursuit of potent next-generation Cdk5 inhibitor, CYC065, in enhancing efficacy of clinical immunotherapeutic trials.
Public Health Relevance Statement: Despite recent successes in immune checkpoint blockade approach in refractory cancers, more than half of all patients still do not benefit from this revolutionary therapeutic modality. Among the responders, severe autoimmunity represents a limiting and sometimes life-threatening complication. We have recently discovered an entirely novel pathway by which tumor cells modulate PD-L1 expression via cyclin-dependent kinase 5 (Cdk5) upon exposure to interferon-gamma, and we have identified candidate serine residues on the co- repressor of PD-L1 gene transcription, IRF2BP2, which may be indirect targets of Cdk5 through other kinases and phosphatases. Success of our current effort may identify additional molecular targets to regulate tumor- specific immune checkpoint.
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| Dorand, R Dixon; Petrosiute, Agne; Huang, Alex Y (2017) Multifactorial regulators of tumor programmed death-ligand 1 (PD-L1) response. Transl Cancer Res 6:S1451-S1454 |
