PDE4D targeting enhances anti-tumor effects of sorafenib in clear cell renal cell carcinoma and attenuates MAPK/ERK signaling in a CRAF-dependent manner.

Clear cell renal cell carcinoma (ccRCC) is the most lethal form of kidney cancer and effective treatment regimens are yet to be established. Tyrosine kinase inhibitors (TKI) have widely been used as ccRCC therapeutics, but their efficacy is limited due to accompanying resistance mechanisms. Previous studies have provided substantial evidence for crosstalk between cAMP and the MAPK/ERK signaling pathway. Low levels of intracellular cAMP have been found in several human malignancies and some data suggest that elevation of cAMP expression can be achieved by phosphodiesterase 4 (PDE4) inhibition, resulting in cell growth arrest and/or cell death. The effects of crosstalk between cAMP and the MAPK/ERK pathway on the development progression in ccRCR, however, remain to be fully understood. In this study, we sought to explore the involvement of PDE4 in ccRCC and to assess its potential as a target for therapeutic intervention. We demonstrated that PDE4D is the predominant subtype of PDE4 expressed in healthy and cancerous renal cell lines, particularly in metastatic Caki-1 cells. We generated a CRISPR/Cas9-mediated PDE4D-KO Caki-1 cell model and showed that PDE4D depletion reduced cell proliferation and recovered cAMP expression in these cells. PDE4D-KO and/or PDE4 inhibition with the FDA approved PDE4 inhibitor, roflumilast, also attenuated MAPK/ERK signaling in a CRAF-dependent manner. Most interestingly, we showed that PDE4D-KO enhanced the effectiveness of the TKI, sorafenib, to stunt cell survival. In conclusion, we provide preliminary evidence of PDE4 involvement in ccRCC and suggest a rationale for dual tyrosine kinase/PDE4D targeting in patients with CRAF-dependent MAPK activation.

Platelet-derived growth factor (PDGF) cross-signaling via non-corresponding receptors indicates bypassed signaling in colorectal cancer.

Platelet-derived growth factor (PDGF) signaling, besides other growth factor-mediated signaling pathways like vascular endothelial growth factor (VEGF) and epidermal growth factor (EGF), seems to play a crucial role in tumor development and progression. We have recently provided evidence for upregulation of PDGF expression in UICC stage I-IV primary colorectal cancer (CRC) and demonstrated PDGF-mediated induction of PI3K/Akt/mTOR signaling in CRC cell lines. The present study sought to follow up on our previous findings and explore the alternative receptor cross-binding potential of PDGF in CRC. Our analysis of primary human colon tumor samples demonstrated upregulation of the PDGFRß, VEGFR1, and VEGFR2 genes in UICC stage I-III tumors. Immunohistological analysis revealed co-expression of PDGF and its putative cross-binding partners, VEGFR2 and EGFR. We then analyzed several CRC cell lines for PDGFRα, PDGFRß, VEGFR1, and VEGFR2 protein expression and found these receptors to be variably expressed amongst the investigated cell lines. Interestingly, whereas Caco-2 and SW480 cells showed expression of all analyzed receptors, HT29 cells expressed only VEGFR1 and VEGFR2. However, stimulation of HT29 cells with PDGF resulted in upregulation of VEGFR1 and VEGFR2 expression despite the absence of PDGFR expression and mimicked the effect of VEGF stimulation. Moreover, PDGF recovered HT29 cell proliferation under simultaneous treatment with a VEGFR or EGFR inhibitor. Our results provide some of the first evidence for PDGF cross-signaling through alternative receptors in colorectal cancer and support anti-PDGF therapy as a combination strategy alongside VEGF and EGF targeting even in tumors lacking PDGFR expression.

KMP01D Demonstrates Beneficial Anti-inflammatory Effects on Immune Cells: An ex vivo Preclinical Study of Patients With Colorectal Cancer.

Background: Colorectal cancer (CRC) is frequently associated with dysbiosis of the gut microbiome which, together with a compromised gut barrier, can result in perioperative endotoxin leakage into the circulation. Constant local and systemic inflammatory activity is suggested to facilitate metastases formation. Previous studies have pointed to the capacity of a colostrum preparation to neutralize endotoxins within the gastrointestinal tract which could ameliorate associated inflammatory responses and tumor recurrence in affected patients. This study aimed to examine the effects of the colostrum preparation, KMP01D, on the inflammatory activity of patient-derived immune cells. Methods: The effects of KMP01D on pro-/anti-inflammatory cytokine responses and apoptosis were examined ex vivo using immune cells from CRC patients (stages I-IV, n = 48). The expression of CD14, CD68, Toll-like receptor (TLR)4, and insulin-like growth factor (IGF)-1 was also analyzed. Results: KMP01D increased interleukin (IL)-10 and IL-13 anti-inflammatory cytokine expression in patient-derived peripheral blood mononuclear cells (PBMCs). Interestingly, KMP01D also decreased the secretion of IL-1ß, IL-6, interferon (IFN)-γ, tumor necrosis factor (TNF)-α, IL-12 inflammatory cytokines, and IGF-1 in these cells. Moreover, CD14 and TLR4 expression involved in endotoxin signaling was downregulated in PBMCs and tumor-derived cells. Apoptosis of immune cells and tumor-derived cells was likewise enhanced with KMP01D. Addition of vitamin D3 as a cofactor demonstrated enhanced anti-inflammatory effects. Conclusions: KMP01D demonstrated beneficial ex vivo effects on inflammatory cytokine responses in PBMCs and enhanced apoptosis of immune cells from CRC patients. In line with previous clinical trials, we present new evidence endorsing KMP01D as a treatment strategy to regulate stage-dependent local and systemic inflammation in CRC patients.

Expression of Tumor-mediated CD137 ligand in human colon cancer indicates dual signaling effects.

CD137-targeting immune therapy, which activates anti-tumor T effector cell responses, seems to be an attractive concept in clinical oncology. Recent evidence has demonstrated that tumor cells besides T cells and antigen-presenting cells are able to express CD137 and CD137L. Here we aimed to identify CD137/CD137L expression in established colon cancer cell lines and primary tumors (UICC stages I-IV) from patients with documented long-term follow-up. CD137/CD137L expression was highly upregulated in early to late-stage tumors while the inverse was observed in patient-derived peripheral blood mononuclear cells. High CD137L expression within primary tumors was mediated by tumor cells and significantly correlated with the occurrence of distant metastases and shortened survival in advanced stages of disease (UICC stage IV). Interestingly, induced tumor cell signaling via CD137L on its surface in vitro resulted in dual effects: (i) reduced tumor cell proliferation suggesting inhibitory signaling in all investigated cancers and (ii) increased epithelial-to-mesenchymal transition signaling events. Taken together CD137/CD137L expression was stage-dependently upregulated with shortened survival in patients with highly CD137L-expressing tumors. Our clinical and experimental data suggest that colon cancer cells predominantly express CD137L and thereby have negative impact on overall survival through a process of reverse signaling. Beside agonistic CD137 antibody therapy to foster T effector cell responses, CD137L-mediated intervention strategies may become instrumental to circumvent relapsed tumor growth through induced epithelial-to-mesenchymal transition and consecutive metastases formation.