Analogs of the hepatocyte growth factor and macrophage-stimulating protein hinge regions act as Met and Ron dual inhibitors in pancreatic cancer cells

Abstract

Pancreatic cancer is among the leading causes of cancer death in the United States with limited effective treatment options. A major contributor to the formation and persistence of pancreatic cancer is the dysregulation of the hepatocyte growth factor (HGF)/Met (HGF receptor) and the macrophage stimulating protein (MSP)/Ron (MSP receptor) systems. These systems normally mediate a variety of cellular behaviors including proliferation, survival, and migration, but are often over-activated in pancreatic cancer and contribute to cancer progression. Previous studies have shown that HGF must dimerize in order to activate Met. Small molecule antagonists with homology to a “hinge” region within the putative dimerization domain of HGF have been developed that bind to HGF and block dimerization, therefore inhibiting Met signaling. Due to the structural and sequence homology between MSP and HGF, we hypothesized that the inhibition of HGF by the hinge analogs may extend to MSP. The primary purpose of this “proof-of-concept” study was to determine if hinge analogs could inhibit cellular responses to both HGF and MSP in pancreatic cancer cells. Our results demonstrated that these compounds inhibited HGF and MSP activity. Hinge analog treatment resulted in decreased Met and Ron activation, and suppressed malignant cell behaviors including proliferation, migration, and invasion in pancreatic cancer cells in vitro. These results suggest that the hinge analogs represent a novel group of molecules that may offer a therapeutic approach for the treatment of pancreatic cancer and warrant further development and optimization.

Norleual, a hepatocyte growth factor and macrophage stimulating protein dual antagonist, increases pancreatic cancer sensitivity to gemcitabine

“…The LM-P pancreatic cell line is derived from KPC (Kras G12D/+ ;LSL-Trp53 R172H/+ ;Pdx-1-Cre mice) mice and expresses mutant K-Ras. Treatment with Norleual increased LM-P sensitivity to gemcitabine (Figure 3C), and in previous studies significantly inhibited cell signaling and malignant behaviors [21], indicating that Norleual demonstrates anti-cancer activity in both K-Ras mutant and wild-type pancreatic cancer cell lines.…”

“…MSP is normally present in serum [15] and while we did not specifically test for activation of the MSP receptor by the serum, we cannot rule out that Norleual activity in the absence of exogenous growth factors may be in part due to inhibition of MSP. Given that we have previously shown Norleual to inhibit MSP-induced signaling and migration [21], and inhibition of MSP-dependent survival in the present study (Figure 1d), it is likely that the effects of Norleual treatment in vitro and in the orthotopic mouse study is to some extent due to inhibition of MSP activity. Regardless of the source in vitro , the pancreatic tumor cells orthotopically implanted in the nude mice had a steady supply of HGF and MSP as these growth factors are readily available in vivo .…”

“…HGF is produced by the tumor stroma including tumor associated fibroblasts [35], and MSP is constitutively secreted into circulation [34]. The U-shaped dose-response curve observed in the survival assays (Figure 1), wherein high and low concentrations failed to significantly inhibit survival, have been observed and discussed previously [21]. While the exact mechanism for this dose-response relationship is unknown, a possible explanation is that Norleual aggregates or there is a low affinity off-target binding site at high concentrations.…”

“…Previous work with the hinge analog Norleual (Nle-Tyr-Leu-ψ-(CH 2 -NH 2 ) 3-4 -His-Pro-Phe), has demonstrated that Norleual is a potent inhibitor of the HGF/Met system, inhibited the pro-survival effects of HGF, and suppressed lung colonization in a murine melanoma cell model [17, 18]. Recent studies have shown that Norleual can inhibit the HGF/Met system and associated behaviors in pancreatic cancer cells including proliferation, migration, and invasion [21]. Interestingly, these studies have also shown that Norleual can inhibit MSP-dependent signaling and migration in pancreatic cancer cells [21], likely due to the substantial sequence homology between HGF and MSP.…”

“…Recent studies have shown that Norleual can inhibit the HGF/Met system and associated behaviors in pancreatic cancer cells including proliferation, migration, and invasion [21]. Interestingly, these studies have also shown that Norleual can inhibit MSP-dependent signaling and migration in pancreatic cancer cells [21], likely due to the substantial sequence homology between HGF and MSP. Considering that previous results indicate Norleual acts a dual antagonist of HGF and MSP, we hypothesized that Norleual treatment may sensitize pancreatic cancer cells to gemcitabine due to inhibition of the pro-survival effects of the MSP/Ron and HGF/Met systems.…”