A new KSRP-binding compound suppresses distant metastasis of colorectal cancer by targeting the oncogenic KITENIN complex

Abstract

Background Distant metastasis is the major cause of death in patients with colorectal cancer (CRC). Previously, we identified KITENIN as a metastasis-enhancing gene and suggested that the oncogenic KITENIN complex is involved in metastatic dissemination of KITENIN-overexpressing CRC cells. Here, we attempted to find substances targeting the KITENIN complex and test their ability to suppress distant metastasis of CRC. Methods We screened a small-molecule compound library to find candidate substances suppressing the KITENIN complex in CRC cells. We selected a candidate compound and examined its effects on the KITENIN complex and distant metastasis through in vitro assays, a molecular docking model, and in vivo tumor models. Results Among several compounds, we identified DKC1125 (Disintegrator of KITENIN Complex #1125) as the best candidate. DKC1125 specifically suppressed KITENIN gain of function. After binding KH-type splicing regulatory protein (KSRP), DKC1125 degraded KITENIN and Dvl2 by recruiting RACK1 and miRNA-124, leading to the disintegration of the functional KITENIN–KSRP–RACK1–Dvl2 complex. A computer docking model suggested that DKC1125 specifically interacted with the binding pocket of the fourth KH-domain of KSRP. KITENIN-overexpressing CRC cells deregulated certain microRNAs and were resistant to 5-fluorouracil, oxaliplatin, and cetuximab. DKC1125 restored sensitivity to these drugs by normalizing expression of the deregulated microRNAs, including miRNA-124. DKC1125 effectively suppressed colorectal liver metastasis in a mouse model. Interestingly, the combination of DKC1125 with 5-fluorouracil suppressed metastasis more effectively than either drug alone. Conclusion DKC1125 targets the KITENIN complex and could therefore be used as a novel therapeutic to suppress liver metastasis in CRC expressing high levels of KITENIN.

A peptide interfering with the dimerization of oncogenic KITENIN protein and its stability suppresses colorectal tumour progression

“…41 In this study, we introduced KDIP, a short peptide that effectively suppressed oncogenic KITENIN in vitro and in vivo through specific inhibition of the dimerization of KITENIN and by reducing the stability of the KITENIN protein. Adding to reports of siRNA, 25 mi-RNA 26 and small-molecular-weight compounds 27 targeting the functional KITENIN complex, our present result is the first report on the inhibition of the oncogenic function of KITENIN by a peptide. RACK1 was previously identified as a novel hypoxiainducible factor-1α (HIF-1α)-interacting protein that regulates HIF-1α stability through competition with HSP90 and recruitment of the Elongin-C/B ubiquitin ligase complex.…”

“… 41 In this study, we introduced KDIP, a short peptide that effectively suppressed oncogenic KITENIN in vitro and in vivo through specific inhibition of the dimerization of KITENIN and by reducing the stability of the KITENIN protein. Adding to reports of siRNA, 25 mi‐RNA 26 and small‐molecular‐weight compounds 27 targeting the functional KITENIN complex, our present result is the first report on the inhibition of the oncogenic function of KITENIN by a peptide.…”

“…In this regard, KITENIN is a promising target for anti‐metastasis therapy. 27 As shown in Figures 6G and S3A , KDIP had a specific effect on the KITENIN dimer and inhibited the downstream signalling of the KITENIN complex. After treatment with KDIP, Myo10 was decreased via proteasomal degradation and thereby the KITENIN dimer lost its stability.…”

“… 33 We previously found that RACK1 serves as an adaptor protein for the molecules involved in the downstream signalling of the KITENIN complex in which RACK1 might control the level of Dvl2 and KITENIN via autophagy‐dependent degradation. 27 To check whether RACK1 directly interacts with KITENIN, we performed an IP experiment using antibody against GFP in Caco2 cells co‐expressing RACK1‐GFP and KITENIN‐V5 and found that RACK1 bound with KITENIN (Figure 3A , left). Also, RACK1 directly interacted with endogenous KITENIN in non‐transfected HCT116 cells (Figure S5A ).…”

“…A syngeneic mouse model of colorectal liver metastasis was established by an infusion of tumour cells into the portal system via intrasplenic injection. 27 In brief, CT‐26/KITENIN‐iRFP‐expressing cells (2 × 10 5 cells/mouse) were injected into the spleen. Splenectomy was performed 15 min after an injection of CT‐26 cells.…”