Overcoming sorafenib evasion in hepatocellular carcinoma using CXCR4-targeted nanoparticles to co-deliver MEK-inhibitors.

Sorafenib is a RAF inhibitor approved for several cancers, including hepatocellular carcinoma (HCC). Inhibition of RAF kinases can induce a dose-dependent "paradoxical" upregulation of the downstream mitogen-activated protein kinase (MAPK) pathway in cancer cells. It is unknown whether "paradoxical" ERK activation occurs after sorafenib therapy in HCC, and if so, if it impacts the therapeutic efficacy. Here, we demonstrate that RAF inhibition by sorafenib rapidly leads to RAF dimerization and ERK activation in HCCs, which contributes to treatment evasion. The transactivation of RAF dimers and ERK signaling promotes HCC cell survival, prevents apoptosis via downregulation of BIM and achieves immunosuppression by MAPK/NF-kB-dependent activation of PD-L1 gene expression. To overcome treatment evasion and reduce systemic effects, we developed CXCR4-targeted nanoparticles to co-deliver sorafenib with the MEK inhibitor AZD6244 in HCC. Using this approach, we preferentially and efficiently inactivated RAF/ERK, upregulated BIM and down-regulated PD-L1 expression in HCC, and facilitated intra-tumoral infiltration of cytotoxic CD8+ T cells. These effects resulted in a profound delay in tumor growth. Thus, this nano-delivery strategy to selectively target tumors and prevent the paradoxical ERK activation could increase the feasibility of dual RAF/MEK inhibition to overcome sorafenib treatment escape in HCC.

CXCR4 inhibition in tumor microenvironment facilitates anti-programmed death receptor-1 immunotherapy in sorafenib-treated hepatocellular carcinoma in mice.

UNLABELLED: Sorafenib, a broad tyrosine kinase inhibitor, is the only approved systemic therapy for advanced hepatocellular carcinoma (HCC) but provides limited survival benefits. Recently, immunotherapy has emerged as a promising treatment strategy, but its role remains unclear in HCCs, which are associated with decreased cytotoxic CD8(+) T-lymphocyte infiltration in both murine and human tumors. Moreover, in mouse models after sorafenib treatment intratumoral hypoxia is increased and may fuel evasive resistance. Using orthotopic HCC models, we now show that increased hypoxia after sorafenib treatment promotes immunosuppression, characterized by increased intratumoral expression of the immune checkpoint inhibitor programmed death ligand-1 and accumulation of T-regulatory cells and M2-type macrophages. We also show that the recruitment of immunosuppressive cells is mediated in part by hypoxia-induced up-regulation of stromal cell-derived 1 alpha. Inhibition of the stromal cell-derived 1 alpha receptor (C-X-C receptor type 4 or CXCR4) using AMD3100 prevented the polarization toward an immunosuppressive microenvironment after sorafenib treatment, inhibited tumor growth, reduced lung metastasis, and improved survival. However, the combination of AMD3100 and sorafenib did not significantly change cytotoxic CD8(+) T-lymphocyte infiltration into HCC tumors and did not modify their activation status. In separate experiments, antibody blockade of the programmed death ligand-1 receptor programmed death receptor-1 (PD-1) showed antitumor effects in treatment-naive tumors in orthotopic (grafted and genetically engineered) models of HCC. However, anti-PD-1 antibody treatment had additional antitumor activity only when combined with sorafenib and AMD3100 and not when combined with sorafenib alone. CONCLUSION: Anti-PD-1 treatment can boost antitumor immune responses in HCC models; when used in combination with sorafenib, anti-PD-1 immunotherapy shows efficacy only with concomitant targeting of the hypoxic and immunosuppressive microenvironment with agents such as CXCR4 inhibitors.

A phase 1/2 and biomarker study of preoperative short course chemoradiation with proton beam therapy and capecitabine followed by early surgery for resectable pancreatic ductal adenocarcinoma.

PURPOSE: To evaluate the safety, efficacy and biomarkers of short-course proton beam radiation and capecitabine, followed by pancreaticoduodenectomy in a phase 1/2 study in pancreatic ductal adenocarcinoma (PDAC) patients. METHODS AND MATERIALS: Patients with radiographically resectable, biopsy-proven PDAC were treated with neoadjuvant short-course (2-week) proton-based radiation with capecitabine, followed by surgery and adjuvant gemcitabine. The primary objective was to demonstrate a rate of toxicity grade ≥ 3 of <20%. Exploratory biomarker studies were performed using surgical specimen tissues and peripheral blood. RESULTS: The phase 2 dose was established at 5 daily doses of 5 GyE. Fifty patients were enrolled, of whom 35 patients were treated in the phase 2 portion. There were no grade 4 or 5 toxicities, and only 2 of 35 patients (4.1%) experienced a grade 3 toxicity event (chest wall pain grade 1, colitis grade 1). Of 48 patients eligible for analysis, 37 underwent pancreaticoduodenectomy. Thirty of 37 (81%) had positive nodes. Locoregional failure occurred in 6 of 37 resected patients (16.2%), and distant recurrence occurred in 35 of 48 patients (72.9%). With median follow-up of 38 months, the median progression-free survival for the entire group was 10 months, and overall survival was 17 months. Biomarker studies showed significant associations between worse survival outcomes and the KRAS point mutation change from glycine to aspartic acid at position 12, stromal CXCR7 expression, and circulating biomarkers CEA, CA19-9, and HGF (all, P<.05). CONCLUSIONS: This study met the primary endpoint by showing a rate of 4.1% grade 3 toxicity for neoadjuvant short-course proton-based chemoradiation. Treatment was associated with favorable local control. In exploratory analyses, KRAS(G12D) status and high CXCR7 expression and circulating CEA, CA19-9, and HGF levels were associated with poor survival.

Dehydro-alpha-lapachone, a plant product with antivascular activity.

Antivascular agents have become a standard of treatment for many malignancies. However, most of them target the VEGF pathway and lead to refractoriness. To improve the diversity of options for antivascular therapy, we applied a high-throughput screen for small molecules targeting cell adhesion. We then assayed the resulting antiadhesion hits in a transgenic zebrafish line with endothelial expression of EGFP (Tg(fli1:EGFP)(y1)) to identify nontoxic molecules with antivascular activity selective to neovasculature. This screen identified dehydro-α-lapachone (DAL), a natural plant product. We found that DAL inhibits vessel regeneration, interferes with vessel anastomosis, and limits plexus formation in zebrafish. Furthermore, DAL induces vascular pruning and growth delay in orthotopic mammary tumors in mice. We show that DAL targets cell adhesion by promoting ubiquitination of the Rho-GTPase Rac1, which is frequently up-regulated in many different cancers.

A safety and survival analysis of neoadjuvant bevacizumab with standard chemoradiation in a phase I/II study compared with standard chemoradiation in locally advanced rectal cancer.

INTRODUCTION: Bevacizumab is increasingly being tested with neoadjuvant regimens in patients with localized cancer, but its effects on metastasis and survival remain unknown. This study examines the long-term outcome of clinical stage II/III rectal cancer patients treated in a prospective phase II study of bevacizumab with chemoradiation and surgery. As a benchmark, we used data from an analysis of 42 patients with locally advanced rectal cancer treated with a contemporary approach of preoperative fluoropyrimidine-based radiation therapy. MATERIALS AND METHODS: Outcome analyses were performed on 32 patients treated prospectively with neoadjuvant bevacizumab, 5-fluorouracil, radiation therapy, and surgery as well as 42 patients treated with standard fluoropyrimidine-based chemoradiation. RESULTS: Overall survival, disease-free survival, and local control showed favorable trends in patients treated with bevacizumab with chemoradiation followed by surgery. Acute and postoperative toxicity appeared acceptable. CONCLUSIONS: Neoadjuvant bevacizumab with standard chemoradiation and surgery shows promising long-term efficacy and safety profiles in locally advanced rectal cancer patients.

Do the dietary supplements epigallocatechin gallate or vitamin e cause a radiomodifying response on tumors in vivo? A pilot study with murine breast carcinoma.

The primary aim of this study was to evaluate a systematic and reproducible assay to examine the potential radiomodifying effects of vitamin E (VE) or epigallocatechin gallate (EGCG), antioxidants commonly consumed by cancer patients as dietary supplements, on tumor control. C3H mice were randomized to a control diet or to the control diet supplemented with VE or EGCG. A tumor control dose 50% (TCD(50)) assay was used to evaluate for a radiomodifying response in stage IV murine cancer (MCa-IV) tumors, implanted in the hindleg of mice, and allowed to grow to 8 mm before receiving a single dose of radiation. The effects of VE and EGCG on intratumoral angiogenesis and apoptosis were evaluated in a group of nonirradiated mice using immunohistochemical staining. Cell proliferation assays were conducted on MCa-IV tumors in vitro. EGCG slowed tumor growth rate by 10%. EGCG and VE slowed tumor regrowth by 24 to 25%. There were no significant differences in TCD(50) values between the groups (control = 73.9 Gy, VE = 77.2 Gy, EGCG = 76.4 Gy); however, normal tissues were protected from late radiation effects (autoamputations) in the VE group. VE and EGCG increased tumor cell apoptosis and decreased tumor cell proliferation but had no effect on microvessel density. In this pilot study, neither VE nor EGCG exerted a significant radiomodifying effect on the MCa-IV tumor. Nonetheless, the suggestion of a small degree of tumor radioprotection by these antioxidant compounds warrants further research. As supplementation with VE radioprotected normal tissue, additional studies on this putative benefit are recommended.