Dendritic Cell Vaccine Loaded with MG-7 Antigen Induces Cytotoxic T Lymphocyte Responses against Gastric Cancer.

Dendritic cells (DCs) are antigen-presenting cells that can activate T cells and initiate a primary immune response. Personalized DC vaccines have demonstrated a modest antitumor potential in some clinical pilot studies. However, those vaccines are difficult to manufacture and have a limited antitumor response. In this study, a lentiviral vector-programmed DC vaccine with high antitumor responses is developed. By transfecting with a lentiviral vector, the DC vaccine is loaded with MG-7 antigen (MG-7Ag). Three representative gastric cancer cell lines, such as KATO-3, MKN45, and SNU16, are used to estimate the in vitro cytotoxic effect of the MG-7Ag DC vaccine. Furthermore, we examine the in vivo antitumor efficacy of specific cytotoxic T lymphocytes (CTLs) induced by the MG-7Ag DC vaccine in patient-derived xenograft (PDX) mice models. The current data demonstrate that the MG-7Ag DC vaccine induced a potent CTL activity. Those CTLs have a significant cytotoxic effect on both KATO-3 and MKN45 with high level of MG-7 expression. In addition, MG-7Ag DC vaccine-mediated CTLs significantly inhibit the growth of tumor xenografts in nude mice. The MG-7Ag DC vaccine activate the cytotoxic effect of lymphocytes and can be employed as a vaccine in gastric cancer immunotherapy.

Enzalutamide-induced and PTH1R-mediated TGFBR2 degradation in osteoblasts confers resistance in prostate cancer bone metastases.

Enzalutamide resistance has been observed in approximately 50% of patients with prostate cancer (PCa) bone metastases. Therefore, there is an urgent need to investigate the mechanisms and develop strategies to overcome resistance. We observed enzalutamide resistance in bone lesion development induced by PCa cells in mouse models. We found that the bone microenvironment was indispensable for enzalutamide resistance because enzalutamide significantly inhibited the growth of subcutaneous C4-2B tumors and the proliferation of C4-2B cells isolated from the bone lesions, and the resistance was recapitulated only when C4-2B cells were co-cultured with osteoblasts. In revealing how osteoblasts contribute to enzalutamide resistance, we found that enzalutamide decreased TGFBR2 protein expression in osteoblasts, which was supported by clinical data. This decrease was possibly through PTH1R-mediated endocytosis. We showed that PTH1R blockade rescued enzalutamide-mediated decrease in TGFBR2 levels and enzalutamide responses in C4-2B cells that were co-cultured with osteoblasts. This is the first study to reveal the contribution of the bone microenvironment to enzalutamide resistance and identify PTH1R as a feasible target to overcome the resistance in PCa bone metastases.

Loss of Myeloid-Specific TGF-ß Signaling Decreases CTHRC1 to Downregulate bFGF and the Development of H1993-Induced Osteolytic Bone Lesions.

The role of myeloid cell-specific TGF-ß signaling in non-small-cell lung cancer (NSCLC)-induced osteolytic bone lesion development is unknown. We used a genetically engineered mouse model, Tgfbr2LysMCre knockout (KO), which has a loss of TGF-ß signaling specifically in myeloid lineage cells, and we found that the area of H1993 cell-induced osteolytic bone lesions was decreased in Tgfbr2LysMCre KO mice, relative to the area in control littermates. The bone lesion areas were correlated with tumor cell proliferation, angiogenesis, and osteoclastogenesis in the microenvironment. The smaller bone lesion area was partially rescued by bFGF, which was expressed by osteoblasts. Interestingly, bFGF was able to rescue the osteoclastogenesis, but not the tumor cell proliferation or angiogenesis. We then focused on identifying osteoclast factors that regulate bFGF expression in osteoblasts. We found that the expression and secretion of CTHRC1 was downregulated in osteoclasts from Tgfbr2LysMCre KO mice; CTHRC1 was able to promote bFGF expression in osteoblasts, possibly through the Wnt/ß-catenin pathway. Functionally, bFGF stimulated osteoclastogenesis and inhibited osteoblastogenesis, but had no effect on H1993 cell proliferation. On the other hand, CTHRC1 promoted osteoblastogenesis and H1993 cell proliferation. Together, our data show that myeloid-specific TGF-ß signaling promoted osteolytic bone lesion development and bFGF expression in osteoblasts; that osteoclast-secreted CTHRC1 stimulated bFGF expression in osteoblasts in a paracrine manner; and that CTHRC1 and bFGF had different cell-specific functions that contributed to bone lesion development.

TGF-ß receptors: In and beyond TGF-ß signaling.

Transforming growth factor ß (TGF-ß) plays an important role in normal development and homeostasis. Dysregulation of TGF-ß responsiveness and its downstream signaling pathways contribute to many diseases, including cancer initiation, progression, and metastasis. TGF-ß ligands bind to three isoforms of the TGF-ß receptor (TGFBR) with different affinities. TGFBR1 and 2 are both serine/threonine and tyrosine kinases, but TGFBR3 does not have any kinase activity. They are necessary for activating canonical or noncanonical signaling pathways, as well as for regulating the activation of other signaling pathways. Another prominent feature of TGF-ß signaling is its context-dependent effects, temporally and spatially. The diverse effects and context dependency are either achieved by fine-tuning the downstream components or by regulating the expressions and activities of the ligands or receptors. Focusing on the receptors in events in and beyond TGF-ß signaling, we review the membrane trafficking of TGFBRs, the kinase activity of TGFBR1 and 2, the direct interactions between TGFBR2 and other receptors, and the novel roles of TGFBR3.

Mechanisms and Approaches for Overcoming Enzalutamide Resistance in Prostate Cancer.

Enzalutamide, a second-generation small-molecule inhibitor of the androgen receptor (AR), has been approved for patients who failed with androgen deprivation therapy and have developed castration-resistant prostate cancer. More than 80% of these patients develop bone metastases. The binding of enzalutamide to the AR prevents the nuclear translocation of the receptor, thus inactivating androgen signaling. However, prostate cancer cells eventually develop resistance to enzalutamide treatment. Studies have found resistance both in patients and in laboratory models. The mechanisms of and approaches to overcoming such resistance are significant issues that need to be addressed. In this review, we focus on the major mechanisms of acquired enzalutamide resistance, including genetic mutations and splice variants of the AR, signaling pathways that bypass androgen signaling, intratumoral androgen biosynthesis by prostate tumor cells, lineage plasticity, and contributions from the tumor microenvironment. Approaches for overcoming these mechanisms to enzalutamide resistance along with the associated problems and solutions are discussed. Emerging questions, concerns, and new opportunities in studying enzalutamide resistance will be addressed as well.

Design, synthesis and pharmacological evaluation of ALK and Hsp90 dual inhibitors bearing resorcinol and 2,4-diaminopyrimidine motifs.

Rather than by directly focusing on the ever-changing ALK mutants, here we report an alternative strategy to overcome the drug resistance caused by treatment of ALK inhibitors by developing ALK and Hsp90 dual targeting inhibitors. Since Hsp90 is a molecular chaperone that regulates the maturation, activation and stability of numerous "client proteins" including ALK, dual targeting ALK and Hsp90 may bring more benefits and efficacy against drug resistance of ALK inhibitors. By using our previously developed ALK inhibitor 6 and the clinical Hsp90 inhibitors AUY922 or AT13387 as the templates, we developed several series of resorcinol tethered 2,4-diaminopyrimidines as ALK/Hsp90 dual inhibitors bearing various linkers at different linking sites. Compound 10h and 10j showed high potency against ALK (17.3 vs 9.8 nM) and Hsp90α (100 vs 40 nM). They also have high potency against ALK resistant mutants, especially the gatekeeper mutation ALKL1196M. Both compounds showed strong antiproliferative activity against the ALK-addictive H3122 cells (11 vs 13 nM). The dual functioning mechanism is further confirmed by their down-regulation of the Hsp90 clients ALK and AKT, and up-regulation of the chaperone protein Hsp70 in H3122 cells.

Design, Synthesis, and Biological Evaluation of the First c-Met/HDAC Inhibitors Based on Pyridazinone Derivatives.

Simultaneous blockade of more than one pathway is considered to be a promising approach to overcome the low efficacy and acquired resistance of cancer therapies. Thus, a novel series of c-Met/HDAC bifunctional inhibitors was designed and synthesized by merging pharmacophores of c-Met and HDAC inhibitors. The most potent compound, 2m, inhibited c-Met kinase and HDAC1, with IC50 values of 0.71 and 38 nM, respectively, and showed efficient antiproliferative activities against both EBC-1 and HCT-116 cells with greater potency than the reference drug Chidamide. Western blot analysis revealed that compound 2m inhibited phosphorylation of c-Met and c-Met downstream signaling proteins and increased expression of Ac-H3 and p21 in EBC-1 cells in a dose-dependent manner. Our study presents novel compounds for the further exploration of dual c-Met/HDAC pathway inhibition achieved with a single molecule.

Inhibition of autophagy aggravated 4-nitrophenol-induced oxidative stress and apoptosis in NHPrE1 human normal prostate epithelial progenitor cells.

4-Nitrophenol (PNP), a well-established human carcinogen, has been proven to have detrimental effects on reproductive system of male rats in previous studies. The molecular mechanisms involved PNP-induced damage remain to be established. Autophagy can exert protective effects on various cytotoxic factors that induce injury. In the present study, we aim to investigate whether autophagy is induced by PNP and the function of autophagy in PNP-induced injury in NHPrE1, a normal human prostate epithelial progenitor cell line. Our results indicate that PNP induced oxidative stress as evidenced by increased MDA levels and decreased activity of SOD and GSH-Px. PNP also increased apoptosis of NHPrE1 cells as evidenced by western blot and Hoechst 33258 staining and activated autophagy in NHPrE1 cells detected by RT-PCR and western blot. Inhibition of autophagy by 3-MA further increased PNP-induced oxidative stress and apoptosis of NHPrE1 cells. We also found that PNP-induced apoptosis was suppressed by N-acetylcysteine, suggesting oxidative stress may play an important role in PNP cytotoxicity. Furthermore, phosphorylation of mTOR protein was inhibited by PNP, indicating that PNP might induce autophagy in NHPrE1 cells via inhibiting mTOR pathway. In conclusion, these results suggest that activation of autophagy should play a protective role in PNP-induced oxidative stress and apoptosis of NHPrE1 cells, which might be mediated through mTOR pathway.

Prolonged Hoarseness Caused by Arytenoid Dislocation After Anterior Cervical Corpectomy and Fusion.

STUDY DESIGN: A case of arytenoid dislocation after anterior cervical corpectomy and fusion (ACCF) is reported. OBJECTIVE: To emphasize that arytenoid dislocation could be a possible cause of prolonged hoarseness in patients after ACCF. SUMMARY OF BACKGROUND: Prolonged hoarseness is a common postoperative complication of cervical surgeries, especially in the anterior approach. Postoperative hoarseness is usually associated with paresis of the recurrent laryngeal nerve (RLN). However, other causes such as arytenoids dislocation, which is often misdiagnosed as RLN palsy, should not be ignored either. METHODS: We reported one case of arytenoid dislocation after ACCF and reviewed the related literatures. RESULTS: One patient treated with ACCF experienced prolonged postoperative hoarseness. Arytenoid dislocation was confirmed by laryngoscopy examination and three-dimensional computed tomography (CT) scan. To deal with the problem, a closed reduction of cricoarytenoid joint was performed under general anesthesia. Fortunately, the motion of vocal fold became nearly back to normal after surgery and the patient recovered uneventfully. He was satisfied with the clinical outcome at the final follow-up. CONCLUSION: Arytenoid dislocation should never be ignored in the differential diagnosis of prolonged postoperative hoarseness after ACCF. This situation can be confirmed by CT scan, vocal cord electromyography (EMG), fiberoptic laryngoscopy, or strobovideolaryngoscopy. Once the diagnosis is established, appropriate treatment should be considered immediately. LEVEL OF EVIDENCE: 3.