Exploring ALK fusion in colorectal cancer: a case series and comprehensive analysis.

Anaplastic lymphoma kinase (ALK) fusion-positive colorectal cancer (CRC) is a rare and chemotherapy-refractory subtype that lacks established and effective treatment strategies. Additionally, the efficacy and safety of ALK inhibitors (ALKi) in CRC remain undetermined. Herein, we examined a series of ALK-positive CRC patients who underwent various lines of ALKi treatment. Notably, we detected an ALK 1196M resistance mutation in a CRC patient who received multiple lines of chemotherapy and ALKi treatment. Importantly, we found that Brigatinib and Lorlatinib demonstrated some efficacy in managing this patient, although the observed effectiveness was not as pronounced as in non-small cell lung cancer cases. Furthermore, based on our preliminary analyses, we surmise that ALK-positive CRC patients are likely to exhibit inner resistance to Cetuximab. Taken together, our findings have important implications for the treatment of ALK-positive CRC patients.

A Temporal PROTAC Cocktail-Mediated Sequential Degradation of AURKA Abrogates Acute Myeloid Leukemia Stem Cells.

AURKA is a potential kinase target in various malignancies. The kinase-independent oncogenic functions partially disclose the inadequate efficacy of the kinase inhibitor in a Phase III clinical trial. Simultaneously targeting the catalytic and noncatalytic functions of AURKA may be a feasible approach. Here, a set of AURKA proteolysis targeting chimeras (PROTACs) are developed. The CRBN-based dAurA383 preferentially degrades the highly abundant mitotic AURKA, while cIAP-based dAurA450 degrades the lowly abundant interphase AURKA in acute myeloid leukemia (AML) cells. The proteomic and transcriptomic analyses indicate that dAurA383 triggers the "mitotic cell cycle" and "stem cell" processes, while dAurA450 inhibits the "MYC/E2F targets" and "stem cell" processes. dAurA383 and dAurA450 are combined as a PROTAC cocktail. The cocktail effectively degrades AURKA, relieves the hook effect, and synergistically inhibits AML stem cells. Furthermore, the PROTAC cocktail induces AML regression in a xenograft mouse model and primary patient blasts. These findings establish the PROTAC cocktail as a promising spatial-temporal drug administration strategy to sequentially eliminate the multifaceted functions of oncoproteins, relieve the hook effect, and prevent cancer stem cell-mediated drug resistance.

Arabidopsis ETHYLENE INSENSITIVE 3 directly regulates the expression of PG1ß-like family genes in response to aluminum stress.

The genes in the subfamily PG1ß (beta subunit of poly-galacturonase isoenzyme 1) have a clear effect on the biosynthesis pathway of pectin, a main component of the cell wall. However, the detailed functions of the PG1ß-like gene members in Arabidopsis (AtPG1-3) have not yet been determined. In this study, we investigated their functional roles in response to aluminum (Al) stress. Our results indicate that the PG1ß-like gene members are indeed involved in the Al-stress response and they can modulate its accumulation in roots to achieve optimum root elongation and hence better seedling growth. We found that transcription factor EIN3 (ETHYLENE INSENSITIVE 3) alters pectin metabolism and the EIN3 gene responds to Al stress to affect the pectin content in the root cell walls, leading to exacerbation of the inhibition of root growth, as reflected by the phenotypes of overexpressing lines. We determined that EIN3 can directly bind to the promoter regions of PG1-3, which act downstream of EIN3. Thus, our results show that EIN3 responds to Al stress in Arabidopsis directly through regulating the expression of PG1-3. Hence, EIN3 mediates their functions by acting as a biomarker in their molecular biosynthesis pathways, and consequently orchestrates their biological network in response to Al stress.

Efficacy of cystectasia in the treatment of ketamine-induced bladder contracture.

BACKGROUND: The treatment of ketamine-induced bladder contractures remains poorly studied. We therefore evaluated the efficacy of cystectasia with a sodium hyaluronate balanced solution in this kind of bladder contracture. METHODS: Eighteen patients presenting with ketamine-induced bladder contracture between July 2010 and February 2018 were selected and analysed. Ketamine was discontinued in all patients, who were then treated with weekly cystectasia (0.09% sodium hyaluronate balanced solution) 3 times. The volume of the first perfusion was twice the preoperatively measured bladder capacity, and the volume of the subsequent two perfusions was increased by 100 mL each time. The Pelvic Pain and Urgency/Frequency (PUF) symptom score, O'Leary-Sant Interstitial Cystitis (IC) Symptom Index (ICSI), IC Problem Index (ICPI), Quality of Life (QOL) score, and bladder capacity were recorded before surgery and 3 and 12 months after the 3rd expansion. RESULTS: No significant complications were observed during the 3 expansions. Fourteen patients completed the full follow-up schedule. Preoperatively and at the 3- and 12-month follow-up evaluations performed after the 3rd expansion, the PUF symptom scores were 20.4±3.6, 11.5±3.1, and 13.2±3.3, respectively; the mean ICSI was 13.6±2.8, 7.7±2.3, and 8.2±2.5, respectively; the mean ICPI was 10.6±2.6, 7.3±2.1, and 7.7±2.5, respectively; and the mean QOL scores were 6.0±0, 2.1±0.5, and 2.7±0.8, respectively; and the mean bladder catheter volume was 83±27, 234±56, and 228±52 mL, respectively. There were significant differences between all preoperative and postoperative values. CONCLUSIONS: Cystectasia with a sodium hyaluronate balanced solution is an effective treatment modality for ketamine-induced bladder contracture.

Ethylene insensitive3-like2 (OsEIL2) confers stress sensitivity by regulating OsBURP16, the ß subunit of polygalacturonase (PG1ß-like) subfamily gene in rice.

The transcription factors EIN3 (ETHYLENE-INSENSITIVE 3) and EILs (EIN3-Likes) play important roles in plant development and defense responses; however, their mechanism in these processes remain unclear. Here, we report that OsEIL2, an EIN3-like transcription factor from rice (Oryza sativa), plays important roles in abiotic stress and leaf senescence. OsEIL2 is a nuclear-localized protein with transactivation activity in the C-terminus (amino acids 344-583) and can be induced by NaCl, polyethylene glycol (PEG), dark, and abscisic acid (ABA) treatment. Transgenic plants of overexpressing OsEIL2 (OsEIL2-OX) show reduced tolerance to salt and drought stress compared with the controls. While the transgenic plants of overexpressing OsEIL2-RNA interference (OsEIL2-RNAi) exhibit enhanced tolerance to salt and drought stress compared with the controls. Moreover, seedlings of OsEIL2-overexpressing transgenic plants exhibit delayed leaf development and an accelerated dark-induced senescence phenotype, whereas OsEIL2-RNAi plants display the opposite phenotype. We further found that OsEIL2 functions upstream of OsBURP14 and OsBURP16. OsBURP14 and OsBURP16 are the members of the ß subunit of polygalacturonase subfamilies. OsBURP16 overexpression reduced pectin content and cell adhesion and increased abiotic stress sensitivity in rice. OsEIL2 binds directly to the promoter of OsBURP14 and OsBURP16 and activates their transcript levels. We also found that OsEIL2 overexpression decreased the pectin content by increasing polygalacturonase (PG) activity. Taken together, these results revealed a new mechanism of OsEIL2 in abiotic stress responses. These findings provide new insights into plant resistance to abiotic stress.

Strained cyclooctyne as a molecular platform for construction of multimodal imaging probes.

Small-molecule-based multimodal and multifunctional imaging probes play prominent roles in biomedical research and have high clinical translation ability. A novel multimodal imaging platform using base-catalyzed double addition of thiols to a strained internal alkyne such as bicyclo[6.1.0]nonyne has been established in this study, thus allowing highly selective assembly of various functional units in a protecting-group-free manner. Using this molecular platform, novel dual-modality (PET and NIRF) uPAR-targeted imaging probe: (64)Cu-CHS1 was prepared and evaluated in U87MG cells and tumor-bearing mice models. The excellent PET/NIRF imaging characteristics such as good tumor uptake (3.69%ID/g at 2 h post-injection), high tumor contrast, and specificity were achieved in the small-animal models. These attractive imaging properties make (64)Cu-CHS1 a promising probe for clinical use.

HMDO-promoted peptide and protein synthesis in ionic liquids.

Hexamethyldisiloxane (HMDO) has been developed to efficiently promote the metal-free direct coupling of an amino function of one cysteine-free peptide or protein and a C-terminal thioester of the second peptide in ionic liquids. The amide-coupling reaction proceeds smoothly under mild conditions to afford the corresponding products in good to excellent yields (63-94%). Peptide couplings were also achieved using in-situ-generated thioesters by the thioesterification of oxo esters.

Targeted delivery of doxorubicin through conjugation with EGF receptor-binding peptide overcomes drug resistance in human colon cancer cells.

BACKGROUND AND PURPOSE: Induction of multidrug resistance by doxorubicin (DOX), together with non-specific toxicities, has restricted DOX-based chemotherapy. Recently, we demonstrated that DOX conjugated with an EGF receptor-binding peptide (DOX-EBP) had enhanced anticancer efficacy and reduced systemic toxicity when targeting EGF receptor-overexpressing tumours. Here we investigated whether DOX-EBP is able to overcome drug resistance and the underlying molecular mechanisms. EXPERIMENTAL APPROACH: DOX-resistant SW480/DOX cells were derived from non-resistant SW480 cells by stepwise exposure to increasing concentrations of DOX, and P-glycoprotein overexpression induced by DOX was confirmed by Western blotting. Cytotoxicity and intracellular distribution of drugs were evaluated by MTT assay and fluorescence microscopy respectively. EGF receptor-mediated endocytosis was determined in EGF receptor and endocytosis inhibition assays. Drug accumulation in tumour cells and murine xenografts was determined by HPLC. KEY RESULTS: The cytotoxicity and accumulation of DOX-EBP in SW480/DOX cells were almost the same as in SW480 cells, but those of free DOX were reduced. DOX-EBP accumulation was prevented by inhibitors of both EGF receptors and endocytosis, suggesting EGF receptors mediate endocytotic uptake. Tumour accumulation of DOX-EBP was significantly higher than free DOX in mice, and the levels of DOX-EBP were similar in DOX-resistant and non-resistant tumour tissues. Importantly, DOX-EBP, but not free DOX, was effective at inhibiting solid tumour growth and increased survival rate in both sensitive and resistant models. CONCLUSION AND IMPLICATIONS: DOX-EBP can overcome DOX resistance of tumour cells and increase in vivo antitumour efficacy. Therefore, it has the potential to be a potent therapeutic agent for treating drug-resistant cancers.

Biological evaluation of a novel doxorubicin-peptide conjugate for targeted delivery to EGF receptor-overexpressing tumor cells.

Epidermal growth factor receptor (EGFR) is overexpressed in a variety of epithelial malignancies and thus can be used for EGFR-targeted therapy to improve antitumor efficacy. Therefore we synthesized a novel conjugate of doxorubicin (DOX) with an EGFR-binding peptide (NH2-CMYIEALDKYAC-COOH; EBP) via an ester bond at position 14 of DOX through a glutarate spacer. To confirm that the DOX-EBP conjugate is capable of targeting tumor cells overexpressing EGFR, we compared the cellular accumulation, intracellular distribution and in vitro cytotoxicity of DOX-EBP and free DOX. After treating with equimolar concentration of DOX-EBP or free DOX, the conjugate accumulated at significantly higher levels in EGFR-overexpressing cells than in non-EGFR-overexpressing cells, while the intracellular accumulation of free DOX was almost the same in all the cells. However, the intracellular accumulation of DOX-EBP was significantly reduced in EGFR-overexpressing cells preincubated with inhibitory anti-EGFR monoclonal antibody, demonstrating the involvement of EGFR pathway in the transport of the conjugate. Confocal fluorescence microscopy reveals that the conjugate was distributed in cytoplasmic and perinuclear areas during the first 30 min, whereas the free DOX was accumulated in both cytoplasm and nuclei. After 24 h, however, the DOX signal in the cells treated with DOX-EBP was also distributed in the nuclei, suggesting the release of DOX from the conjugate and entry into the nuclei. Biodistribution and in vivo antitumor experiments, together with in vitro cytotoxicity, indicate that the therapeutic competence of DOX-EBP was due to its increased accumulation in EGFR-expressing tumor cells. Furthermore, the survival of tumor-bearing mice treated with DOX-EBP was significantly higher than that with free DOX. These data demonstrate the enhanced anticancer efficacy and reduced systemic toxicity of DOX-EBP conjugate with targeting ability to EGFR-overexpressing tumor cells.