Lik-Cheung Cheng

The EML4-ALK fusion gene is involved in various histologic types of lung cancers from nonsmokers with wild-type EGFR and KRAS

The echinoderm microtubule‐associated protein‐like 4‐anaplastic lymphoma kinase (EML4‐ALK) fusion gene resulting from the chromosome inversion inv(2)(p21;p23) recently was identified in nonsmall cell lung cancer (NSCLC). The authors of this study investigated the frequency, genetic and clinicopathologic profiles of EML4‐ALK in Chinese patients with NSCLC.

A novel KIF5B-ALK variant in nonsmall cell lung cancer

The anaplastic lymphoma kinase (ALK) gene is involved frequently in chromosomal translocations, resulting in fusion genes with different partners found in various lymphoproliferative conditions. It was recently reported in nonsmall cell lung cancer (NSCLC) that the fusion protein encoded by echinoderm microtubule‐associated protein‐like 4‐ALK (EML4‐ALK) fusion gene conferred oncogenic properties. The objective of the current study was to identify other possible ALK fusion genes in NSCLC.

Acacetin, a Natural Flavone, Selectively Inhibits Human Atrial Repolarization Potassium Currents and Prevents Atrial Fibrillation in Dogs

Background— The development of atrium-selective antiarrhythmic agents is a current strategy for inhibiting atrial fibrillation (AF). The present study investigated whether the natural flavone acacetin from the traditional Chinese medicine Xuelianhua would be an atrium-selective anti-AF agent. Methods and Results— The effects of acacetin on human atrial ultrarapid delayed rectifier K+ current (IKur) and other cardiac ionic currents were studied with a whole-cell patch technique. Acacetin suppressed IKur and the transient outward K+ current (IC50 3.2 and 9.2 &mgr;mol/L, respectively) and prolonged action potential duration in human atrial myocytes. The compound blocked the acetylcholine-activated K+ current; however, it had no effect on the Na+ current, L-type Ca2+ current, or inward-rectifier K+ current in guinea pig cardiac myocytes. Although acacetin caused a weak reduction in the hERG and hKCNQ1/hKCNE1 channels stably expressed in HEK 293 cells, it did not prolong the corrected QT interval in rabbit hearts. In anesthetized dogs, acacetin (5 mg/kg) prolonged the atrial effective refractory period in both the right and left atria 1 to 4 hours after intraduodenal administration without prolongation of the corrected QT interval, whereas sotalol at 5 mg/kg prolonged both the atrial effective refractory period and the corrected QT interval. Acacetin prevented AF induction at doses of 2.5 mg/kg (50%), 5 mg/kg (85.7%), and 10 mg/kg (85.7%). Sotalol 5 mg/kg also prevented AF induction (60%). Conclusions— The present study demonstrates that the natural compound acacetin is an atrium-selective agent that prolongs the atrial effective refractory period without prolonging the corrected QT interval and effectively prevents AF in anesthetized dogs after intraduodenal administration. These results indicate that oral acacetin is a promising atrium-selective agent for the treatment of AF.

Double EGFR mutants containing rare EGFR mutant types show reduced in vitro response to gefitinib compared with common activating missense mutations

Epidermal growth factor receptor (EGFR) mutations are common in lung adenocarcinomas, especially from nonsmoking women of Asian descent. We have previously shown EGFR mutations occur in >70% of lung adenocarcinoma from nonsmokers in our population with a complex mutational profile, including 13% of EGFR double mutations. In this study, we investigated the in vitro gefitinib response of four EGFR double mutants identified in untreated patients, including Q787R+L858R, E709A+G719C, T790M+L858R, and H870R+L858R. The phosphorylation profiles of EGFR and downstream effectors AKT, STAT3/5, and ERK1/2 were compared by immunoblot analyses among the single and double mutants transfected into H358 cells. Results showed that mutants responded to in vitro gefitinib treatment with different sensitivities. The G719C and L858R single mutants showed the highest gefitinib sensitivity compared with the corresponding coexisting single mutants E709A, Q787R, H870R, and T790M. The double mutants E709A+G719C, Q787R+L858R, and H870R+L858R showed attenuated responses to gefitinib in the EGFR and downstream effector phosphorylation profiles compared with G719C or L858R alone. T790M+L858R showed strong resistance to gefitinib. Clinically, the patient whose tumor contained H870R+L858R showed tumor stabilization by 250 mg oral gefitinib daily but cerebral metastasis developed 6 months later. Correlation with the in vitro phosphorylation profile of H870R+L858R suggested that treatment failure was probably due to inadequate suppression of EGFR signaling by the drug level attainable in the cerebrospinal fluid at the given oral dosage. Overall, the findings suggested that rare types of EGFR substitution mutations could confer relative gefitinib resistance when combined with the common activating mutants. [Mol Cancer Ther 2009;8(8):2142–51]

Src Promotes Survival and Invasion of Lung Cancers with Epidermal Growth Factor Receptor Abnormalities and Is a Potential Candidate for Molecular-Targeted Therapy

Molecular-targeted therapy using tyrosine kinase inhibitors against epidermal growth factor receptor (EGFR) is an effective therapy for non–small cell lung cancer that harbor EGFR mutations. This study aimed to investigate the role of Src, a close EGFR associator, as a drug target in NSCLC cells with different EGFR genomic statuses. Src inhibition was achieved using 4-(4′-Phenoxyanilino)-6,7-dimethoxyquinazolinee (SKI-1) and the specificity of action was verified by RNA interference. The results showed that SKI-1 induced significant apoptosis in a dose-dependent manner in cancer cells with high basal Src activation. Activation of FAK and p130Cas was involved in Src-mediated invasion in SKI-1–sensitive cells. SKI-1 inhibited phosphorylation of EGFR as well as EGFR downstream effectors, such as signal transducers and activators of transcription 3/5, extracellular signal-regulated kinase 1/2 and AKT in the mutant cells but not the wild-type cells. This inhibition profile of EGFR implicates that induction of apoptosis and sensitivity of mutant cells to SKI treatment is mediated by EGFR and EGFR downstream pathways. Cotreatment with SKI-1 and gefitinib enhanced apoptosis in cancer cells that contained EGFR mutation and/or amplification. SKI-1 treatment alone induced significant apoptosis in H1975 cells known to be resistant to gefitinib. Src phosphorylation was shown by immunohistochemistry in around 30% of primary lung carcinomas. In 152 adenocarcinomas studied, p-Src was associated with EGFR mutations (P = 0.029). Overall, the findings indicated that Src could be a useful target for treatment of non–small cell lung cancer. Besides EGFR genomic mutations, other forms of EGFR and related family member abnormalities such as EGFR amplification might enhance SKI sensitivity. (Mol Cancer Res 2009;7(6):923–32)

Evidence for functional expression of TRPM7 channels in human atrial myocytes

Transient receptor potential melastatin-7 (TRPM7) channels have been recently reported in human atrial fibroblasts and are believed to mediate fibrogenesis in human atrial fibrillation. The present study investigates whether TRPM7 channels are expressed in human atrial myocytes using whole-cell patch voltage-clamp, RT-PCR and Western blotting analysis. It was found that a gradually activated TRPM7-like current was recorded with a K+- and Mg2+-free pipette solution in human atrial myocytes. The current was enhanced by removing extracellular Ca2+ and Mg2+, and the current increase could be inhibited by Ni2+ or Ba2+. The TRPM7-like current was potentiated by acidic pH and inhibited by La3+ and 2-aminoethoxydiphenyl borate. In addition, Ca2+-activated TRPM4-like current was recorded in human atrial myocytes with the addition of the Ca2+ ionophore A23187 in bath solution. RT-PCR and Western immunoblot analysis revealed that in addition to TRPM4, TRPM7 channel current, mRNA and protein expression were evident in human atrial myocytes. Interestingly, TRPM7 channel protein, but not TRPM4 channel protein, was significantly increased in human atrial specimens from the patients with atrial fibrillation. Our results demonstrate for the first time that functional TRPM7 channels are present in human atrial myocytes, and the channel expression is upregulated in the atria with atrial fibrillation.

Management of symptomatic congenital tracheal stenosis in neonates and infants by slide tracheoplasty: a 7-year single institution experience.

OBJECTIVE Congenital tracheal stenosis (CTS) is rare. When it presents early in life, its management can be challenging. Of the surgical techniques that have been devised to correct long-segment CTS, slide tracheoplasty (ST) appears to be superior. We present our 7-year-experience of ST for the treatment of symptomatic CTS in neonates and infants. METHODS The hospital records of all 14 neonates and infants who underwent ST between 2001 and 2008 at our hospital were retrieved. Patient characteristics, trachea morphology, co-existing anomalies, operative procedures, techniques, outcomes and clinical courses were reviewed. RESULTS Patients underwent ST at age 4 days to 22 months (median: 2.4 months). Five (36%) required intermittent ventilator support prior to surgery. All ST was done under cardiopulmonary bypass. The associated cardiovascular anomalies in 10 infants (71%) were also corrected at the same operation. All survived the initial surgical procedures. The in-hospital mortality for the group was 14.3%. The median periods of postoperative intensive care unit and hospital stay of the 12 children, who were successfully extubated within 7 postoperative days, were 9 days (range: 6-28 days) and 28 days (range: 14-375 days), respectively. Follow-up of all 12 midterm survivors was complete, ranging from 5 months to 5.6 years (median: 40 months). A total of four patients had been found to have tracheobronchial malacia postoperatively and were managed with stenting. Of the remaining 10 survivors who had no residual or recurrent airway problems, and no cardiovascular residuum or sequela, two had gross developmental delay. CONCLUSIONS In the management of symptomatic infants with CTS, our limited experience suggests that meticulously performed ST together with vigilant pre- and postoperative care can provide satisfactory short and midterm solution to the airway problem. Some incidental residual clinical problems appear to be unavoidable.

Modulation of human cardiac transient outward potassium current by EGFR tyrosine kinase and Src-family kinases

AIMS The human cardiac transient outward K(+) current I(to) (encoded by Kv4.3 or KCND3) plays an important role in phase 1 rapid repolarization of cardiac action potentials in the heart. However, modulation of I(to) by intracellular signal transduction is not fully understood. The present study was therefore designed to determine whether/how human atrial I(to) and hKv4.3 channels stably expressed in HEK 293 cells are regulated by protein tyrosine kinases (PTKs). METHODS AND RESULTS Whole-cell patch voltage-clamp, immunoprecipitation, western blotting, and site-directed mutagenesis approaches were employed in the present study. We found that human atrial I(to) was inhibited by the broad-spectrum PTK inhibitor genistein, the selective epidermal growth factor receptor (EGFR) kinase inhibitor AG556, and the Src-family kinases inhibitor PP2. The inhibitory effect was countered by the protein tyrosine phosphatase inhibitor orthovanadate. In HEK 293 cells stably expressing human KCND3, genistein, AG556, and PP2 significantly reduced the hKv4.3 current, and the reduction was antagonized by orthovanadate. Interestingly, orthovanadate also reversed the reduced tyrosine phosphorylation level of hKv4.3 channels by genistein, AG556, or PP2. Mutagenesis revealed that the hKv4.3 mutant Y136F lost the inhibitory response to AG556, while Y108F lost response to PP2. The double-mutant Y108F-Y136F hKv4.3 channels showed no response to either AG556 or PP2. CONCLUSION Our results demonstrate that human atrial I(to) and cloned hKv4.3 channels are modulated by EGFR kinase via phosphorylation of the Y136 residue and by Src-family kinases via phosphorylation of the Y108 residue; tyrosine phosphorylation of the channel may be involved in regulating cardiac electrophysiology.

Artificial chordae: A simple clip and tie technique

1. Ilbawi AM, Spicer DE, Bharati S, Cook A, Anderson RH. Morphologic study of the ascending aorta and aortic arch in hypoplastic left hearts: surgical implications. J Thorac Cardiovasc Surg. 2007;134:99-105. 2. Mahle WT, Rychik J, Weinberg PM, Cohen MS. Growth characteristics of the aortic arch after the Norwood operation. J Am Coll Cardiol. 1998;32:1951-4. 3. Forbess JM, Cook N, Roth SJ, Serraf A, Mayer JE Jr, Jonas RA. Ten-year institutional experience with palliative surgery for hypoplastic left heart syndrome. Risk factors related to stage I mortality. Circulation. 1995; 92(9 Suppl):II262-6. FIGURE 1. A, Intraoperative image showing diminutive aorta. B, Angiogram revealing narrowed ostium to native ascending aorta.

Functional transient receptor potential canonical type 1 channels in human atrial myocytes

Transient receptor potential (TRP) channels are not well understood in human atrium, and the present study was therefore designed to investigate whether TRPC channels would mediate the nonselective cation current reported previously and are involved in the formation of store-operated Ca2+ entry (SOCE) channels in human atrial myocytes using approaches of whole-cell patch voltage-clamp, RT-PCR, Western blotting, co-immunoprecipitation, and confocal scanning approaches, etc. We found that a nonselective cation current was recorded under K+-free conditions in human atrial myocytes, and the current was inhibited by the TRP channel blocker La3+. Thapsigargin enhanced the current, and its effect was suppressed by La3+ and prevented by pipette inclusion of anti-TRPC1 antibody. Endothlin-1 and angiotensin II enhanced the current that could be inhibited by La3+. Gene and protein expression of TRPC1 channels were abundant in human atria. In addition, mRNA and protein of STIM1 and Orai1, components of SOCE channels, were abundantly expressed in human atria. Co-immunoprecipitation analysis demonstrated an interaction of TRPC1 with STIM1 and/or Orai1. Ca2+ signaling mediated by SOCE channels was detected by a confocal microscopy technique. These results demonstrate the novel evidence that TRPC1 channels not only mediate the nonselective cation current, but also form SOCE channels in human atria as a component. TRPC1 channels can be activated by endothelin-1 or angiotensin II, which may be involved in the atrial electrical remodeling in patients with atrial fibrillation.