Monitoring epidermal growth factor receptor C797S mutation in Japanese non-small cell lung cancer patients with serial cell-free DNA evaluation using digital droplet PCR.

Osimertinib is a third-generation epidermal growth factor receptor tyrosine kinase inhibitor (EGFR-TKI) that is effective in treating both naïve and T790M-mutated EGFR-TKI-resistant non-small cell lung cancer patients. The EGFR C797S mutation is the major osimertinib resistance mechanism. The present study monitored the EGFR C797S mutation during osimertinib treatment in Japanese patients using droplet digital PCR (ddPCR). In our first cohort, C797S detection was validated with tumor specimens and/or plasma samples from 26 patients using ddPCR with custom-designed probes detecting and discriminating T790M and C797S in cis and trans positions. In our second cohort, 18 patients with EGFR-T790M who were going to start osimertinib were analyzed using ddPCR by collecting the plasma samples every month from the beginning of the course of osimertinib. In the first cohort, C797S was detected in 15.4% of patients. C797S and T790M in cis and trans positions were distinguished using ddPCR. In the second cohort, serial cfDNA evaluation revealed that the rate of EGFR mutation changes with disease state. Increases of EGFR mutation were detected, including C797S several months before the diagnosis of disease progression. As with the first cohort, C797S and T790M in cis and trans position were distinguished by ddPCR at disease progression. Coincidentally, in the first cohort, next generation sequencing detected NRAS Q61K mutation and the resistance with NRAS Q61K mutation was overcome by trametinib. In the second cohort, serial cfDNA analysis was useful for evaluating bone oligo-progression and local radiation therapy.

Alcohol Dehydrogenase Activity Converts 3″-Hydroxy-geranylhydroquinone to an Aldehyde Intermediate for Shikonin and Benzoquinone Derivatives in Lithospermum erythrorhizon.

Shikonin derivatives are red naphthoquinone pigments produced by several boraginaceous plants, such as Lithospermum erythrorhizon. These compounds are biosynthesized from p-hydroxybenzoic acid and geranyl diphosphate. The coupling reaction that yields m-geranyl-p-hydroxybenzoic acid has been actively characterized, but little is known about later biosynthetic reactions. Although 3″-hydroxy-geranylhydroquinone produced from geranylhydroquinone by CYP76B74 has been regarded as an intermediate of shikonin derivatives, the next intermediate has not yet been identified. This study describes a novel alcohol dehydrogenase activity in L. erythrorhizon cell cultures. This enzyme was shown to oxidize the 3″-alcoholic group of (Z)-3″-hydroxy-geranylhydroquinone to an aldehyde moiety concomitant with the isomerization at the C2'-C3' double bond from the Z-form to the E-form. An enzyme oxidizing this substrate was not detected in other plant cell cultures, suggesting that this enzyme is specific to L. erythrorhizon. The reaction product, (E)-3″-oxo-geranylhydroquinone, was further converted to deoxyshikonofuran, another meroterpenoid metabolite produced in L. erythrorhizon cells. Although nonenzymatic cyclization occurred slowly, it was more efficient in the presence of crude enzymes of L. erythrorhizon cells. This activity was detected in both shikonin-producing and nonproducing cells, suggesting that the aldehyde intermediate at the biosynthetic branch point between naphthalene and benzo/hydroquinone ring formation likely constitutes a key common intermediate in the synthesis of shikonin and benzoquinone products, respectively.

Drug resistance mechanisms in Japanese anaplastic lymphoma kinase-positive non-small cell lung cancer and the clinical responses based on the resistant mechanisms.

The treatment for anaplastic lymphoma kinase (ALK)-positive lung cancer has been rapidly evolving since the introduction of several ALK tyrosine kinase inhibitors (ALK-TKI) in clinical practice. However, the acquired resistance to these drugs has become an important issue. In this study, we collected a total of 112 serial biopsy samples from 32 patients with ALK-positive lung cancer during multiple ALK-TKI treatments to reveal the resistance mechanisms to ALK-TKI. Among 32 patients, 24 patients received more than two ALK-TKI. Secondary mutations were observed in 8 of 12 specimens after crizotinib failure (G1202R, G1269A, I1171T, L1196M, C1156Y and F1245V). After alectinib failure, G1202R and I1171N mutations were detected in 7 of 15 specimens. G1202R, F1174V and G1202R, and P-gp overexpression were observed in 3 of 7 samples after ceritinib treatment. L1196M + G1202R, a compound mutation, was detected in 1 specimen after lorlatinib treatment. ALK-TKI treatment duration was longer in the on-target treatment group than that in the off-target group (13.0 vs 1.2 months). In conclusion, resistance to ALK-TKI based on secondary mutation in this study was similar to that in previous reports, except for crizotinib resistance. Understanding the appropriate treatment matching resistance mechanisms contributes to the efficacy of multiple ALK-TKI treatment strategies.

Overcoming resistance by ALK compound mutation (I1171S + G1269A) after sequential treatment of multiple ALK inhibitors in non-small cell lung cancer.

BACKGROUND: Anaplastic lymphoma kinase (ALK) fusion genes are found in 3%-5% of non-small cell lung cancers (NSCLCs). ALK inhibitors show a very high response rate to ALK-positive NSCLCs. However, the emergence of acquired resistance is inevitable. In this study, we investigated the drugs for overcoming resistance especially compound mutations after sequential treatment with crizotinib, alectinib, and lorlatinib. METHOD: Next-generation sequencing (NGS) and Sanger sequencing were performed on a liver biopsy tissue obtained from a clinical case. Ba/F3 cells in which mutant EML4-ALK were overexpressed were prepared, and cell viability assay and immunoblotting were performed to check the sensitivity of five independent ALK inhibitors. RESULTS: I1171S + G1269A double mutation was identified by NGS and Sanger sequencing on a liver biopsy tissue from a patient who relapsed on lorlatinib treatment. Ceritinib and brigatinib-but not other ALK inhibitors-were active against the compound mutations in the cell line model. CONCLUSIONS: With the sequential ALK inhibitors treatment, cancer cells accumulate new mutations in addition to mutations acquired previously. The identified compound mutation (I1171S + G1269A) was found to be sensitive to ceritinib and brigatinib, and indeed the patient's tumor partially responded to ceritinib. KEY POINTS: ALK compound mutation was found in a clinical sample that was resistant to lorlatinib after sequential ALK-tyrosine kinase inhibitor (TKI) treatment. Ceritinib and brigatinib are potential overcoming drugs against ALK I1171S + G1269A double mutation.

Paragonimus westermani infection mimicking recurrent lung cancer: A case report.

Herein, we report a case of Paragonimus westermani infection, which required differentiation from recurrent lung cancer. A 66-year old Japanese man with a history of lung cancer who had undergone a lobectomy was referred to our clinic for treatment of cough, sputum, dyspnea, and a right pulmonary nodule. He had previously eaten seafood he visited China. P. westermani infection was confirmed by the presence of antibody against P. westermani antigen in the patient's serum and eggs in his sputum. Eventually, molecular identification by PCR-restriction fragment length polymorphism analysis and sequencing confirmed that the patient was infected with triploid forms of P. westermani.

Cellulitis and Bacteremia due to Corynebacterium striatum Identified by Matrix-assisted Laser Desorption Ionization-Time of Flight Mass Spectrometry.

Corynebacterium striatum has been described as a pathogen in immunocompromised patients; however, correctly identifying Corynebacterium spp. is often difficult, and cases of cellulitis caused by C. striatum are only rarely reported. We herein describe a case of cellulitis and bacteremia due to C. striatum identified by matrix-assisted laser desorption ionization-time of flight mass spectrometry. Antimicrobial susceptibility testing was performed using the Strepto-Haemo Supplement method, and vancomycin was replaced by a narrow-spectrum oral amoxicillin.

Helicobacter fennelliae Bacteremia: Three Case Reports and Literature Review.

Helicobacter fennelliae is a gram-negative, spiral bacillus that appears as thin-spread colonies on sheep blood agar and is similar to Helicobacter cinaedi. H fennelliae is diagnosed by genetic testing, which is not readily available in all laboratories. Therefore, H fennelliae bacteremia has only been reported sporadically, and little is known about its clinical characteristics.We describe 3 cases of H fennelliae bacteremia with gastrointestinal symptoms, including nausea, vomiting, and diarrhea. Isolates could be differentiated from H cinaedi by biochemical reaction testing, including nitrate reduction and alkaline phosphatase hydrolysis.We retrospectively reviewed 24 cases of H fennelliae bacteremia reported in the literature. Most of the patients had immunosuppressive backgrounds, including solid tumors, hematological malignancies, and autoimmune diseases. Although gastrointestinal symptoms were common, cellulitis was not often observed in patients with H fennelliae bacteremia.Clinicians should bear in mind that H fennelliae may be a differential diagnosis in patients with gastrointestinal manifestations and gram-negative, spiral bacilli. In addition, biochemical reactions, such as nitrate reduction and alkaline phosphatase hydrolysis, are useful in differentiating H fennelliae from H cinaedi.

Evaluation of Streptococcus pneumoniae in bile samples: A case series review.

Although Streptococcus pneumoniae is an important pathogen of humans, pneumococcal cholangitis is rare because of the rapid autolysis of S. pneumoniae. The aim of this case series was to review patients with bile cultures positive for S. pneumoniae. This study was a single center retrospective case series review of patients with S. pneumoniae in their bile at a tertiary-care cancer center between September 2002 and August 2015. Subjects consisted of all patients in whom S. pneumoniae was isolated in their bile during the study period. Bile specimens for culture were obtained from biliary drainage procedures such as endoscopic retrograde biliary drainage, endoscopic nasobiliary drainage, and percutaneous transhepatic biliary drainage. There were 20 patients with bile cultures positive for S. pneumoniae during the study period. All patients presented with extrahepatic obstructive jaundice due to hepatopancreatobiliary tumors. Nineteen of 20 patients underwent the placement of plastic intrabiliary tubes. The mean time between the first-time drainage and the positive culture was 26 days (range 0-313 days). Although 12 of 20 patients met our definition of cholangitis, 5 were clinically treated with antibiotics based on a physician's assessment of whether there was a true infection. The present study is the largest case series of patients with S. pneumoniae in their bile. Based on our findings, the isolation of S. pneumoniae from bile may be attributed to the placement of biliary drainage devices.

The low incidence of bacteremia after esophageal endoscopic submucosal dissection (ESD) obviates the need for prophylactic antibiotics in esophageal ESD.

BACKGROUND: Although a high incidence of bacteremia after esophageal endoscopic procedures has been reported, the incidence of bacteremia associated with esophageal endoscopic submucosal dissection (ESD) remains unknown. Therefore, we investigated the incidence of bacteremia associated with esophageal ESD. METHODS: From April 2013 to March 2014, patients who underwent esophageal ESD were enrolled prospectively. Two sets of blood cultures were collected from patients at the following time points: (1) immediately after ESD; (2) the next morning; and (3) when fever ≥38 °C was present after ESD. RESULTS: A total of 424 blood culture sets were collected from 101 patients. Six patients had positive blood cultures immediately after ESD (4 %, 7/202 sets). Another patient had a positive blood culture the next morning (0.5 %, 1/202 sets). Ten patients (10 %) developed a post-ESD fever ≥38 °C, and blood cultures from these patients were all negative (0/20 sets). The seven patients with positive blood cultures had no post-ESD fever or infectious symptoms. Growth of Bacteroides thetaiotaomicron was only observed in one patient (1 %) with positive blood cultures immediately after ESD, and this patient was diagnosed with transient bacteremia. The other six patients were considered to have contaminants in their blood cultures. Thus, the incidence of bacteremia after esophageal ESD was 1 % [95 % confidence interval (CI) 0-5 %]. No patient had infectious symptoms, and none required antibiotics after ESD. CONCLUSIONS: The incidence of bacteremia after esophageal ESD was low and post-ESD fever was not associated with bacteremia. We conclude that use of routine prophylactic antibiotics to patients undergoing esophageal ESD is unnecessary. CLINICAL TRIAL REGISTRY NUMBER: UMIN000012908.

Microbiology of Pelvic Lymphocyst Infection after Lymphadenectomy for Malignant Gynecologic Tumors.

BACKGROUND: Pelvic lymphocyst infection is a rare complication after lymphadenectomy for malignant gynecologic tumors. Although medical therapy is a useful addition to surgical drainage, the appropriate antibiotic regimen is unknown because few studies have examined the causative organisms. The purpose of this case series was to identify the micro-organisms infecting pelvic lymphocysts. METHODS: This was a single-center, retrospective, case-series review conducted at a tertiary-care cancer center between October 2002 and March 2013. The participants included all patients who experienced their first pelvic lymphocyst infection after undergoing pelvic lymphadenectomy for cervical, endometrial, or ovarian cancer and exhibited positive lymphocyst fluid culture. Computed tomography- or sonography-guided percutaneous aspiration procedures were performed to obtain lymphocyst fluid for culture. RESULTS: During the study period, 878 patients underwent lymphadenectomy for gynecologic malignant tumors, and 13 developed a pelvic lymphocyst infection documented microbiologically. Cultures identified Staphylococcus aureus (three patients), S. epidermidis (one patient), Streptococcus agalactiae (three patients), Enterococcus (two patients), Escherichia coli (one patient), and anaerobic bacteria (three patients). They were all monomicrobial infections. CONCLUSIONS: Our study and other smaller ones suggest that lymphocyst infections following pelvic lymphadenectomy for malignant gynecologic tumors usually are monomicrobial and caused by gram-positive cocci, including Staphylococcus, Streptococcus, and Enterococcus, and anaerobes such as Bacteroides fragilis. These bacteria should be considered when selecting empiric antibiotic therapy.

Surveillance of extended-spectrum ß-lactamase-producing Escherichia coli and Klebsiella pneumoniae at a comprehensive cancer center in Japan, 2009-2013.

We examined the results of surveillance of extended-spectrum ß-lactamase (ESBL)-producing Escherichia coli and Klebsiella pneumoniae, which are pathogens known to cause nosocomial outbreaks, at a comprehensive cancer center in Japan over a 5-year period. We found that the admission prevalence and the incidence of ESBL-producing E coli increased during the study period, in contrast with ESBL-producing K pneumoniae, in which the parameters remained low throughout the study period.

Infection control for extrapulmonary tuberculosis at a tertiary care cancer center.

Extrapulmonary tuberculosis (TB) can be infectious when diagnostic or therapeutic procedures are performed on infected lesions. We retrospectively describe infection control and evaluate isolation failure rates during the manipulation of active extrapulmonary TB lesions at a comprehensive cancer center over a 5-year period. Among patients with a high suspicion of cancer, extrapulmonary TB was not suspected, and airborne precautions often were not used when manipulating infected lesions.

Cryptococcus gattii genotype VGIIb infection in Japan.

This report describes a case of Cryptococcus gattii VGIIb infection of the pulmonary and central nervous systems in an immunocompetent Japanese man with a travel history, and it hypothesizes the place where he was infected with C. gattii using the genotype information.

[Evaluation of anesthetic management and renal function in open and endovascular repair of abdominal aortic aneurysm].

BACKGROUND: Since 2007, the number of patients receiving endovascular aneurysm repairs (EVARs) is increasing in Japan. Although EVAR is less invasive and has a lower short-term mortality, it has no long-term advantages and may lead to deterioration of renal function. METHODS: We retrospectively evaluated anesthetic management and renal function in patients undergoing EVAR and open repair (OR) between July 2010 and June 2011. RESULTS: Sixty-three patients (EVAR 33, OR 30) were studied. The average age of patients was significantly older in the EVAR group, and the duration of surgery and anesthesia were longer in the OR group. Despite lower blood loss in the EVAR group compared with the OR group, a massive hemorrhage (1,563 g) occurred in the EVAR group. The renal function of the EVAR group did not deteriorate within 1 year after surgery. However, the rate of acute kidney injuries (AKI) was higher in patients with renal dysfunction before operation than in patients with normal renal function. CONCLUSIONS: Although EVAR is less invasive than OR, anesthesiologists should pay attention to pre-operative comorbidity and massive hemorrhage during the operation. To avoid postoperative renal dysfunction, it is important to protect the kidney during surgery.

Raoultella ornithinolytica bacteremia in cancer patients: report of three cases.

Raoultella ornithinolytica is a Gram-negative aerobic bacillus reclassified in the new genus from the Klebsiella species based on new genetic approaches; however, human infections caused by R. ornithinolytica are rare. We herein report three cases of R. ornithinolytica bacteremia associated with biliary tract infections in cancer patients. R. ornithinolytica can be a causative pathogen of biliary tract infection in cancer patients.