The complexity of EGFR exon 19 deletion and L858R mutant cells as assessed by proteomics, transcriptomics, and metabolomics.

Most lung adenocarcinoma-associated EGFR tyrosine kinase mutations are either an exon 19 deletion (19Del) or L858R point mutation in exon 21. Although patients whose tumors contain either of these mutations exhibit increased sensitivity to tyrosine kinase inhibitors, progression-free and overall survival appear to be longer in patients with 19Del than in those with L858R. In mutant-transfected Ba/F3 cells, 19Del and L858R were compared by multi-omics analyses including proteomics, transcriptomics, and metabolomics. Proteome analysis identified increased plastin-2, TKT, PDIA5, and ENO1 expression in L858R cells, and increased EEF1G expression in 19Del cells. RNA sequencing showed significant differences between 19Del and L858R cells in 112 genes. Metabolome analysis showed that amino acids, adenylate, guanylate, NADPH, lactic acid, pyruvic acid glucose 6-phosphate, and ribose 5-phosphate were significantly different between the two mutant cells. Because GSH was increased with L858R, we combined osimertinib with the GSH inhibitor buthionine sulfoximine in L858R cells and observed synergistic effects. The complexity of EGFR 19Del and L858R mutant cells was demonstrated by proteomics, transcriptomics, and metabolomics analyses. Therapeutic strategies for lung cancer with different EGFR mutations could be considered because of their different metabolic phenotypes.

Targeting ROR1 in combination with osimertinib in EGFR mutant lung cancer cells.

Lung cancer that exhibits epidermal growth factor receptor (EGFR) gene mutation is sensitive to EGFR-tyrosine kinase inhibitors (TKIs), such as osimertinib. Receptor tyrosine kinase-like orphan receptor 1 (ROR1) may be involved in overcoming EGFR-TKI resistance. Growth inhibition, colony formation, apoptosis, and mRNA/protein levels in four osimertinib-sensitive and resistant cell lines transfected with small interfering RNA (siRNA) targeting ROR1 (siROR1) were evaluated. Cell growth and colony formation were suppressed and apoptosis was increased in all cell lines treated with siROR1. Although EGFR, AKT, and ERK phosphorylation were not suppressed in all cell lines, TGF-ß2, AXL, CDH2, PARP1, PEG10, and TYMS mRNA expression levels were reduced. The combination of osimertinib with siROR1 was effective for the four cell lines, particularly in the two osimertinib-sensitive lines. In conclusion, targeting ROR1 in combination with osimertinib in EGFR mutant lung cancer may be a novel therapeutic option.

[Relationship between Coombs-positive autoimmune hemolytic anemia and development of malignant tumors: a retrospective study].

Autoimmune hemolytic anemia (AIHA) is secondary to underlying diseases, such as autoimmune diseases and lymphoid malignancies. Recently, solid cancers have also been reported to be associated with AIHA, although there is not much information available. In this study, we retrospectively examined the correlation between AIHA and onset of malignancy in 100 patients diagnosed with AIHA based on the broad definition of AIHA at our hospital and cooperating institutions from January 1, 1995 to May 31, 2016. Malignancies were detected in 52 of the 100 patients (hematological malignancies: 39 patients; solid cancers: 22 patients; total malignancies including multiple primary malignancies: 67 patients). Of the 67 patients with malignancies, 28 were diagnosed with malignancies within 6 months of AIHA diagnosis. All patients with cold agglutinin disease (CAD) were associated with malignancies. Compared with warm AIHA, solid cancers were significantly more common among the patients with CAD. These findings emphasize the importance of investigating the malignancies upon diagnosis of AIHA.

Publication of lung cancer clinical trials in the Japanese Clinical Trial Registry.

INTRODUCTION: Since June 2005, the University Hospital Medical Information Network-Clinical Trial Registry (UMIN-CTR) has been an International Committee of Medical Journal Editors (ICMJE)-approved clinical trial registry in Japan. The number of clinical trials registered in the UMIN-CTR has increased annually. To date, no report exists regarding the publishing of clinical trials registered in the UMIN-CTR. Therefore, we evaluated the publication frequency of clinical trials registered in the UMIN-CTR in Japan. METHODS: We targeted trials that assessed the treatment effect of chemotherapy or molecular targeting drugs for lung cancer. We included trials registered between June 2005 and January 2010, and identified published trials through a computer-based search of MEDLINE and Google Scholar. The cumulative publication rate of the trials was calculated using Kaplan-Meier analysis. RESULTS: In our study, 179 trials met the inclusion criteria. Of these, 46.4% (83/179) trials were published by the end of the cut-off period. With regard to publication, differences existed between the information recorded in the UMIN-CTR database and the actual searched results. The publication rate between groups was insignificantly different; however, whether a clinical study group did or did not conduct a trial differed significantly (53.3% vs. 36.1%; P = 0.024). Phase II studies with positive results were more likely to be published (84.4%); however, the overall publication rate was low (41.8%), which may reflect publication bias. CONCLUSIONS: The UMIN-CTR fundamentally functions as the unique ICMJE-approved clinical trial registry in Japan. However, it seems insufficient to require it as the official clinical database.

[Hereditary red cell membrane disorders in Japan: comparison with other countries].

Red cell membrane disorders are the most common type of inherited hemolytic disorders in the Japanese population. In hereditary spherocytosis (HS), the primary presentation is a loss of membrane surface area, leading to reduced deformability because of defects in the membrane proteins ankyrin, band 3, ß-spectrin, α spectrin, or protein 4.2 (P4.2). Complete P4.2 deficiencies, which are inherited in an autosomal recessive manner, comprise a unique HS subgroup and are common in Japanese, but rare in other populations. In contrast, the principle presentation in hereditary elliptocytosis (HE) is mechanical weakness of the erythrocyte membrane skeleton due to defects in α-spectrin, ß-spectrin, or protein 4.1. Although α-spectrin mutations are the most frequent cause of HE in Caucasian, African, and Mediterranean populations, these mutations are rare in the Japanese population, in which P4.1 deficiencies are instead most common. Furthermore, hereditary stomatocytoses (HSt) are disorders of monovalent cation permeability in the red cell membrane.

[Band 3 deficiency as a cause of hereditary spherocytosis].

Band 3 protein accounts for the largest percentage of whole erythrocyte membrane proteins. Abnormalities in this protein are closely associated with pathologies including hereditary spherocytosis (HS), Southeast Asian ovalocytosis and distant renal tubular acidosis. Currently, EMA binding capacity measurement in erythrocytes is the most useful screening test for diagnosing HS. We have also demonstrated reduced EMA binding capacity in patients with HS who have deficiencies of membrane proteins such as ankyrin not directly binding to EMA and who have as yet undetectable abnormalities of membrane proteins. However, even patients with hereditary elliptocytosis, who have a partial spectrin deficiency, were found to show reduced EMA binding capacity. Six of 7 had spherocytic elliptocytosis. Therefore, it is necessary to meticulously diagnose HS by ruling out all other possibilities.

Varied clinical course of aplastic crisis in hereditary spherocytosis.

This study is the first to report a familial case involving differing clinical courses of aplastic crisis triggered by parvovirus B19 in two patients with HS, although similar eosin-5-maleimide-binding test and sodium dodecylsulfate-polyacrylamide gel electrophoresis results had been obtained for both. One patient had short-term mild symptoms, whereas the other patient developed severe anemia that required blood transfusion, experienced fever for 13 days, and did not have any rash. The severity of aplastic crisis is reported to be correlated with the severity of the underlying hemolytic anemia; the present findings show that the severity of infection should also be considered as an important predictive factor of the severity of aplastic crisis.

A rapid screening and production method using a novel mammalian cell display to isolate human monoclonal antibodies.

Antibody display methods are increasingly being used to produce human monoclonal antibodies for disease therapy. Rapid screening and isolation of specific human antibody genes are valuable for producing human monoclonal antibodies showing high specificity and affinity. In this report, we describe a novel mammalian cell display method in which whole human IgG is displayed on the cell surface of CHO cells. Cells expressing antigen-specific human monoclonal IgGs with high affinity on the cell surface after normal folding and posttranscriptional modification were screened using a cell sorter. The membrane-type IgG-expressing CHO cells were then converted to IgG-secreting cells by transfection with a plasmid coding Cre recombinase. This mammalian cell display method was applied to in vitro affinity maturation of monoclonal C9 IgG specific to the human high-affinity IgE receptor (FcεRIα). The CDR3 of the C9 heavy chain variable region gene was randomly mutated and inserted into pcDNA5FRT/IgG. A C9 IgG (CDRH3r)-expressing CHO cell display library consisting of 1.1×10(6) independent clones was constructed. IgG-displaying cells showing high reactivity to FcεRIα antigen were screened by the cell sorter, resulting in the establishment of a CHO cell line producing with higher reactivity than the parent C9 IgG.

Clinical Features of Patients With Hematological Malignancies Treated at the Palliative Care Unit.

Background: In Japan, the number of patients with aggressive hematological malignancies (PHMs) admitted at the palliative care unit (PCU) in their end-of-life (EOL) stage was fewer than that of patients with solid tumors due to several reasons. The assessment of patient characteristics and the methods of survival prediction among PHMs in the EOL stage are warranted. Objectives: This study aimed to identify the current medical status and the method of survival prediction among PHMs treated at the PCU. Setting/Subjects/Measurements: We retrospectively analyzed the clinical data of 25 PHMs treated at our PCU between January 2017 and December 2020. The association between survival time and the palliative prognostic score (PAP) and palliative prognostic index (PPI) was analyzed. Results: The average age of the PHMs was higher than that of patients with lung cancer as a control. The median survival time of the PHMs was shorter than the control group. Most PHMs could not receive standard chemotherapy, and the most common cause of death was disease-related organ failure. Significant associations were observed between the survival time and each PAP/PPI value in patients with malignant lymphoma, but not in those with leukemia. Conclusion: The PHMs in the PCU had a lower median survival time than the control group. These results were induced by the result of patient selection to avoid treatment-related severe toxicity. The survival prediction using the PAP and PPI was less accurate in patients with leukemia.

Herpes zoster in lung cancer patients treated with PD-1/PD-L1 inhibitors.

Background: Herpes zoster (HZ) occurs mostly in elderly and immunocompromised individuals. Immune reconstitution may be associated with the pathogenesis of HZ. As immune checkpoint inhibitor (ICI) treatment amplifies the immune response, use of ICI may increase the incidence of HZ. There have been few studies of HZ in lung cancer patients treated with ICI. This study was performed to investigate the frequency of HZ in lung cancer patients who received ICI or cytotoxic chemotherapeutic agents. Methods: We searched the electronic medical records for lung cancer patients receiving anticancer drug therapy at our hospital, who developed HZ between April 2011 and June 2020. Results: The review identified 80 patients with a history of ICI treatment (ICI group) and 356 who had been treated with cytotoxic chemotherapeutic agents alone (non-ICI group). Among the 20 patients who developed HZ, 4 (5.0%) belonged to the ICI group and 16 (4.5%) to the non-ICI group (P=0.782). After exclusion of patients aged 65 years and older, to avoid effects of advanced age on the results, the ICI and non-ICI groups consisted of 24 and 81 patients, respectively. In total, 3 of the 24 patients (12.5%) in the ICI group and 1 of the 81 (1.2%) patients in the non-ICI group developed HZ (P=0.0365). Conclusions: There was no significant difference in the rate of HZ between lung cancer patients treated with ICI and those treated with cytotoxic chemotherapy alone. However, patients younger than 65 years treated with ICI might be at increased risk of HZ. Because this is a retrospective small study, further prospective observational studies are needed.

IgA Vasculitis in a Lung Cancer Patient During Chemoradiotherapy.

A 72-year-old man with locally advanced lung squamous cell carcinoma experienced red purpura on the lower legs and hematuria when the disease progressed during definitive chemoradiotherapy. He had renal dysfunction and proteinuria. Biopsy specimens of the skin lesion and kidney revealed immunoglobulin A vasculitis. Potential causes such as paraneoplastic syndrome and cancer treatment have been proposed. The administration of steroids rapidly improved the symptoms. The presentation of immunoglobulin A vasculitis is accompanied by malignancies. Clinicians should keep this syndrome in mind, even during curative-intent treatment.

Lung cancer metastasis to the pancreas mimicking autoimmune pancreatitis.

The unique radiological manifestation mimicking autoimmune pancreatitis caused by lung cancer metastasis to the pancreas has not previously been reported. The incidence of pancreatic secondary tumors has previously been reported to be approximately 15% in autopsy cases of malignant tumors, and it is unusual for thoracic oncologists to find that the second common primary tumor site of metastatic pancreas tumor is the lung.

Do Elderly Lung Cancer Patients Aged ≥75 Years Benefit from Immune Checkpoint Inhibitors?

Lung cancer patients ≥75 years represent nearly 40% of all lung cancer patients and continue to increase. If elderly patients have a good performance status and adequate organ function, they can be treated the same as non-elderly patients. However, few comparative studies limited to elderly patients (≥75 years) have been conducted. We review the evidence on using immune check inhibitors for the treatment of elderly patients (≥75 years old) with advanced non-small cell lung cancer. Prospective randomized or non-randomized, retrospective, registrational, insurance-based, and community-based studies have shown that elderly (≥75 years) and non-elderly patients are similarly treated with immune check inhibitors effectively and safely. However, such analyses have not shown that immune check inhibitors are significantly more effective than chemotherapy alone. In addition, patient selection might be critically performed to administer immune check inhibitors in the elderly because they are more likely to have a poor performance status with comorbidities, which lead to little benefit, even in non-elderly patients. There is a need for more evidence showing the benefit of immune check inhibitors in non-small cell lung cancer patients ≥75 years.

Targeting ROR1 in combination with pemetrexed in malignant mesothelioma cells.

OBJECTIVE: Receptor tyrosine kinase-like orphan receptor 1 (ROR1) is overexpressed in a subset of malignant cells. However, it remains unknown whether ROR1 is targetable in malignant mesothelioma (MM). Therefore, in this study, we investigated the effects of ROR1 inhibition in mesothelioma cells. MATERIALS AND METHODS: Growth inhibition, colony formation, apoptosis, and mRNA/protein levels using siRNA-transfected MM cells were evaluated. Cluster analysis using Gene Expression Omnibus repository of transcriptomic information was also performed. RESULTS: Our results indicated that in three (H2052, H2452, and MESO-1) among four MM cell lines, ROR1 inhibition had anti-proliferative and apoptotic effects and suppressed the activation of AKT and STAT3. Although growth inhibition by siROR1 was minimal in another mesothelioma cell line (H28), colony formation was significantly suppressed. Microarray, quantitative polymerase chain reaction, and Western blot analyses showed that there were differences in the suppression of mRNA and proteins between H2452 and H28 cells transfected with siROR1 compared with those transfected with control siRNA. Cluster analysis further showed that MM tumors had relatively high ROR1 expression, although the cluster in them was different from that in MM cell lines. Thymidylate synthase, a target of pemetrexed, was downregulated in H2452 cells transfected with siROR1. Accordingly, a combination of pemetrexed with siROR1 was found to be effective in the three MM cell lines we studied. CONCLUSION: Our findings may provide novel therapeutic insight into the treatment of advanced MM.

Hereditary spherocytosis in 3 children coexisting with UDP-glucuronyl transferase 1A1 deficiency.

Simultaneous presence of hemolytic anemia and bilirubin UDP-glucuronosyltransferase deficiency is a possible cause of misdiagnosis. Seven-year-old and 17-year-old brothers and a 15-year-old sister consecutively suffered from aplastic crises. Although few spherocytes were present, the siblings and their mother had diagnoses of hereditary spherocytosis with flow cytometric analysis of eosin-5'-maleimide-labeled red blood cells in addition to osmotic fragility test. However, inappropriately high values of bilirubin compared with mild hemolysis persisted. Further analysis of UDP-glucuronyltransferase 1A1 revealed all 3 siblings were heterozygous for A(TA)7TAA-P229Q. We report here the importance of careful evaluation of mild hereditary spherocytosis masking UDP-glucuronyltransferase 1A1 deficiency.

[Development of treatment adherence by jellification of itraconazole oral solution].

There have been some reports on the efficacy and tolerability of itraconazole (ITCZ) as prophylaxis for fungal infection after HSCT, and guidelines recommend itraconazole as a standard drug for prophylaxis of fungal infection in HSCT patients. However, it is not uncommon for patients undergoing HSCT to develop anorexia and taste disturbance. There are some cases where the bitter taste of ITCZ oral solution leads to interruption of administration because the patient refuses to take this medicine. Therefore, we investigated the clinical utility and influence on continuing treatment adherence by jellification of ITCZ. Compared with ITCZ oral solution, jellified ITCZ was extremely easy for most patients to take, and it was suggested that jellified ITCZ can make it easier for patients to continue treatment if they have difficulty with administration because of the bitter taste of ITCZ oral solution. Furthermore, it was confirmed that the plasma concentration of ITCZ was suitable for prophylaxis even with jellified ITCZ. This also suggested that the efficacy of ITCZ would be maintained by using jellified formation. For long-term antifungal therapy in patients with a high risk of fungal infection such as those having HSCT, it is very important for successful prophylaxis to maintain good adherence.

Drug-induced interstitial lung disease in the treatment of malignant lymphoma as a potential diagnostic marker: a comparison of serum Krebs von Lungen-6 and thymus and activation-regulated chemokine/CC chemokine ligand 17.

PURPOSE: Cure-oriented treatment of malignant lymphoma (ML) is possible even in an advanced stage; however, the progression of drug-induced interstitial lung disease (DILD) sometimes accounts for poor clinical outcomes. This study aims to assess the incidence and clinical characteristics of DILD among patients with ML and compares the serum level of Krebs von den Lungen-6 (KL-6) with that of circulating thymus and activation-regulated chemokine (TARC)/CC chemokine ligand 17 (CCL17) as a diagnostic biomarker for DILD. PATIENTS AND METHODS: Between July 2011 and August 2016, we enrolled 36 patients with ML who were undergoing systemic chemotherapy at our hospital. Then, we evaluated the serum concentration of KL-6 and TARC/CCL17 by a sandwich-type electrochemiluminescence immunoassay and enzyme-linked immunosorbent assay, respectively. RESULTS: DILD developed in 22.2% of patients with ML. All patients recovered immediately after the discontinuation of causative drug and/or glucocorticoid therapy. Although the sensitivity of both TARC/CCL17 and KL-6 was almost equal, the mean concentration of serum KL-6 after the progression of interstitial lung disease was significantly higher than that before progression. CONCLUSION: DILD developed in patients who were treated with first-line rituximab combined regimen. Remarkably, TARC/CCL17 and KL-6 seemed approximately equal as a predictive biomarkers for DILD; however, KL-6 was more specific than TARC/CCL17.