Impact of Complete Surgical Resection of Metastatic Lesions in Patients with Advanced Renal Cell Carcinoma in the Era of Tyrosine Kinase Inhibitors and Immune Checkpoint Inhibitors.

Complete metastasectomy (CM) in metastatic renal cell carcinoma (mRCC) has demonstrated efficacy in the cytokine era, but its effectiveness in the era of tyrosine kinase inhibitors (TKIs) and immune checkpoint inhibitors (ICIs) remains unclear. A multi-institutional database included clinicopathological data of 367 patients with mRCC. Patients were divided into two groups: the CM group and the non-CM group. These two groups were compared before and after propensity score matching (PSM). Cox proportional hazard models were used to detect factors associated with disease-free survival (DFS) and overall survival (OS) from mRCC diagnosis. The CM group showed a significant association with longer overall survival compared to the non-CM group in the PSM-unadjusted cohorts (p < 0.001, hazard ratio 0.49, 95% confidence interval 0.35-0.69), but no superiority was noted in the adjusted cohorts. The median DFS after CM was 24 months, with no significant differences based on relapse timing. Notably, the international metastatic RCC database consortium risk categories and metastatic burden were associated with DFS. This study supports the potential of CM in mRCC management during the TKI/ICI era, although limitations including sample size and selection bias need to be considered.

Highly potent, orally active novel small-molecule HPK1 inhibitor DS21150768 induces anti-tumor responses in multiple syngeneic tumor mouse models.

Augmenting T-cell activity is a promising approach to enhance the efficacy of cancer immunotherapy treatment. Hematopoietic progenitor kinase 1 (HPK1) is predominantly expressed in immune cells and negatively regulates T-cell receptor signaling. It is reported that inhibition of the kinase function of HPK1 results in tumor growth suppression by enhancing cancer immunity. Thus, developing HPK1 inhibitors has attracted considerable attention as a future cancer immunotherapy approach. However, despite recent progress in HPK1 biology and pharmacology, various challenges still remain, such as developing HPK1 inhibitors with favorable pharmacological profiles and identifying tumor characteristics that can be applied to define susceptibility to HPK1 inhibition. Here, we present the identification and pharmacological evaluation of DS21150768, a potent small-molecule HPK1 inhibitor with a novel chemical scaffold. DS21150768 shows remarkable inhibition of HPK1 kinase activity, and in vitro studies demonstrated its potent activity to enhance T-cell function. DS21150768 is orally bioavailable and shows sustained plasma exposure, which leads to enhanced cytokine responses in vivo. We conducted a comparison of the anti-tumor efficacy of DS21150768 alone or in combination with anti-PD-1 antibody in 12 different mouse cancer cell models, and observed that the treatments suppressed tumor growth in multiple models. Furthermore, Gene Set Enrichment Analysis demonstrated significant enrichment of immune-related gene signatures in the tumor models responsive to DS21150768 treatment. Our results provide a path forward for the future development of HPK1 inhibitors and fundamental insights into biomarkers of HPK1-targeted therapy.

Significant Improvement of Prognosis After the Advent of Immune Checkpoint Inhibitors in Patients with Advanced, Unresectable, or Metastatic Urothelial Carcinoma: A Propensity Score Matching and Inverse Probability of Treatment Weighting Analysis on Real-World Data.

PURPOSE: The treatment landscape for advanced, unresectable, or metastatic urothelial carcinoma (aUC) has shifted substantially since the advent of immune checkpoint inhibitors (ICIs). We investigated the extent to which pembrolizumab therapy is superior to conventional chemotherapy as a second-line treatment. PATIENTS AND METHODS: A multicenter-derived database registered 454 patients diagnosed with aUC between 2008 and 2020. Of these, 94 patients (21%) who received second-line pembrolizumab and 75 (17%) who received second-line chemotherapy but never received third-line or later ICI therapy were included. We compared overall survival (OS) from the initial date of first-line chemotherapy between two groups by adjusting for prognostic factors through propensity score matching (PSM) and inverse probability of treatment weighting (IPTW). The IPTW-adjusted hazard ratio and 95% confidence interval were estimated using a multivariate Cox regression analysis. To identify patients who were more likely to benefit from second-line pembrolizumab than from chemotherapy, we performed a subgroup analysis for OS with an IPTW-adjusted model. RESULTS: The PSM-adjusted comparison showed a significant improvement in the prognosis with second-line pembrolizumab use (P = 0.01). The OS benefit with the advent of pembrolizumab was 8 months (18 months vs 26 months). Multivariable analyses using IPTW adjustment demonstrated that lymph node metastasis (P = 0.001), lung metastasis (P = 0.013), and bone metastasis (P = 0.003) were poor independent prognostic factors, and pembrolizumab use (P = 0.021) was a favorable independent prognostic factor. Subgroup analyses revealed that pembrolizumab was associated with survival benefits over chemotherapy in all subgroups, including young patients (age <70 years), those who received radical surgery, and those without visceral metastasis. CONCLUSION: We demonstrated a significant improvement in prognosis after the advent of pembrolizumab for patients with aUC. ICIs should not be restricted based on patient characteristics.

[A Case of Nephrocutaneous and Enterorenal Fistula Associated with Pyonephrosis].

A 57-year-old woman was referred to our hospital with a palpable mass in the left lumbar area. Computerized tomography revealed a diffusely enlarged destructed left kidney with impacted ureteropelvic junction stones and intense inflammatory stranding of the perirenal fat. This infiltration extended into the subcutaneous tissue. Since she refused to undergo nephrectomy, we performed transurethral ureterolithotripsy (TUL) two times. Retrograde ureterography before the third TUL showed communication between the renal pelvis and the jejunum. We performed a left-sided nephrectomy with a wedge resection of the jejunum. This is a rare case of nephrocutaneous and enterorenal fistula caused by pyonephrosis.

[Retrospective Analysis of Severe Renal Infections Requiring Nephrectomy to Control Infection].

Patients who contract severe renal infections often suffer from urosepsis. Therefore, early diagnosis and treatment are required. Sometimes, the treatment with antibiotics is not enough for control of the infections. Most of the patients also require surgical interventions including transurethral drainage and nephrectomy. Twenty-two patients with severe renal infections treated between April 2010 and October 2019 at our institute were evaluated retrospectively. Eleven patients had undergone nephrectomy. Open nephrectomy was performed on 10 patients. Laparoscopic nephrectomy was attempted in the other patient but was converted to open nephrectomy because severe adhesion was found around the tissues. Nephrectomy was performed by the retroperitoneal approach on 9 patients and by the transperitoneal approach on 2 patients. The reteroperitoneal approach was used on two patients who suffered postoperative colon perforation. Inflammatory involvement of renal pelvis, hilum and adjacent structures leads to dense fibrotic reaction and obliteration of tissue planes, which makes the nephrectomy procedure challenging. Therefore, it is important to choose the most appropriate nephrectomy procedure for each patient when they have contracted severe renal infections.

[Transmission of SARS-CoV-2 through contact with a SARS-CoV-2-infected individual in the presymptomatic or asymptomatic state in Japan: A case series].

Objectives　There is little evidence supporting the transmission of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) from presymptomatic or asymptomatic SARS-CoV-2-infected individuals in Japan, where the incidence of SARS-CoV-2 infection is lower than that in other developed countries. This study aimed to determine whether SARS-CoV-2 transmission can occur from presymptomatic or asymptomatic SARS-CoV-2-infected individuals.Methods　We surveyed all directors of Japanese public health centers for index cases and secondary patients who possibly contracted SARS-CoV-2 infection from a presymptomatic or asymptomatic SARS-CoV-2-infected individual who came under their care before June 20, 2020. The professional staff at the centers routinely perform contact tracing of infected persons based on the guidelines of the Infection Control Act. Four authors independently reviewed reports of 9 index cases of SARS-CoV-2-infected individuals with 17 secondary patients from 8 prefectures and examined the cases to determine whether transmission from a SARS-CoV-2-infected individual in the presymptomatic or asymptomatic state occurred.Results　We reported 7 index cases with 13 secondary patients. 1) An elderly woman acquired SARS-CoV-2 infection from her sustained asymptomatic granddaughter at home, 2) 4 guests and 1 accompanying child waiting at a hair salon acquired infection from the presymptomatic female hair stylist, 3) 2 inpatients acquired infection from a presymptomatic nurse while providing nursing care in close contact, 4) an elderly couple acquired SARS-CoV-2 infection from their presymptomatic relative who was in the 50s during household care at their home, 5) a man acquired SARS-CoV-2 infection from a presymptomatic adult neighbor in an enclosed space with poor ventilation, 6) a presymptomatic man had transmitted infection to another man at a coffee shop while having a discussion on business, and 7) a man in his 50s acquired SARS-CoV-2 infection from a presymptomatic man during 50 minutes of close contact at their office and in a car. These secondary patients had no other likely routes of infection. The interval between the date of symptom onset in the presymptomatic index case and the secondary patient ranged from 2 to 6 days. The incidence rates at the time these infections occurred in the corresponding prefectures ranged from 0.00 to 6.56 cases/1 million person-days.Conclusion　We report the first case of SARS-CoV-2 transmission from a sustained asymptomatic index case in Japan. All secondary patients came into close contact with presymptomatic index cases in areas with poor ventilation.

[A Case Report of Severe Ischemic Priapism Treated Aggressively by Proximal Shunt].

Priapism is defined as a full or partial erection for >4 hours without sexual stimulation or orgasm. Ischemic priapism is an emergency disease that urgently requires surgery to reduce pressure in the corpora cavernosa. We encountered a case of severe ischemic priapism that required a proximal shunt. The patient was a 71-year-old man who had been diagnosed with benign prostatic hyperplasia ; hence, oral administration of tamsulosin had been initiated at a local hospital. His penis became gradually rigid and presented with prolonged erection. Then he consulted our hospital. Penile aspiration was impossible, and phenylephrine injection was not effective. He was subsequently transferred to our hospital. He did not have severe pain but had complete erection. We could not correctly evaluate blood gas from the corpora cavernosa and color Doppler ultrasonography showed little blood flow in the corpora cavernosa. We judged that the patient had ischemic priapism. The erection could not be controlled by distal shunt plastic surgery performed using the T shunt and Al-Ghorab methods. Therefore, a proximal shunt plastic surgery as described by Sacher was performed. The surgery was effective, but it caused erectile dysfunction.

Author Correction: Genome defense against integrated organellar DNA fragments from plastids into plant nuclear genomes through DNA methylation.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Phosphorus Localization and Its Involvement in the Formation of Concentrated Uranium in the Renal Proximal Tubules of Rats Exposed to Uranyl Acetate.

Although the kidneys comprise a critical target of uranium exposure, the dynamics of renal uranium distribution have remained obscure. Uranium is considered to function physiologically in the form of uranyl ions that have high affinity for phosphate groups. The present study applied microbeam-based elemental analysis to precisely determine the distribution of phosphorus and uranium in the kidneys of male Wistar rats exposed to uranium. One day after a single subcutaneous injection of uranyl acetate (2 mg/kg), areas of concentrated phosphorus were scattered in the S3 segments of the proximal tubule of the kidneys, whereas the S3 segments in control rats and in rats given a lower dose of uranium (0.5 mg/kg) contained phosphorus without concentrated phosphorus. Areas with concentrated phosphorus contained uranium 4- to 14-fold more than the mean uranium concentration (126-472 vs. 33.1 ± 4.6 µg/g). The chemical form of uranium in the concentrated phosphorus examined by XAFS was uranium (VI), suggesting that the interaction of uranyl ions with the phosphate groups of biomolecules could be involved in the formation of uranium concentration in the proximal tubules of kidneys in rats exposed to uranium.

Divergence of RNA editing among Arabidopsis species.

RNA editing altered the RNA sequence by replacing the C nucleotide to U in the organellar genomes of plants. RNA editing status sometimes differed among distant species. The pattern of conservation and variation of RNA editing status made it possible to evaluate evolutionary mechanisms impacting functional aspects of RNA editing. In this study, divergence of RNA editing in the chloroplast genome among Arabidopsis species was analyzed to determine 9 losses and 1 gain in RNA editing. All changes in A. thaliana lineage resulted from changes to the chloroplast genome sequence, whereas changes in the A. lyrata / halleri lineage were possibly due to exclusive changes in the nuclear editing factors. One loss of RNA editing in A. lyrata was caused by a deficiency in the PPR gene OTP80. The changes in RNA editing occurred approximately every two million years and were not observed at functionally important sites. These results highlight the conserved nature of RNA editing status suggesting the importance of RNA editing during evolution.

Genome defense against integrated organellar DNA fragments from plastids into plant nuclear genomes through DNA methylation.

Nuclear genomes are always faced with the modification of themselves by insertions and integrations of foreign DNAs and intrinsic parasites such as transposable elements. There is also substantial number of integrations from symbiotic organellar genomes to their host nuclear genomes. Such integration might have acted as a beneficial mutation during the evolution of symbiosis, while most of them have more or less deleterious effects on the stability of current genomes. Here we report the pattern of DNA substitution and methylation on organellar DNA fragments integrated from plastid into plant nuclear genomes. The genome analyses of 17 plants show homology-dependent DNA substitution bias. A certain number of these sequences are DNA methylated in the nuclear genome. The intensity of DNA methylation also decays according to the increase of relative evolutionary times after being integrated into nuclear genomes. The methylome data of epigenetic mutants shows that the DNA methylation of organellar DNA fragments in nuclear genomes are mainly dependent on the methylation maintenance machinery, while other mechanisms may also affect on the DNA methylation level. The DNA methylation on organellar DNA fragments may contribute to maintaining the genome stability and evolutionary dynamics of symbiotic organellar and their host's genomes.

DNA Methylation Diversification at the Integrated Organellar DNA-Like Sequence.

Plants have a lot of diversity in epigenetic modifications such as DNA methylation in their natural populations or cultivars. Although many studies observing the epigenetic diversity within and among species have been reported, the mechanisms how these variations are generated are still not clear. In addition to the de novo spontaneous epi-mutation, the intra- and inter-specific crossing can also cause a change of epigenetic modifications in their progenies. Here we report an example of diversification of DNA methylation by crossing and succeeding selfing. We traced the inheritance pattern of epigenetic modification during the crossing experiment between two natural strains Columbia (Col), and Landsberg electa (Ler) in model plant Arabidopsis thaliana to observe the inheritance of DNA methylation in two organellar DNA-like sequence regions in the nuclear genome. Because organellar DNA integration to the nuclear genome is common in flowering plants and these sequences are occasionally methylated, such DNA could be the novel source of plant genome evolution. The amplicon sequencing, using bisulfite-converted DNA and a next-generation auto-sequencer, was able to efficiently track the heredity of DNA methylation in F1 and F2 populations. One region showed hypomethylation in the F1 population and succeeding elevation of DNA methylation with large variance in the F2 population. The methylation level of Col and Ler alleles in F2 heterozygotes showed a significant positive correlation, implying the trans-chromosomal effect on DNA methylation. The results may suggest the possible mechanism causing the natural epigenetic diversity within plant populations.

Genome-Wide Analysis of Parent-of-Origin Allelic Expression in Endosperms of Brassicaceae Species, Brassica rapa.

Uniparental gene expression, observed in both animals and plants, is termed genomic imprinting. Genomic imprinting is a well-known epigenetic phenomenon regulated through epigenetic modifications such as DNA methylation and histone modifications. Recent genome-wide studies of endosperm transcription have revealed the rapid change of imprinted genes between species, suggesting the flexibility of this phenomenon. Although the functional significance and evolutionary trends of imprinted genes are still obscure, it can be clarified by inter-species comparisons. In this study, we analyzed the pattern of genomic imprinting in Brassica rapa, a species related to Arabidopsis thaliana. Compared with the ancient karyotype of A. thaliana and B. rapa, B. rapa has a triplicated genome. Many imprinted genes, beyond the estimated number previously reported in other species, were observed. Several imprinted genes have been conserved among species in Brassicaceae. We also observed rapid molecular evolution of imprinted genes compared to non-imprinted genes in B. rapa. Especially, imprinted gene overlapping between species showed more rapid molecular evolution and preferential expression in endosperms. It may imply that a small number of imprinted genes have retained functional roles among diverged species and have been the target of natural selection.

The complete chloroplast genome of Sisymbrium irio.

The complete chloroplast genome of Sisymbrium irio was determined. The length of the complete chloroplast genome is 154,001 bp. The whole chloroplast genome consists of 83,891 bp long single copy (LSC) and 17,630 bp small single copy (SSC) regions, separated by a pair of 26,240 bp inverted repeat (IR) regions. The S. irio chloroplast genome encodes 112 annotated known unique genes including 79 protein-coding genes, 30 tRNA genes, and four rRNA genes. The phylogenetic position of S. irio is sister to Brassiceae and Thlaspideae.

Analysis of nuclear mitochondrial DNAs and factors affecting patterns of integration in plant species.

Sequences homologous to organellar DNA that have been integrated into nuclear genomes are referred to as nuclear mitochondrial DNAs (NUMTs) and nuclear plastid DNAs (NUPTs). NUMTs in nine plant species were analyzed to reveal the integration patterns and possible factors involved. The cumulative lengths of NUMTs in two-thirds of species analyzed were greater than those of NUPTs observed in a previous study. The age distribution of NUMTs was similar to that of NUPTs, suggesting similar mechanisms for integration and degradation of both NUPTs and NUMTs. Nuclear genome size and the cumulative length of NUMTs showed a significant positive correlation for older but not younger NUMTs. The same correlation was also found between nuclear genome size and older NUPTs in 17 species. These results suggested that genome size is a key factor to determine the cumulative length of relatively older NUPTs/NUMTs. Although the factor(s) determining the cumulative length of younger NUPTs/NUMTs is unclear, these sequences may be more deleterious, which could explain the different manner of determining the cumulative length of younger NUPTs/NUMTs in nuclear genomes. In addition, a relationship between the cumulative length of integrated NUMTs and complexity of mitochondrial genomes (i.e., the number of repeats) was found. The results indicate that the structural complexity of both NUMTs and their original mitochondrial sequences affects integration and degradation processes.

Gene duplication, silencing and expression alteration govern the molecular evolution of PRC2 genes in plants.

PRC2 genes were analyzed for their number of gene duplications, dN/dS ratios and expression patterns among Brassicaceae and Gramineae species. Although both amino acid sequences and copy number of the PRC2 genes were generally well conserved in both Brassicaceae and Gramineae species, we observed that some rapidly evolving genes experienced duplications and expression pattern changes. After multiple duplication events, all but one or two of the duplicated copies tend to be silenced. Silenced copies were reactivated in the endosperm and showed ectopic expression in developing seeds. The results indicated that rapid evolution of some PRC2 genes is initially caused by a relaxation of selective constraint following the gene duplication events. Several loci could become maternally expressed imprinted genes and acquired functional roles in the endosperm.

Patterns of genomic integration of nuclear chloroplast DNA fragments in plant species.

The transfer of organelle DNA fragments to the nuclear genome is frequently observed in eukaryotes. These transfers are thought to play an important role in gene and genome evolution of eukaryotes. In plants, such transfers occur from plastid to nuclear [nuclear plastid DNAs (NUPTs)] and mitochondrial to nuclear (nuclear mitochondrial DNAs) genomes. The amount and genomic organization of organelle DNA fragments have been studied in model plant species, such as Arabidopsis thaliana and rice. At present, publicly available genomic data can be used to conduct such studies in non-model plants. In this study, we analysed the amount and genomic organization of NUPTs in 17 plant species for which genome sequences are available. The amount and distribution of NUPTs varied among the species. We also estimated the distribution of NUPTs according to the time of integration (relative age) by conducting sequence similarity analysis between NUPTs and the plastid genome. The age distributions suggested that the present genomic constitutions of NUPTs could be explained by the combination of the rapidly eliminated deleterious parts and few but constantly existing less deleterious parts.

Importance of gene duplication in the evolution of genomic imprinting revealed by molecular evolutionary analysis of the type I MADS-box gene family in Arabidopsis species.

The pattern of molecular evolution of imprinted genes is controversial and the entire picture is still to be unveiled. Recently, a relationship between the formation of imprinted genes and gene duplication was reported in genome-wide survey of imprinted genes in Arabidopsis thaliana. Because gene duplications influence the molecular evolution of the duplicated gene family, it is necessary to investigate both the pattern of molecular evolution and the possible relationship between gene duplication and genomic imprinting for a better understanding of evolutionary aspects of imprinted genes. In this study, we investigated the evolutionary changes of type I MADS-box genes that include imprinted genes by using relative species of Arabidopsis thaliana (two subspecies of A. lyrata and three subspecies of A. halleri). A duplicated gene family enables us to compare DNA sequences between imprinted genes and its homologs. We found an increased number of gene duplications within species in clades containing the imprinted genes, further supporting the hypothesis that local gene duplication is one of the driving forces for the formation of imprinted genes. Moreover, data obtained by phylogenetic analysis suggested "rapid evolution" of not only imprinted genes but also its closely related orthologous genes, which implies the effect of gene duplication on molecular evolution of imprinted genes.

Evolution of the ONSEN retrotransposon family activated upon heat stress in Brassicaceae.

A Ty1/Copia-like retrotransposon, ONSEN, is activated by heat stress in Arabidopsis thaliana, and its de novo integrations that were observed preferentially within genes implies its regulation of neighboring genes. Here we show that ONSEN related copies were found in most species of Brassicaceae, forming a cluster with each species in phylogenetic tree. Most copies were localized close to genes in Arabidopsis lyrata and Brassica rapa, suggesting conserved integration specificity of ONSEN family into genic or open chromatin. In addition, we found heat-induced transcriptional activation of ONSEN family in several species of Brassicaceae. These results suggest that ONSEN has conserved transcriptional activation promoted by environmental heat stress in some Brassicaceae species.

Genetic structure and putative selective sweep in the pioneer tree, Zanthoxylum ailanthoides.

Zanthoxylum ailanthoides Siebold & Zucc. is one of the most frequently encountered pioneer trees in Japanese warm-temperate evergreen oak forests. Our previous study in one region of Japan suggested high levels of population differentiation and putative natural selection acting on one of the nuclear loci analyzed. Here, we extend our analysis to study the genetic structure of 10 populations of Z. ailanthoides across Japan using 9 simple sequence repeat (SSR) loci for a better understanding of its genetic structure. First, the southernmost population (Kagoshima) in the samples was found to have the highest genetic diversity, suggesting there was a glacial refugium at or near the location of the population. Second, relatively strong genetic differentiation was found among populations, and there was a positive correlation between genetic distances and geographic distances (Mantel test; P < 0.001). Based on this information, we analyzed nucleotide variation at the putatively selected locus homologous to the gene encoding the ADP-glucose pyrophosphorylase large subunit (agpL). Despite the strong genetic differentiation among populations suggested by the SSR loci, the agpL locus was monomorphic in almost all populations analyzed. The results of this study strongly supported the possibility of a selective sweep at or near the agpL locus.