Mucosal melanoma of the head and neck: a retrospective analysis of 34 cases in Japan.

PURPOSE: Mucosal melanoma of the head and neck (MMHN) is a rare condition. This study aimed to investigate oncological outcomes of surgical intervention in patients with MMHN. MATERIALS AND METHODS: The study included 34 patients with MMHN who underwent surgical resection as initial treatment at 10 institutions in Japan between July 2005 and June 2015. Results: The 5-year overall survival (OS), local control rate (LCR), disease-free survival (DFS), and disease-specific survival (DSS) rates were 48.7%, 53.4%, 32.4%, and 55.1%, respectively. Based on multivariate analysis, no independent prognostic factors for the 5-year OS and DSS were found. Based on univariate analysis, the 5-year LCR was worse in patients with lesions in the nasal cavity and paranasal sinuses than in the oral cavity and pharynx. However, no differences in oncological outcomes were identified in relation to primary sites, and postoperative radiotherapy (PORT) and adjuvant systemic therapy did not contribute to improvements in the 5-year OS. CONCLUSIONS: No independent prognostic factors for the 5-year OS or DSS were identified. Regional or distant recurrences are often identified, regardless of local control with surgical resection. Difficult control of MMHN with conventional therapeutic strategies, such as surgical intervention, PORT, and systemic therapy, has been suggested.

Diastereo- and Enantioselective Reductive Mannich-type Reaction of α,ß-Unsaturated Carboxylic Acids to Ketimines: A Direct Entry to Unprotected ß2,3,3 -Amino Acids.

Stereoselective construction of unprotected ß-amino acids is a significant challenge owing to the lack of methods for the catalytic generation of highly enantioenriched carboxylic acid enolates. In this study, a novel copper-catalyzed diastereo- and enantioselective reductive Mannich-type reaction of α,ß-unsaturated carboxylic acids was developed, which provides a direct and scalable synthetic method for enantioenriched ß2,3,3 -amino acids with vicinal stereogenic centers. The protocol features in situ generation of transiently protected carboxylic acids by a hydrosilane and their diastereo- and enantioselective reductive coupling with ketimines. The synthetic utility of this process was demonstrated by a gram-scale reaction and the transformation of ß-amino acids.

How long should urologists observe patients with prostate cancer after radical prostatectomy? Weibull analysis.

OBJECTIVES: There is no recommended observation time for patients who have undergone radical prostatectomy for prostate cancer. This study was undertaken to determine the postoperative observation time by investigating the hazard rate for prostate-specific antigen failure and other-cause death using Weibull analysis. METHODS: We included 612 patients who underwent laparoscopic radical prostatectomy for localized prostate cancer between June 2002 and December 2017. Risk classification was categorized by the D'Amico risk classification, and the patients were divided into three age groups: <60, 60-69 and ≥70 years. The hazard rates at each point were derived using Weibull analysis. The optimal observation time after laparoscopic radical prostatectomy was determined as the intersection point at which the hazard rate of other-cause death overtakes the hazard rate of prostate-specific antigen failure. RESULTS: In all groups classified by age, the hazard rate of other-cause deaths increased over time. In contrast, the hazard rate of prostate-specific antigen failure decreased gradually. The ≥70 years age group showed the highest hazard rate. The hazard rate of prostate-specific antigen failure was highest in the high-risk group. The patients aged ≥70 and 60-69 years in the low-risk group were recommended 6 years 6 months and 14 years 8 months, respectively, for observation. The remaining patients were recommended >25 years of postsurgical observation. CONCLUSIONS: The observation time after laparoscopic radical prostatectomy could be estimated by comparing the estimated hazard rates of prostate-specific antigen failure and other-cause death based on Weibull analysis. Urologists should pay attention to age and risk classifications for optimal postoperative observation.

Conditional Survival and Time of Biochemical Recurrence of Localized Prostate Cancer in Japanese Patients Undergoing Laparoscopic Radical Prostatectomies.

PURPOSE: Using conditional survival (CS) analysis, we investigated whether the duration of survival without biochemical recurrence (BCR) of prostate cancer after laparoscopic radical prostatectomies (LRP) affected the BCR rate. We also investigated the impact of well-known risk factors for BCR. METHODS: Between 2002 and 2014, 627 consecutive patients underwent LRPs at our institution. Prostate-specific antigen (PSA) concentrations above 0.2  ng/mL were defined as BCR. Conditional BCR-free survival rates were determined through Kaplan-Meier analysis. Assessment of potential BCR risk factors was performed using a Cox proportional hazard analysis. RESULTS: The 10-year BCR-free rates after LRP increased to 82.4%, 84.5%, 86.6%, 90.1%, and 94.7% in patients surviving 1, 2, 3, 5, and 7.5 years without BCR, respectively. Multivariate analyses of age, PSA concentrations, neoadjuvant therapy, and pathological findings were performed for all patients. In all patients, positive surgical margins (PSM) and Gleason Grade Groups (GG) ≥ 4 were independent risk factors for BCR (p < 0.001, hazard ratio [HR] = 2.45; and p < 0.001, HR = 2.83, respectively,). Similarly, PSM and GG ≥ 4 were significant risk factors in patients surviving 1-5 years without BCR. No clear risk factors were observed in patients surviving > 5 years without BCR after LRPs. CONCLUSIONS: The BCR-free rate increased with time after LRP. It is recommended that patients with PSM, GG ≥ 4, or with both factors are strictly monitored for 5 years postoperatively. CS analysis is particularly useful for predicting the postoperative course of patients.

Functional outcomes of transurethral thulium laser enucleation versus bipolar transurethral resection for benign prostatic hyperplasia over a period of 12 months: A prospective randomized study.

OBJECTIVE: To evaluate functional outcomes over a year for benign prostatic hyperplasia treated with thulium laser enucleation of the prostate or bipolar transurethral resection of the prostate. METHODS: In this study, we prospectively included 140 patients who were treated for benign prostatic hyperplasia from April 2017 to February 2019. A randomization list was used for non-blind assignment to treatment groups (thulium laser enucleation of the prostate or bipolar transurethral resection of the prostate). The International Prostate Symptom Score, International Prostate Symptom Score quality of life, Overactive Bladder Symptom Score, uroflowmetry and International Index of Erectile Function-5 were used for the assessment of outcomes. Scores were taken before treatment, and at 1, 3, 6, 9 and 12 months after treatment. RESULTS: Preoperatively, characteristics of patients in the thulium laser enucleation of the prostate group (n = 70) and the bipolar transurethral resection of the prostate group (n = 70) did not significantly differ. The thulium laser enucleation of the prostate group had a lower median decrease in hemoglobin (0.10 g/dL vs 0.69 g/dL, P < 0.0001) and shorter median catheterization (2 days vs 3 days, P < 0.0001) than the bipolar transurethral resection of the prostate group. Both groups had significantly improved International Prostate Symptom Score, International Prostate Symptom Score quality of life, Overactive Bladder Symptom Score, maximum flow rate and residual urine compared with preoperative values. In the thulium laser enucleation of the prostate group, erectile function was significantly impaired at 1 month post-treatment compared with pretreatment, but improved to baseline at 3 months; however, the transurethral resection of the prostate group did not recover. However, total energy (P = 0.001) and laser irradiation time (P = 0.025) were significantly different between the patients who had preserved erectile function and those who did not after the treatment in the thulium laser enucleation of the prostate group. CONCLUSIONS: Thulium laser enucleation of the prostate is superior to bipolar transurethral resection of the prostate in resection efficacy, hemostasis, catheterization time and preservation of erectile function. However, excessive laser irradiation time in thulium laser enucleation of the prostate has the possibility to delay improvement of overactive bladder symptoms and decrease sexual function.

A retrospective multicenter study of sublingual gland carcinoma in Japan.

OBJECTIVE: Salivary gland carcinoma is rare among head and neck cancers. Sublingual gland carcinoma, a type of salivary gland carcinoma, is even rarer; therefore, the number of cases at a single institute is too small for sufficient evaluation of tumor characteristics. We conducted a multicenter, retrospective analysis of sublingual gland carcinomas in patients who visited 12 institutions associated with the Kyoto Hospital and Affiliated Facilities Head and Neck Clinical Oncology Group. METHODS: Thirteen previously untreated patients who visited the institutions between 2006 and 2015 were enrolled. The overall survival (OS) and disease-free survival (DFS) rates for all patients and by disease stage were analyzed. Statistical analyses were performed for all patients with respect to disease stage. RESULTS: Eight of thirteen patients were diagnosed with adenoid cystic carcinoma on pathological study. A significant difference in OS rate was observed between patients with Stage I-III and Stage IV disease; however, the difference in DFS rate by disease stage was not significant. CONCLUSION: Stage IV disease was identified as a poor prognostic factor in patients with sublingual gland carcinoma. However, even patients with Stage I-III disease experienced relatively short DFS. Distant metastasis is a serious problem among patients with sublingual gland carcinoma.

Survival in patients with submandibular gland carcinoma - Results of a multi-institutional retrospective study.

OBJECTIVE: Clinical studies demonstrating the prognostic factors in submandibular gland carcinoma are limited because the tumor is relatively rare. The aim of this study was to identify clinical outcomes and prognostic factors in submandibular gland carcinoma. METHODS: The study included 65 patients with submandibular gland carcinoma who underwent initial surgical treatment at the Kyoto University and its affiliated hospitals. RESULTS: The 3-year overall survival (OS), disease specific survival, locoregional control (LRC), and no distant metastasis (NDM) rates were 74.2%, 74.2%, 90.0%, and 64.8%, respectively. In the current follow-up study, 16 patients died of the disease, 5 patients were alive with recurrence, 43 patients were alive without disease, and 1 patient died of unrelated disease without recurrence. All patients who died of the disease had developed distant metastasis. Based on univariate analysis, tumor grade (high grade) and lymph node metastases (≥N2) were significant prognostic factors for OS and LRC. It also revealed tumor grade (high grade), T classification (≥T3), and lymph node metastases (≥N2) were significant for distant metastasis. Multivariate analysis showed the following significant prognostic factors: lymph node metastases (≥N2) for OS, LRC, and NDM, and high tumor grade for NDM. CONCLUSION: Our study suggested death of submandibular gland carcinoma occurred mainly due to distant metastasis. The significant predictors of distant metastasis were lymph node metastases (≥N2) and tumor grade (high grade).

Unknown primary squamous cell carcinoma of the head and neck: retrospective analysis of 80 cases.

OBJECTIVES: The management of patients with cervical metastasis in head and neck cancer of unknown primary (HNCUP) remains controversial. This current multicenter retrospective study investigated the treatment outcomes of patients with HNCUP. METHODS: The study included patients who were treated curatively at 12 institutions in Japan from January 2006 to December 2015. RESULTS: Eighty patients with HNCUP were included. The median follow-up period was 34 months. The three-year overall survival (OS), disease-specific survival (DSS), regional relapse-free survival (RRFS), local progression-free survival (LPFS), and distant metastasis-free survival (DMFS) rates were 72.5%, 80.3%, 74.0%, 89.7%, and 86.9%, respectively. Nodal status was a significant factor for OS, DSS, RRFS, and DMFS; and extracapsular extension (ECE) was significant for OS and DSS. There was a distinct difference between the survival rates of patients with N1-2a and N2b-3 disease. RT was a significant positive factor for LPFS (3-year LPFS, RT 93.0% vs. no RT 83.0%, p = .043). CONCLUSIONS: For N2a as well as N1 disease without ECE, a single treatment modality, including ND or RT alone is acceptable. When ND alone is performed, thorough monitoring should be continued during follow-up to identify the emergence of the primary lesion.

Survival in patients with parotid gland carcinoma - Results of a multi-center study.

BACKGROUND: Parotid gland carcinoma is a rare malignancy, comprising only 1-4% of head and neck carcinomas; therefore, it is difficult for a single institution to perform meaningful analysis on its clinical characteristics. The aim of this study was to update the clinical knowledge of this rare disease by a multi-center approach. METHODS: The study was conducted by the Kyoto University Hospital and Affiliated Facilities Head and Neck Clinical Oncology Group (Kyoto-HNOG). A total of 195 patients with parotid gland carcinoma who had been surgically treated with curative intent between 2006 and 2015 were retrospectively reviewed. Clinical results including overall survival (OS), disease-free survival (DFS), disease-specific survival (DSS), local control rate (LCR), regional control rate (RCR), and distant metastasis-free survival (DMFS) were estimated. Univariate and multivariate analyses were performed to identify prognostic factors. RESULTS: The median patient age was 63years old (range 9-93years), and the median observation period was 39months. The OS, DFS, DSS, LCR, RCR, and DMFS at 3years were 85%, 74%, 89%, 92%, 88%, and 87%, respectively. Univariate analysis showed age over 74, T4, N+, preoperative facial palsy, high grade histology, perineural invasion, and vascular invasion were associated with poor OS. N+ and high grade histology were independent factors in multivariate analysis. In subgroup analysis, postoperative radiotherapy was associated with better OS in high risk patients. CONCLUSION: Nodal metastases and high grade histology are important negative prognostic factors for OS. Postoperative radiotherapy is recommended in patients with advanced high grade carcinoma.

Adenoid cystic carcinoma of the head and neck: a retrospective multicenter study.

BACKGROUND: Adenoid cystic carcinoma of the head and neck (ACCHN) is rare and difficult to study effective treatment at one institute. Our aim is to identify prognostic factors for this disease by conducting a multicenter study at 11 institutions in Japan. METHODS: A retrospective multicenter study of ACCHN was performed. One hundred and three patients were identified between 2006 and 2015. The overall survival (OS) rate for all patients was calculated, and OS, locoregional control (LRC) rate, or no distant metastasis (NDM) rate was calculated for patients in that the surgery was performed without distant metastasis (DM). Statistical analyses were performed. RESULTS: A significant difference with multivariate analysis was observed in patients in sublingual glands, stage IV and the use of radiation therapy ≥60Gy (sufficient RT) in OS for all patients. A significant difference was observed in the use of sufficient postoperative RT in the OS and the LRC rate, and in pathological surgical margins in the NDM rate. CONCLUSION: Sublingual glands or stage IV was a poorer, and sufficient RT was a better prognostic factor for ACCHN. Sufficient RT was effective to prevent local recurrence after surgical resection. Positive surgical margins caused an increase in DM.

[Prognosis of Papillary Thyroid Carcinoma with Local Invasion].

OBJECTIVE: The aim of this study was to detect prognostic factors in patients with locally advanced papillary thyroid carcinoma. PATIENTS: The study included 72 patients (T4a/T4b 72/0, N0/N1a/N1b 25/15/32, M0/M1 68/4, mean follow-up 8.1 4.4 years) who underwent initial surgical treatment at Osaka Red Cross Hospital between April 1993 and April 2011. RESULTS: Eleven patients died of PTC, 3 patients with recurrence died of unrelated disease and 10 patients are alive with recurrence. The overall 5-year survival rate was 88.3%, and the 10-year survival rate was 73.4%. The disease-specific 5-year survival rate was 91.4%, and the 10-year survival rate was 88.6%. The 5-year local control rate was 94.1%, and the 10-year local control rate was 85.4%. Patients with distant metastasis (M1), tracheal invasion and/or multiple organs invasion showed a significantly worse disease-specific survival rate based on a univariate analysis, which also revealed that tracheal invasion, laryngeal invasion, esophageal invasion and multiple organs invasion were risk factors linked to the development of distant metastasis during follow-up (recurrence as distant metastasis). The following were found to be clinically significant risk factors, based on the multivariate analysis among tracheal invasion, laryngeal invasion, esophageal invasion and recurrent laryngeal nerve invasion : Tracheal invasion was a risk factor for disease-specific survival, and tracheal invasion and laryngeal invasion were risk factors for recurrence as distant metastasis. CONCLUSIONS: In this study, distant metastasis, multiple organs invasion, tracheal invasion and/or laryngeal invasion were shown to be higher risk factors.

8-(3-chloro-4-methoxybenzyl)-8H-pyrido[2,3-d]pyrimidin-7-one derivatives as potent and selective phosphodiesterase 5 inhibitors.

A novel series of highly selective phosphodiesterase 5 (PDE5) inhibitors was found. 8H-Pyrido[2,3-d]pyrimidin-7-one derivatives bearing an (S)-2-(hydroxymethyl)pyrrolidin-1-yl group at the 2-position and a 3-chloro-4-methoxybenzyl group at the 8-position exhibited potent PDE5 inhibitory activities and high PDE5 selectivity over PDE6. Among the synthesized compounds, the 5-methyl analogue (5b) showed the most potent relaxant effect on isolated rabbit corpus cavernosum with an EC30 value of 0.85 nM.

The discovery of avanafil for the treatment of erectile dysfunction: a novel pyrimidine-5-carboxamide derivative as a potent and highly selective phosphodiesterase 5 inhibitor.

Novel pyrimidine-5-carboxamide derivatives bearing a 3-chloro-4-methoxybenzylamino group at the 4-position were identified as potent and highly selective phosphodiesterase 5 inhibitors. Among them, we successfully found 10j (avanafil) which exhibited a potent relaxant effect on isolated rabbit cavernosum (EC30=2.1 nM) and a high isozyme selectivity.

Design and synthesis of novel 5-(3,4,5-trimethoxybenzoyl)-4-aminopyrimidine derivatives as potent and selective phosphodiesterase 5 inhibitors: scaffold hopping using a pseudo-ring by intramolecular hydrogen bond formation.

5-(3,4,5-Trimethoxybenzoyl)-4-amimopyrimidine derivatives were found as a novel chemical class of potent and highly selective phosphodiesterase 5 inhibitors. A pseudo-ring formed by an intramolecular hydrogen bond constrained the conformation of 3-chloro-4-methoxybenzylamino and 3,4,5-trimethoxybenzoyl substituents and led to the discovery of T-6932 (19a) with a potent PDE5 inhibitory activity (IC50 = 0.13 nM) and a high selectivity over PDE6 (IC50 ratio: PDE6/PDE5 = 2400). Further modification at the 2-position of T-6932 resulted in the finding of 26, which exhibited potent relaxant effects on isolated rabbit corpus cavernosum (EC30 = 11 nM) with a high PDE5 selectivity over PDE6 (IC50 ratio: PDE6/PDE5 = 2800).

MitoFish and MitoAnnotator: a mitochondrial genome database of fish with an accurate and automatic annotation pipeline.

Mitofish is a database of fish mitochondrial genomes (mitogenomes) that includes powerful and precise de novo annotations for mitogenome sequences. Fish occupy an important position in the evolution of vertebrates and the ecology of the hydrosphere, and mitogenomic sequence data have served as a rich source of information for resolving fish phylogenies and identifying new fish species. The importance of a mitogenomic database continues to grow at a rapid pace as massive amounts of mitogenomic data are generated with the advent of new sequencing technologies. A severe bottleneck seems likely to occur with regard to mitogenome annotation because of the overwhelming pace of data accumulation and the intrinsic difficulties in annotating sequences with degenerating transfer RNA structures, divergent start/stop codons of the coding elements, and the overlapping of adjacent elements. To ease this data backlog, we developed an annotation pipeline named MitoAnnotator. MitoAnnotator automatically annotates a fish mitogenome with a high degree of accuracy in approximately 5 min; thus, it is readily applicable to data sets of dozens of sequences. MitoFish also contains re-annotations of previously sequenced fish mitogenomes, enabling researchers to refer to them when they find annotations that are likely to be erroneous or while conducting comparative mitogenomic analyses. For users who need more information on the taxonomy, habitats, phenotypes, or life cycles of fish, MitoFish provides links to related databases. MitoFish and MitoAnnotator are freely available at http://mitofish.aori.u-tokyo.ac.jp/ (last accessed August 28, 2013); all of the data can be batch downloaded, and the annotation pipeline can be used via a web interface.

CentroidHomfold-LAST: accurate prediction of RNA secondary structure using automatically collected homologous sequences.

Although secondary structure predictions of an individual RNA sequence have been widely used in a number of sequence analyses of RNAs, accuracy is still limited. Recently, we proposed a method (called 'CentroidHomfold'), which includes information about homologous sequences into the prediction of the secondary structure of the target sequence, and showed that it substantially improved the performance of secondary structure predictions. CentroidHomfold, however, forces users to prepare homologous sequences of the target sequence. We have developed a Web application (CentroidHomfold-LAST) that predicts the secondary structure of the target sequence using automatically collected homologous sequences. LAST, which is a fast and sensitive local aligner, and CentroidHomfold are employed in the Web application. Computational experiments with a commonly-used data set indicated that CentroidHomfold-LAST substantially outperformed conventional secondary structure predictions including CentroidFold and RNAfold.

Identification of a high incidence region for retroviral vector integration near exon 1 of the LMO2 locus.

Therapeutic retroviral vector integration near the oncogene LMO2 is thought to be a cause of leukemia in X-SCID gene therapy trials. However, no published studies have evaluated the frequency of vector integrations near exon 1 of the LMO2 locus. We identified a high incidence region (HIR) of vector integration using PCR techniques in the upstream region close to the LMO2 transcription start site in the TPA-Mat T cell line. The integration frequency of the HIR was one per 4.46 x 10(4) cells. This HIR was also found in Jurkat T cells but was absent from HeLa cells. Furthermore, using human cord blood-derived CD34+ cells we identified a HIR in a similar region as the TPA-Mat T cell line. One of the X-linked severe combined immunodeficiency (X-SCID) patients that developed leukemia after gene therapy had a vector integration site in this HIR. Therefore, the descriptions of the location and the integration frequency of the HIR presented here may help us to better understand vector-induced leukemogenesis.

Ubiquitin-independent binding of Hrs mediates endosomal sorting of the interleukin-2 receptor beta-chain.

Several lines of evidence have revealed that ubiquitylation of membrane proteins serves as a signal for endosomal sorting into lysosomes or lytic vacuoles. The hepatocyte growth factor-regulated tyrosine kinase substrate (Hrs) interacts with ubiquitylated cargoes through its ubiquitin-interacting-motif domain (UIM domain), and plays an essential early role in endosomal sorting. Here, we show that the C-terminal region of Hrs, which does not contain the UIM domain, can bind to interleukin-2 receptor beta (IL-2Rbeta). We found a direct interaction between bacterially expressed IL-2Rbeta and Hrs in GST pull-down assays, indicating that their binding is independent of ubiquitin. Trafficking and degradation assays revealed that, similarly to wild-type IL-2Rbeta, an IL-2Rbeta mutant lacking all the cytoplasmic lysine residues is sorted from Hrs-positive early endosomes to LAMP1-positive late endosomes, resulting in degradation of the receptor. By contrast, an IL-2Rbeta mutant lacking the Hrs-binding region passes through early endosomes and is mis-sorted to compartments positive for the transferrin receptor. The latter mutant exhibits attenuated degradation. Taken together, these results indicate that precise sorting of IL-2Rbeta from early to late endosomes is mediated by Hrs, a known sorting component of the ubiquitin-dependent machinery, in a manner that is independent of UIM-ubiquitin binding.

Quantitation of human parvovirus B19 DNA by real-time polymerase chain reaction.

BACKGROUND: There have been no reports on either the serial quantification of genomic copies in the various parvovirus B19 infections or the comparison of the viral amount in erythema infectiosum, unlike with that in other parvovirus B19 infections. METHODS: A total of 19 children with parvovirus B19 infection were classified into a group of seven (group A) with erythema infectiosum and a group of 12 (group B) without erythema infectiosum, and their serum levels of parvovirus B19 DNA were quantified by real-time polymerase chain reaction. A total of 30 boys and girls with some symptoms but no parvovirus B19 infection served as a control group (group C). RESULTS: The amount of parvovirus B19 DNA differed significantly between groups A and C (P < 0.01) and between groups B and C (P < 0.01). The amount of viral DNA was significantly higher in group B than in group A (P < 0.01). Sequential determination showed that the amount of viral DNA in group B rapidly decreased over several days. Erythema infectiosum developed in two patients of group B on the 6th and 29th days after onset when the amount of viral DNA was similar to that in group A. CONCLUSIONS: The amount of parvovirus B19 DNA correlated well with the stage of infection, and its quantitation was useful for determining disease status and prognosis. In parvovirus B19 infection, the viremia is associated with rare but varied pathological states different from erythema infectiosum, such as transient aplastic crisis, hemophagocytic syndrome, lupus-like syndrome, and papular-purpuric gloves and socks syndrome.

Murine leukemia virus vector integration favors promoter regions and regional hot spots in a human T-cell line.

Genomic analysis of integration will be important in evaluating the safety of human gene therapy with retroviral vectors. Here, we investigated MLV vector integration sites in human T-cells, since they are amenable to gene transfer studies, and have been used therapeutically in clinical trials. We mapped 340 MLV vector integration sites in the infected human T-cell clones we established. The data showed that MLV preferred integration near the transcription start sites (+/-5kb), near CpG islands (+/-1kb), and within the first intron of RefSeq genes. We also identified MLV integration hot spots that contained three or more integrations within a 100kb region. RT-PCR revealed that mRNA-levels of T-cell clones that contained MLV integrations near transcription start sites or introns were dysregulated compared to the uninfected cells. These studies help define the profile of MLV integration in T-cells and the risks associated with MLV-based gene therapy.