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Objective: To report outcomes of multicenter series of penile cancer patients undergoing robot-assisted video endoscopic inguinal lymph node dissection (RA-VEIL). Materials and Methods: In this retrospective analysis from 3 tertiary care centers in India, consecutive intermediate-/high-risk carcinoma penis (CaP) patients with nonpalpable inguinal lymphadenopathy and/or nonbulky (<3 cm) mobile inguinal lymphadenopathy undergoing RA-VEIL were included. Patients with matted/bulky (>3 cm) and fixed lymphadenopathy were excluded. Demographic, clinical, and intraoperative data were recorded. Perioperative complications were graded by the Clavien-Dindo classification (CDC). The International Society of Lymphology (ISL) {0-III} grading was used for the assessment of lymphedema. Incidence and pattern of recurrences were assessed on follow-up. Results: From January 1, 2011, to September 30, 2023, 115 patients (230 groins) underwent bilateral RA-VEIL for CaP. The median age of the cohort was 60 (50-69) years. Clinically palpable (either unilateral or bilateral) inguinal lymphadenopathy was seen in 54 patients (47%). The "per groin" median operative time was 120 (100-140) minutes with median lymph node yield of 12 (9-16). No complications were recorded in 87.8% groins operated, with major complications (CDC 3) seen in 2.6% groins. At a median follow-up of 13.5 months, 13 patients had documented recurrences and there were 10 cancer-related deaths. No port-site recurrences were observed. No/minimal lymphedema (ISL 0/I) was seen in 94% legs. Conclusion: RA-VEIL demonstrates safety and oncologic efficacy in penile cancer patients presenting with clinically nonpalpable and/or nonbulky inguinal lymphadenopathy, with favorable functional outcomes.

Background: The incidence of prostate cancer is increasing worldwide. A significant proportion of patients develop metastatic disease and are initially prescribed androgen deprivation therapy (ADT). However, subsequent sequences of treatments in real-world settings that may improve overall survival remain an area of active investigation., Materials and Methods: Data were collected from 384 patients presenting with de novo metastatic prostate cancer from 2011 to 2015 at a tertiary cancer center. Patients were categorized into surviving (n = 232) and deceased (n = 152) groups at the end of 3 years. Modified sequence pattern mining techniques (Generalized Sequential Pattern Mining and Sequential Pattern Discovery using Equivalence Classes) were applied to determine the exact order of the most frequent sets of treatments in each group., Results: Degarelix, as the initial form of ADT, was uniquely in the surviving group. The sequence of ADT followed by abiraterone and docetaxel was uniquely associated with a higher 3-year overall survival. Orchiectomy followed by fosfestrol was found to have a unique niche among surviving patients with a long duration of response to the initial ADT. Patients who received chemotherapy followed by radiotherapy and those who received radiotherapy followed by chemotherapy were found more frequently in the deceased group., Conclusions: We identified unique treatment sequences among surviving and deceased patients at the end of 3 years. Degarelix should be the preferred form of ADT. Patients who received ADT followed by abiraterone and chemotherapy showed better results. Patients requiring palliative radiation and chemotherapy in any sequence were significantly more frequent in the deceased group, identifying the need to offer such patients the most efficacious agents and to target them in clinical trial design., Competing Interests: No conflict of interest has been declared by the author., (Copyright © 2023 The Authors. Published by Wolters Kluwer Health, Inc.)

Introduction: Transrectal ultrasound (TRUS) guided systematic prostate biopsy is conventionally used for the diagnosis of carcinoma prostate (CaP). However, magnetic resonance imaging (MRI) guided biopsies have been shown to have superior diagnostic performance. MRI-TRUS fusion biopsy improves the detection by combining the systematic and the targeted biopsies (TB). In this study, we evaluated the role of fusion biopsy in the detection of CaP as well as clinically significant carcinoma prostate (CsCaP)., Methods: In this retrospective study, the patients who underwent fusion biopsy from January 2016 to July 2022 were evaluated. Patients underwent multiparametric MRI and the suspicious lesions were reported as per the Prostate Imaging Reporting and Data System (PIRADS) version 2. The clinical, imaging, and biopsy parameters were recorded and evaluated., Results: A total of 330 patients with PIRADS ≥3 underwent MRI-TRUS fusion biopsy and prostate cancer was detected in 187 patients (56.67%). With an increase in the PIRADS score, there was a significant rise in the detection of CaP ( P < 0.001) and CsCaP ( P < 0.0000001). Prostatitis was observed in 13%-18.1% of the patients with a lesion on MRI irrespective of the PIRADS score. The systematic and TB were comparable for the detection of CaP ( P = 0.88) and CsCaP ( P = 0.26). With a prostate-specific antigen density (PSAD) cutoff of 0.15 ng/mL/cc and 0.22 ng/mL/cc, biopsy could be safely avoided in 14.2% and 20.3% of the patients, missing only 0.3% of CaP and 0.9% of CsCaP, respectively. Different subgroups based on PSA levels, prostate volume, lesion dimension, and PIRADS score did not show a significant difference between the systematic and the targeted cores for the detection of CsCaP., Conclusion: This single center study of MRI-TRUS fusion prostate biopsy shows that in men with clinical suspicion of prostate cancer a pre-biopsy MRI and MRI-TRUS fusion combined systematic and targeted prostate biopsy improves the detection of prostate cancer and CsCaP. Patients with a PIRADS 3 lesion with a PSA density <0.22 can safely avoid prostate biopsy, without a significant risk of missing clinically significant prostate cancer., Competing Interests: There are no conflicts of interest., (Copyright: © 2023 Indian Journal of Urology.)

Introduction: We retrospectively compared surgical and oncological outcomes of robot-assisted (RA) radical nephroureterectomy (RNU) in patients of upper-tract urothelial carcinoma with a cohort of patients who underwent the same procedure using a laparoscopic approach., Methods: Data of 63 consecutive patients who underwent RNU with bladder cuff excision (BCE) from 2011 to 2022 at a single tertiary care institution was retrospectively retrieved from the electronically maintained institutional database. Twenty-six cases underwent RNU with a laparoscopic approach, whereas 37 were done by RA approach. Demographic, clinical, surgical, and pathologic details and survival analyses were reported and compared. The tetrafecta of RNU, which include the performance of a BCE, lymphadenectomy, no positive surgical margin, and no major surgical complication, was also reviewed., Results: The mean age and body mass index of the robotic and laparoscopic groups were 61.5 years versus 62.7 years and 23.8 versus 24.9 kg/m2, respectively ( P = 0.710 and 0.309). The Charlson Comorbidity Index and upper-tract tumor site distribution were comparable between the groups. There was no significant difference in the distribution of T stage, N stage, presence of multifocality, or lymphovascular invasion between the two groups. Although the rate of concomitant carcinoma in situ was higher in laparoscopic cohort, 42.8% versus 10.8% in robotic cohort ( P = 0.004). The laparoscopic group had higher blood transfusion rates (50 vs. 13.5%, P = 0.002) and longer median hospital stays (7 vs. 4 days, P = 0.000). The median follow-up time was 21.5 versus 27 months in the laparoscopic and robotic groups. The RA group was significantly better in the achievement of the tetrafecta outcomes. The 5-year urinary bladder recurrence-free survival (UB RFS) and elsewhere RFS between the laparoscopic and robotic cohorts were 65% versus 72% and 56% versus 70%, respectively ( P = 0.510 and 0.190). The laparoscopic cohort had worse 5-year cancer-specific survival and overall survival (64% vs. 90% and 58% vs. 74%, P = 0.04 and 0.08)., Conclusion: The robotic approach to RNU and BCE has significantly lower transfusion rates, lower hospital stays, and significantly better cancer-specific survival rates., Competing Interests: There are no conflicts of interest., (Copyright: © 2023 Indian Journal of Urology.)

Background: Sunitinib remains the first-line treatment for favorable risk metastatic clear cell renal cell cancer (mccRCC). It was conventionally given in the 4/2 schedule; however, toxicity necessitated trying the 2/1 regimen. Regional variations in treatment response and toxicity are known, and there is no data from the Indian subcontinent about the outcomes of the alternative dosing schedule., Methods: Clinical records of all consecutive adult patients who received sunitinib as first-line therapy for histologically proven mccRCC following cytoreductive nephrectomy from 2010-2018 were reviewed. The primary objective was to determine the progression-free survival (PFS), and the secondary objectives were to evaluate the response rate (objective response rate and clinical benefit rate), toxicity, and overall survival. A list of variables having a biologically plausible association with outcome was drawn and multivariate inverse probability treatment weights (IPTW) analysis was done to determine the absolute effect size of dosing schedules on PFS in terms of "average treatment effect on the treated" and "potential outcome mean.", Results: We found 2/1 schedule to be independently associated with higher PFS on IPTW analysis such that if every patient in the subpopulation received sunitinib by the 2/1 schedule, the average time to progression was estimated to be higher by 6.1 months than the 4/2 schedule. We also found 2/1 group to have a lower incidence than the 4/2 group for nearly all ≥ grade 3 adverse effects. Other secondary outcomes were comparable between both treatment groups., Conclusion: Sunitinib should be given via the 2/1 schedule in Indian patients., (Copyright © 2024 Copyright: © 2024 Indian Journal of Cancer.)

Cutaneous radiation-associated angiosarcoma (cRAA) is a rare and aggressive secondary cutaneous angiosarcoma (cAS) with poor survival. cRAA has been mostly reported in breast carcinoma patients. Owing to its rarity, there is scanty literature available and no treatment guidelines. To the best of our knowledge, this is the first report of cRAA after multimodality treatment of carcinoma penis. A sixty-eight-year-old gentleman, a known case of carcinoma penis, underwent total penectomy with perineal urethrostomy and bilateral radical inguinopelvic lymph node dissection 6 years ago. He received adjuvant radiotherapy to the pelvis and bilateral groin. He presented with a bleeding plaque-like lesion with ulceration over the left lower abdomen (within previous radiation field) which rapidly progressed in size over the past 2 months. On examination, the lesion bled profusely on touch. Contrast MRI was suggestive of lobulated exophytic enhancing cutaneous lesion free from underlying muscle. Wedge biopsy was suggestive of cutaneous angiosarcoma. He underwent wide local excision with local perforator flap reconstruction from the right lower abdomen. Histopathology was suggestive of cutaneous angiosarcoma which showed immunoexpression of CD31, ERG1, cMYC suggestive of cRAA. cRAA is a very aggressive disease with 5-year survival of 15-34%. To the best of our knowledge, this is the first ever reported case of cRAA of lower abdomen after multimodality management of carcinoma penis. It masquerades with other benign and less aggressive radiation-induced skin lesions. cMYC immunoexpression is specific for secondary cAS. Wide local resection with negative margin provides the best outcome., Competing Interests: Conflict of InterestThe authors declare no conflict of interests., (© The Author(s), under exclusive licence to Indian Association of Surgical Oncology 2023. Springer Nature or its licensor (e.g. a society or other partner) holds exclusive rights to this article under a publishing agreement with the author(s) or other rightsholder(s); author self-archiving of the accepted manuscript version of this article is solely governed by the terms of such publishing agreement and applicable law.)

Genetic purity of seeds is one of the critical aspects in the seed industry. Molecular seed testing laboratories are utilizing PCR based diagnostic tools for genetic purity analysis. High quality DNA is an essential prerequisite for such analyses. Here, we demonstrate a robust and inexpensive DNA extraction method to isolate genomic DNA from variety of crops. Current method (M2) was compared with four commonly used DNA isolation methods for PCR-based genetic characterization and High Resolution Melt (HRM) based hybridity analysis of cotton, okra, tomato and maize using SSR markers. DNA extracted through current method showed excellent yield and quality as compared to other methods. High quality, PCR ready DNA was isolated within 30-50 min and displayed best results for genetic purity analysis using HRM. In contrast, several genomic DNA samples extracted using other methods were found unsuitable for HRM analysis. Our method can be a perfect choice in seed industry, where thousands of samples are processed every day. Notably, using our method single technician can extract DNA from 96 leaf samples within 30-50 min, at a cost of only $0.11/sample. Overall, current DNA extraction method is a reliable and cost-effective solution for large-scale genotyping experiments in the agricultural industry., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier Inc. All rights reserved.)

Competing Interests: There are no conflicts of interest.

Objectives: To present our intermediate to long-term oncological and functional outcomes of robot-assisted retroperitoneal lymph node dissection (RA-RPLND) in post-chemotherapy (PC) residual mass in testicular cancers. To the best of our knowledge, this is the largest single-centre experience of RA-RPLND for in such setting., Methods: Prospectively maintained database of carcinoma testis patients undergoing RA-RPLND from February 2012 to September 2021 was reviewed. Patient demographics, tumour stage and risk groups and chemotherapy details were recorded. Intraoperative details and post-operative complications were also noted. Pathological outcomes included were lymph node yield and histopathology report. Further, follow-up was done for recurrence and antegrade ejaculation status., Results: Total of 37 cases were done for PC residual masses. International germ cell cancer collaborative group good, intermediate and poor risk proportion was 18 (48.6%), 14 (37.8%) and 5 (13.5%), respectively. Bilateral full template dissection, unilateral modified template dissection and residual mass excision was performed in 59.5% (22/37), 35.1% (13/37) and 5.4% (2/37) patients, respectively. The median size of the excised residual mass was 3.45 cm interquartile range (IQR 2-6 cm), with the largest being 9 cm. The median lymph nodal yield was 19. The most common histology was necrosis (n = 24, 65%), followed by teratoma (n = 11, 30%) and viable malignancy (n = 2, 5%). Antegrade ejaculation was reported in 32 patients (86.4%). After a median follow-up of 41 (IQR 14-64) months, only one patient had a recurrence., Conclusions: RA-PC-RPLND is thus a safe, feasible and oncologically effective option for selected patients. With increasing experience, larger masses can also be dealt with efficiently., Competing Interests: None

Purpose: Despite widespread acceptance of robotics in urology, literature on using the minimally invasive approach for management of post robotic surgical complications is limited. Here we describe our experience with tips and tricks for robotic re-exploration of post-operative in house complications following robotic pelvic uro-oncologic surgery., Methods: A retrospective query of prospectively maintained database was done for all patients who underwent robotic - radical cystoprostatectomy (RCP, 437 patients) and radical prostatectomy (RP, 649 patients), from Jan 2015 or March 2021. Clinical details were collected for all who underwent a second robotic procedure during the same hospital admission for any complication related to the primary surgery., Results: Following RCP, 5 patients were re-explored for intestinal obstruction. Surgery was successfully completed in all with a median console time of 80 minutes. Median time to the passage of flatus and discharge from hospital following relook surgery was 3 and 6 days, respectively. Following RP, 3 patients underwent robotic re-exploration (two for reactionary hemorrhage, one for rectal injury). All three cases were managed with a median console time of 75 minutes. Robotic re-exploration was accomplished without extending the skin incision of the index surgery and we did not find an increased incidence of infectious or wound related complications., Conclusion: Robotic re-exploration for select post robotic urologic pelvic oncology surgery complications in the immediate and early post-operative period is feasible in the hands of experienced surgeons. Our experience can help others adopt robotics in such scenarios., Competing Interests: None

Aims: This study aims to assess the survival and identify the prognostic factors in ovarian cancer patients treated with surgery and carboplatin/paclitaxel based first-line chemotherapy (CT)., Settings and Design: The electronic medical records of all ovarian cancer patients registered during January 2009 and December 2017 were screened retrospectively., Subjects and Methods: A total of 440 cases were included in accordance with the inclusion/exclusion criteria of study. The comprehensive data regarding demography, treatment, chemotoxicities, recurrence, and others were collated and analyzed., Statistical Analysis Used: Cox regression analysis was used for univariate and multivariate analyses of prognostic factors., Results: The median age at diagnosis was 50.6 years. All cases had got CT-related morbidity but no associated mortality. The median recurrence-free survival (RFS) and mean overall survival (OS) were 30 (95% confidence interval [CI]: 24.65-35.38) months and 40.4 months, respectively. A significant difference was observed among the RFS (P < 0.001); and OS (P = 0.036) in relation to the stage of disease. Furthermore, patients who relapsed post first-line CT had 36%, 9%, 3% recurrence in second-, third-, and fourth-line CT regimens, respectively. Multivariate analysis proved the histology, low-grade serous, to be the favorable prognostic factor for RFS (hazard ratio = 0.18; 95% CI: 0.04-0.82)., Conclusions: Surgery and first-line CT with carboplatin/paclitaxel lead-to-moderate long-term survival in ovarian cancer. The likelihood of relapse is fairly high as stage advances. Low-grade serous histology is an independent prognostic factor for RFS., Competing Interests: None

Inguinal lymph node status is the single most important prognostic factor for survival in patients with carcinoma penis. Various modifications and alternatives to open inguinal lymph node dissection have been developed as the same is associated with high postoperative morbidity such as wound infection, skin flap necrosis, lymphorrhea, and lymphedema. Robot-assisted video endoscopic inguinal lymph node dissection (RA-VEIL) has the potential to accomplish thorough inguinal lymph node dissection with definitively reduced postoperative morbidity. In this video, we demonstrate our technique of RA-VEIL: The fascia lata first approach and highlight our technical modifications of the conventionally described procedure.

Objectives: To compare overall survival (OS) between adjuvant radiation, chemotherapy and chemoradiation (CCRT) postsurgery for node-positive patients with carcinoma penis., Methods: Prospectively maintained registry for 45 patients receiving adjuvant treatment following lymph node dissection from 2011 to 2017, having minimum 6 months follow-up and more than 2 positive inguinal nodes was analyzed. Patients without pelvic nodal positivity (n= 32) were treated by radiotherapy (RT) (n = 25) or chemotherapy (n = 7); CCRT (n = 6) or chemotherapy (n = 7) was used in patients with positive pelvic nodes (n = 13). Data was collected for age, comorbidities, body mass index, tobacco exposure, treatment modality, tumor grade, pathological T and N stage, and extra-nodal extension. OS was compared between different treatment modalities stratifying patients with and without pelvic nodal positivity. Multivariate cox proportional hazard analysis was used to narrow down remaining variables and Inverse Probability Treatment Weights modeling was used to determine average treatment effect., Results: About 12 of 14 patients in the chemotherapy group received both cisplatin and paclitaxel. Pathological T stage, N stage and extra-nodal extension had significant association with OS on multivariate analysis. Among patients with nodal positivity restricted to groin the estimated average OS when all patients received adjuvant RT was 1,438 days (95% confidence interval [CI] 1,256-1,619 days, Pvalue <0.0001). The estimated average OS if all patients received chemotherapy was lower by 1,007 days (95% CI 810-1,202 days, P value <0.0001). Among patients with positive pelvic nodes the estimated average OS when all patients received adjuvant CCRT was 467 days (95% CI 368-566 days, P value <0.0001). The estimated average OS difference if all patients received chemotherapy was 17 days (95% CI -144 to 178 days, Pvalue 0.21)., Conclusion: In patients with nodal positivity limited to groin, adjuvant RT proved superior to chemotherapy. Among patients with pelvic nodal positivity, CCRT offers no significant OS advantage over combination chemotherapy., Competing Interests: Conflict of interest The authors declare that they have no conflict of interest., (Copyright © 2020 Elsevier Inc. All rights reserved.)

Objectives: To document the management of advanced prostate cancer including diagnosis, prognosis, treatment, and care, in real-world practice in Asia using the United in Fight against prOstate cancer (UFO) registry., Patients and Methods: We established a multi-national, longitudinal, observational registry of patients with prostate cancer presenting to participating tertiary care hospitals in eight Asian countries. A total of 3636 eligible patients with existing or newly diagnosed high-risk localised prostate cancer (HRL), non-metastatic biochemically recurrent prostate cancer (M0), or metastatic prostate cancer (M1), were consecutively enrolled and are being followed-up for 5 years. Patient history, demographic and disease characteristics, treatment and treatment decisions, were collected at first prostate cancer diagnosis and at enrolment. Patient-reported quality of life was prospectively assessed using the European Quality of Life-five Dimensions, five Levels (EQ-5D-5L) and Functional Assessment of Cancer Therapy for Prostate Cancer questionnaires. In the present study, we report the first interim analysis of 2063 patients enrolled from study start (15 September 2015) until 18 May 2017., Results: Of the 2063 enrolled patients, 357 (17%), 378 (19%), and 1328 (64%) had HRL, M0 or M1 prostate cancer, respectively. The mean age at first diagnosis was similar in each group, 56% of all patients had extracapsular extension of their tumour, 28% had regional lymph node metastasis, and 53% had distant metastases. At enrolment, 62% of patients had at least one co-morbidity (mainly cardiovascular disease or diabetes), 91.8% of M1 patients had an Eastern Cooperative Oncology Group performance score of <2 and the mean EQ-5D-5L visual analogue score was 74.6-79.6 across cohorts. Treatment of M1 patients was primarily with combined androgen blockade (58%) or androgen-deprivation therapy (either orchidectomy or luteinising hormone-releasing hormone analogues) (32%). Decisions to start therapy were mainly driven by treatment guidelines and disease progression. Decision to discontinue therapy was most often due to disease progression (hormonal drug therapy) or completion of therapy (chemotherapy)., Conclusion: In the UFO registry of advanced prostate cancer in Asia, regional differences exist in prostate cancer treatment patterns that will be explored more deeply during the follow-up period; prospective follow-up is ongoing. The UFO registry will provide valuable descriptive data on current disease characteristics and treatment landscape amongst patients with prostate cancer in Asia., (© 2019 The Authors BJU International Published by John Wiley & Sons Ltd on behalf of BJU International.)

Objective: Levels of proinflammatory (TNF A) and anti-inflammatory (IL-10) cytokines play a key role in the progression of inflammation as well as cancer disease. We were investigating the potential association of single-nucleotide polymorphisms (SNPs)/haplotypes in proinflammatory (TNF A) and anti-inflammatory (IL-10) cytokines locus with the development of PCa in Indian population., Materials and Methods: We had genotyped 235 BPH/PCa samples (130 BPH and 105 cancer) along with 115 control samples for proinflammatory (TNF A -238G/A and -308G/A) and anti-inflammatory (IL-10 -1082A/G, -819C/T and -592C/A) cytokines SNPs in the gene promoter region using ARMS-PCR method., Results: Allelic frequencies of TNF A and IL-10 SNPs were found to be significantly associated with the risk of prostate cancer and BPH when compared to controls (p = 0.05). Further haplotypic analysis showed that two haplotypes of TNF A (AG and AA) and IL-10 gene (CCG and CTG) were serving as risk haplotypes for prostate cancer development. IL-10 risk haplotypes were found to be positively associated with aggressiveness of prostate cancer. We also noticed successively increasing percentage of TNF A and IL-10 risk haplotypes with life style habits like smoking (10 and 26%) and alcohol consuming (9 and 27%)., Conclusions: According to our data, TNF A -238G>A and IL-10 -1082A>G, -819C>T and -592C>A may be associated with the development of prostate cancer and BPH. We could also notice higher frequency of TNF A and IL-10 risk haplotypes in smoker and alcohol user. Interestingly, IL-10 risk haplotype was positively associated with aggressiveness of tumor. This information can be used for the early diagnosis of disease and to improve tissue-specific treatment's efficacy which will be moving ultimately towards the discovery of personalized therapy.

Testicular germ cell tumors (GCTs) comprise 2% of all human male malignancies and are the most common solid tumors in men between ages 15 and 35 years. Risk of contralateral testicular GCT is between 1 and 5%. Partial orchidectomy (PO) was originally described in 1984 by Richie. The evolving indications include metachronous tumors and tumor in solitary testicles. Also, small non-palpable lesions detected only by ultrasonography (USG) in asymptomatic patients is another indication. Salvagability is only chosen for tumors less than 2 cm in size. The key feature of PO is an inguinal approach with early vascular control using a rubber tourniquet before testicular mobilization into the field to avoid systemic tumor seeding. After, mass excision with a margin mandatory frozen section is done to assess adequacy of resection. Intra-op USG may be beneficial in small non-palpable lesions. Post op tumor markers are assessed and patients are taught self-examination of testis. Recent series shows that PO is safe and gives adequate oncological control. Carcinoma in situ (CIS) in the affected testis at PO or after testicular sparing surgery remains a challenge. At most centers, 20 Gy is recommended when adjuvant local radiation treatment is chosen to treat CIS. But this dose may hamper Androgen production. Radical orchiectomy remains the gold standard and should be discussed as part of informed consent. It is mandatory to highlight the risks of local recurrence and CIS, and treatment (observation, radiation, or completion orchiectomy) as well as the need for androgen supplementation and fertility risks before choosing testicular salvage procedures., (© Indian Association of Surgical Oncology 2016.)

Glycosyl hydrolase family 1 β-glucosidases are important enzymes that serve many diverse functions in plants including defense, whereby hydrolyzing the defensive compounds such as hydroxynitrile glucosides. A hydroxynitrile glucoside cleaving β-glucosidase gene (Llbglu1) was isolated from Leucaena leucocephala, cloned into pET-28a (+) and expressed in E. coli BL21 (DE3) cells. The recombinant enzyme was purified by Ni-NTA affinity chromatography. The optimal temperature and pH for this β-glucosidase were found to be 45 °C and 4.8, respectively. The purified Llbglu1 enzyme hydrolyzed the synthetic glycosides, pNPGlucoside (pNPGlc) and pNPGalactoside (pNPGal). Also, the enzyme hydrolyzed amygdalin, a hydroxynitrile glycoside and a few of the tested flavonoid and isoflavonoid glucosides. The kinetic parameters K (m) and V (max) were found to be 38.59 μM and 0.8237 μM/mg/min for pNPGlc, whereas for pNPGal the values were observed as 1845 μM and 0.1037 μM/mg/min. In the present study, a three dimensional (3D) model of the Llbglu1 was built by MODELLER software to find out the substrate binding sites and the quality of the model was examined using the program PROCHEK. Docking studies indicated that conserved active site residues are Glu 199, Glu 413, His 153, Asn 198, Val 270, Asn 340, and Trp 462. Docking of rhodiocyanoside A with the modeled Llbglu1 resulted in a binding with free energy change (ΔG) of -5.52 kcal/mol on which basis rhodiocyanoside A could be considered as a potential substrate.

Background: Intetumumab is a fully human mAb with antiangiogenic, antitumor properties which has shown potential therapeutic effect in castration-resistant prostate cancer (CRPC) patients., Patients and Methods: In a phase 2, randomized, double-blind, multicenter study, men with metastatic CRPC without prior systemic nonhormonal therapy were randomly assigned to 75-mg/m(2) docetaxel (Taxotere) and 5-mg prednisone plus placebo (N = 65) or 10-mg/kg intetumumab (N = 66) q3w. Placebo patients with progressive disease (PD) could cross over to 10-mg/kg intetumumab alone or with docetaxel. The primary end-point was progression-free survival (PFS). The secondary end-points included tumor response (complete response + partial response, CR + PR), prostate-specific antigen (PSA) response, and overall survival (OS)., Results: All efficacy end-points favored placebo over intetumumab, including PFS (median 11.0 versus 7.6 months, P = 0.014), tumor response (20% versus 16%, P = 0.795), PSA response (68% versus 47%, P = 0.018), OS (median 20.6 versus 17.2 months, P = 0.163). Common all-grade adverse events (AEs) with placebo and intetumumab were alopecia (43% versus 26%); diarrhea, leukopenia (both 34% versus 27%); neutropenia (35% versus 23%). Grade ≥ 3 leukopenia (28% versus 17%) and neutropenia (26% versus 18%) occurred more often with placebo than with intetumumab. Intetumumab serum concentrations increased with repeated dosing and did not reach steady-state. Greater decreases in N-telopeptide of type I collagen (NTx), C-telopeptide (CTx) and CTCs occurred with intetumumab than with placebo., Conclusion: The addition of intetumumab to docetaxel resulted in shorter PFS without additional toxicity among CRPC patients.

Aim: To analyze the influence of computed tomography (CT)-guided 3D conformal interstitial brachytherapy (3D-IBT) boost for better coverage of target volumes and study the diametric impact to organ at risk in patients with post-surgery recurrent and residual cervical cancer., Material and Methods: Fourteen consecutive patients with recurrent or residual cervical cancer who were treated with interstitial brachytherapy as a boost were included in the study. All patients received 50.4 Gy external radiation (EBRT) to whole pelvis with conformal technique to reduce the dose to bowel. The clinical target volume (CTV) and organs at risk were contoured on CT scan with gold seeds being a surrogate marker of initial tumor extent implanted before commencing treatment. The median dose of prescription was 10.5-12Gy in 3 fractions. Dose volume histogram was calculated to evaluate the dose that covers 100% and 90% of the target volume and dose to the bladder, rectum and bowel (2 mL, 1 mL volume)., Results:   The median follow-up was 12 months (range 6-18). The doses to CTV (D90, D100) ranged from 1141 to 2014 cGy, and 585 to 969 cGy, respectively. The mean cumulative 2-mL rectal, bladder and bowel doses were 66.70, 73.15 and 61.01Gy, respectively. Rectal toxicity of grade 2 or more had a strong correlation with the dose delivered (Spearman's correlation, 0.950). The local control rate at one year was 92% with failure seen in one patient only., Conclusion: Conformal EBRT supplemented with 3D-IBT seems to be a practical and appropriate approach to give the most optimal therapeutic benefit with the least side-effects in postoperative recurrent and residual cervical cancer patients., (© 2012 The Authors. Journal of Obstetrics and Gynaecology Research © 2012 Japan Society of Obstetrics and Gynecology.)

cDNA and genomic clones of cinnamoyl CoA reductase measuring 1011 and 2992 bp were isolated from a leguminous pulpwood tree Leucaena leucocephala, named as LlCCR. The cDNA exhibited 80-85% homology both at the nucleotide and amino acid levels with other known sequences. The genomic sequence contained five exons and four introns. Sense and antisense constructs of LlCCR were introduced in tobacco plants to up and down-regulate this key enzyme of lignification. The primary transformants showed a good correlation between CCR transcript levels and its activity. Most of the CCR down-regulated lines displayed stunted growth and development, wrinkled leaves and delayed senescence. These lines accumulated unusual phenolics like ferulic and sinapic acids in cell wall. Histochemical staining suggested reduction in aldehyde units and increased syringyl over guaiacyl (S/G) ratio of lignin. Anatomical studies showed thin walled, elongated xylem fibres, collapsed vessels with drastic reduction of secondary xylem. The transmission electron microscopic studies revealed modification of ultrastructure and topochemical distribution of wall polysaccharides and lignin in the xylem fibres. CCR down-regulated lines showed increased thickness of secondary wall layers and poor lignification of S2 and S3 wall layers. The severely down-regulated line AS17 exhibited 24.7% reduction of Klason lignin with an increase of 15% holocellulose content. Contrarily, the CCR up-regulated lines exhibited robust growth, development and significant increase in lignin content. The altered lignin profiles observed in transgenic tobacco lines support a role for CCR down-regulation in improving wood properties of L. leucocephala exclusively used in the pulp and paper industry of India.

Removal of lignin is a major hurdle for obtaining good quality pulp. Leucaena leucocephala (subabul) is extensively used in paper industry in India; therefore, as a first step to generate transgenic plants with low lignin content, cDNA and genomic clones of CCR gene were isolated and characterized. The cDNA encoding CCR (EC 1.2.1.44) was designated as Ll-CCR; the sequence analysis revealed an Open Reading Frame (ORF) of 1005 bp. Phylogenetic analysis showed that Ll-CCR sequence is highly homologous to CCRs from other dicot plants. The 2992 bp genomic clone of Leucaena CCR consists of 5 exons and 4 introns. The haploid genome of L. leucocephala contains two copies as revealed by DNA blot hybridization. Ll-CCR gene was over-expressed in Escherichia coli, which showed a molecular mass of approximately 38 kDa. Protein blot analysis revealed that Ll-CCR protein is expressed at higher levels in root and in stem, but undetectable in leaf tissues. Expression of CCR gene in Leucaena increased up to 15 d in case of roots and stem as revealed by QRT-PCR studies in 0-15 d old seedlings. ELISA based studies of extractable CCR protein corroborated with QRT-PCR data. CCR protein was immuno-cytolocalized around xylem tissue. Lignin estimation and expression studies of 5, 10 and 15 d old stem and root suggest that CCR expression correlates with quantity of lignin produced, which makes it a good target for antisense down regulation for producing designer species for paper industry., (Copyright © 2010 Elsevier Masson SAS. All rights reserved.)

Leucaena leucocephala is a fast growing multipurpose legume tree used for forage, leaf manure, paper and pulp. Lignin in Leucaena pulp adversely influences the quality of paper produced. Developing transgenic Leucaena with altered lignin by genetic engineering demands an optimized regeneration system. The present study deals with optimization of regeneration system for L. leucocephala cv. K636. Multiple shoot induction from the cotyledonary nodes of L. leucocephala was studied in response to cytokinins, thidiazuron (TDZ) and N(6)-benzyladenine (BA) supplemented in half strength MS (½-MS) medium and also their effect on in vitro rooting of the regenerated shoots. Multiple shoots were induced from cotyledonary nodes at varied frequencies depending on the type and concentration of cytokinin used in the medium. TDZ was found to induce more number of shoots per explant than BA, with a maximum of 7 shoots at an optimum concentration of 0.23 µM. Further increase in TDZ concentration resulted in reduced shoot length and fasciation of the shoots. Liquid pulse treatment of the explants with TDZ did not improve the shoot production further but improved the subsequent rooting of the shoots that regenerated. Regenerated shoots successfully rooted on ½-MS medium supplemented with 0.54 µM α-naphthaleneacetic acid (NAA). Rooted shoots of Leucaena were transferred to coco-peat and hardened plantlets showed ≥ 90 % establishment in the green house.

Caffeoyl coenzyme A O-methyltransferase (CCoAOMT) is an important enzyme that participates in lignin biosynthesis especially in the formation of cell wall ferulic esters of plants. It plays a pivotal role in the methylation of the 3-hydroxyl group of caffeoyl CoA. Two cDNA clones that code CCoAOMT were isolated earlier from subabul and in the present study; 3D models of CCoAOMT1 and CCoAOMT2 enzymes were built using the MODELLER7v7 software to find out the substrate binding sites. These two proteins differed only in two amino acids and may have little or no functional redundancy. Refined models of the proteins were obtained after energy minimization and molecular dynamics in a solvated water layer. The models were further assessed by PROCHECK, WHATCHECK, Verify&#95;3D and ERRAT programs and the results indicated that these models are reliable for further active site and docking analysis. The refined models showed that the two proteins have 9 and 10 alpha-helices, 6 and 7 beta-sheets respectively. The models were used for docking the substrates CoA, SAM, SAH, caffeoyl CoA, feruloyl CoA, 5-hydroxy feruloyl CoA and sinapyl CoA which showed that CoA and caffeoyl CoA are binding with high affinity with the enzymes in the presence and absence of SAM. It appears therefore that caffeoyl CoA is the substrate for both the isoenzymes. The results also indicated that CoA and caffeoyl CoA are binding with higher affinity to CCoAOMT2 than CCoAOMT1. Therefore, CCoAOMT2 conformation is thought to be the active form that exists in subabul. Docking studies indicated that conserved active site residues Met58, Thr60, Val63, Glu82, Gly84, Ser90, Asp160, Asp162, Thr169, Asn191 and Arg203 in CCoAOMT1 and CCoAOMT2 enzymes create the positive charge to balance the negatively charged caffeoyl CoA and play an important role in maintaining a functional conformation and are directly involved in donor-substrate binding.

An approximately 4.9 kb Sau3A I genomic DNA fragment from the Streptomyces aureofaciens NRRL 2209 aiding in the biosynthesis of PHB in recombinant Escherichia coli has been sequenced and analysed for phaC gene. The putative phaC(Sa) gene of 2 kb is 79.1% GC rich and encodes a 63.5 kDa protein. It expressed under its own promoter and significant PHA synthase activity was detected in the recombinant E. coli. This is the first putative PHA synthase gene reported from a Streptomyces sp. with serine as the active nucleophile in the conserved lipase box. The phaC(Sa) was found in close proximity to a regulatory gene, which apparently regulated the phaC expression.

Purpose: Prostate-specific antigen (PSA) test has become a widely used screening test in prostate cancer (CaP). However, low specificity of serum PSA leads to many false-positive and false-negative results and clinical uncertainty. Development of CaP-specific diagnostic and prognostic markers is needed. Detection of circulating PSA-expressing cells (CPECs) in blood and bone marrow of CaP patients has potential in molecular diagnosis and prognosis. Our novel observations of the frequent presence of CPECs in CaP patients with organ-confined disease by reverse transcription (RT)-PCR-PSA assay in epithelial cells enriched from peripheral blood (ERT-PCR/PSA) have led us to test the hypothesis that CPECs have diagnostic potential for CaP., Experimental Design: Epithelial cells from peripheral blood of radical prostatectomy patients or prostate biopsy patients were isolated using antiepithelial cell antibody, Ber-EP4-coated magnetic beads, and total RNA specimens from these cells were analyzed for PSA expression by RT-PCR., Results: Peripheral blood specimens of 108 of 135 (80.0%) CaP patients were positive in ERT-PCR/PSA assay. Peripheral blood specimens from 45 control men were virtually negative (97.8%). In the blinded investigation, 84 patients who had biopsy for suspicion of CaP were evaluated by ERT-PCR/PSA assay. Eighteen of 22 (81.8%) patients with biopsy-proven CaP were positive, and 54 of 62 (87.1%) patients with biopsy negative for CaP were negative in this assay (P < 0.001)., Conclusions: Our study provides intriguing novel results showing that the majority of patients with clinically organ-confined CaP contain CPECs. Strong concordance between the biopsy results and ERT-PCR/PSA assay (sensitivity 81.8%; specificity 87.1%) suggests a potentially new diagnostic application of this type of assay in CaP diagnosis.

Recombinant Escherichia coli (ATCC:PTA-1579) harbouring poly(3-hydroxybutyrate) (PHB) synthesising genes from Streptomyces aureofaciens NRRL 2209 accumulates PHB. Effects of different carbon and nitrogen sources on PHB accumulation by recombinant E. coli were studied. Among the carbon sources used glycerol, glucose, palm oil and ethanol supported PHB accumulation. No PHB accumulated in recombinant cells when sucrose or molasses were used as carbon source. Yeast extract, peptone, a combination of yeast extract and peptone, and corn steep liquor were used as nitrogen sources. The maximum PHB accumulation (60% of cell dry weight) was measured after 48 h of cell growth at 37 degrees C in a medium with glycerol as the sole carbon source, and yeast extract and peptone as nitrogen sources. Scanning electron microscopy of the PHB granules isolated from recombinant E. coli revealed these to be spherical in shape with a diameter ranging from 0.11 to 0.35 pm with the mean value of 0.23 +/- 0.06 pm.

An approximately 5.0 kb Sau3A I genomic DNA fragment from Streptomyces aureofaciens NRRL 2209 was cloned in a plasmid vector and introduced into Escherichia coli. The recombinant E. coli accumulated polyhydroxyalkanoates (PHAs) as cytoplasmic inclusions. The accumulated PHA was identified as the isotactic homopolymer of PHB with a molecular weight of 2.85x10(5). Purified PHB granules were spherical with an average size of 1.1 microm and of stable configuration. DSC thermogram suggested high crystalline nature of the polymer. Maximum thermal degradation of the biopolymer occurred between 250 and 340 degrees C. Recombinant E. coli cells preferentially utilized glycerol as the carbon source and accumulated 25-28 times more PHB than the native S. aureofaciens.

Immature zygotic embryo explants of Carica papaya were cultured on MS medium supplemented with 2,4-D (2.0 mg/l) and formed globular embryos on explants without callus formation in 4-6 weeks. Maturation and conversion of somatic embryos was also achieved on the same medium. Cotyledonary stage embryos germinated to 63.66 and 68.33% in cv. honey dew and washington respectively in MS basal medium supplemented ABA (0.5 microm/l). Robust development and proliferation of plantlet roots in vitro was obtained on MS basal medium. Hardened plantlets have 60% survival rate.

Taxol is perhaps the most successful drug used for the treatment of various cancers. Comprehensive literature accumulated on therapeutics of the drug has indicated numerous side effects. In this paper, by use of fluorescence spectroscopy, it is shown that taxol binds to DNA with an affinity constant (Ka) of 1.08 x 10(7) M-1. This binding is accompanied by a large 'red edge excitation shift' (REES) of fluorescence emission maximum in taxol-DNA complex. The results point to an interaction of taxol with its core eight-membered ring in the DNA groove and the three phenyl rings projecting away from the DNA. The drug encompasses about two base pairs of DNA upon binding to it. Systematic studies with taxol analogues confirms such a mode of binding. These interesting findings on hitherto unknown taxol-DNA interactions may have clinical implications in view of its large number of side effects and pharmacokinetics.

The developmental patterns of the nitrogen assimilating enzymes were investigated and compared in the non-shoot forming and the shoot forming callus cultures of sugarane. In the shoot forming tissue the pre-emergence period od shoots was characterised by increasing activities of glutamine synthetase and glutamate synthase. The activity of these enzymes during the corresponding period in the non-shoot forming callus was found to decline. Although the activity of glutamate dehydrogenase in the shoot forming callus during the period of pre-emergence of shoots did not show any appreciable change, in the non-shoot forming callus, it increased during the corresponding period. The developmental patterns of nitrate reductase in both the programmes were identical except for the fact that in the shoot forming tissue the nitrate reductase activity was higher at all times than in the non-shoot forming callus. The data suggest that (a) shoot differentiation occurs concomitant with peak glutamine synthetase, glutamate synthase, and nitrate reductase activity, whereas the glutamate dehydrogenase activity is at its lowest, (b) better mobilization of nitrate occurs in the shoot forming callus and (c) the glutamine synthetase/glutamate synthase pathway becomes operative prior to shoot differentiation., (Copyright © 1984 Gustav Fischer Verlag, Stuttgart. Published by Elsevier GmbH.. All rights reserved.)

The in vivo nitrate reductase activity in 8 day old dark-grown sugarcane callus was over three fold that of the light-grown callus. NADH (0.3 mM) in the reaction system, increased the in vivo nitrate reductase activity by more than two fold both in the dark- and the light-grown callus tissues. The NADH dependence of nitrate reductase activity followed Michaelian kinetics. The apparent Km values for NADH were 0.083 mM and 0.20 mM, respectively, for the dark- and the light-grown callus. In vivo nitrate reductase activity in green sugarcane leaves (field grown) was unaffected by NADH in the reaction system. Under the standard conditions of assay up to 60% of the NADH penetrated into the sugarcane callus within 2 min. No penetration of NADH into the sugarcane leaf discs was, however, recorded under identical conditions.