Preliminary results of hypofractionated radiotherapy in breast cancer in Chandigarh, India: single-centre, non-inferiority, open-label, randomised, phase 3 trial.

Background: Globally, most of the randomised trials with hypofractionation in patients with breast cancer have used 3-dimensional conformal radiotherapy technique (3D-CRT). As facilities for 3D-CRT technique may not be available in low-resource settings, there is a need to see if hypofractionation is feasible and safe with 2-dimensional (2-D) technique. In this study, we compared a 3-week radiation schedule with a 2-week schedule of hypofractionated radiotherapy in patients with breast cancer with 2-D technique. Methods: The current study was an open-label, randomised, phase 3 trial. Patients with breast cancer, stage I-III, post mastectomy or after breast conservative surgery who needed adjuvant locoregional radiotherapy were randomised in the Department of Radiotherapy & Oncology, Post Graduate Institute of Medical Education & Research (PGIMER), Chandigarh, India; to 34Gy in 10 fractions over 2 weeks (2-week arm) or 35Gy in 15 fractions over 3 weeks to the chest wall and 40Gy/15#/3wks to breast and supraclavicular fossa (3-week arm). Boost dose when indicated was 8-10Gy/2-4#/2-4 days in both the arms. Patients were planned on a 2-dimensional (2D) simulator with 2 tangential fields to breast/chest wall and incident supraclavicular fossa field. Acute toxicity was assessed using the Radiation Therapy Oncology Group (RTOG) grading scale. Assessments were carried out weekly during radiotherapy and at 4 weeks after treatment by the physician. Cosmetic outcome was assessed using the Harvard/National Surgical Adjuvant Breast and Bowel Project (NSABP)/RTOG scale. The toxicity rates between the two arms were compared using Fisher's exact tests. The trial was approved by institutional ethics committee and registered with ClinicalTrials.gov, number NCT04075058. Findings: This study included 1121 eligible patients from June 2015 to December 2020. Median follow-up was 35 months (6-84 months). Mean age was 48 years (24-75 years). The patient characteristics were comparable between the two arms except for more mastectomies in the 3-week arm and more node-positive patients in the 2-week arm. There were more oestrogen receptor-positive tumors in the 3-week arm. Acute skin toxicities were comparable between the two arms. Grade 2 and 3 skin toxicity was 100 (18%) and 82 (15%); and 16 (3%) and 12 (2%) in the 3-week and 2-week arm (p = 0.21), respectively. Cosmetic outcome was assessed as Excellent or Good for 89% of patients in the 3-week arm as compared to 94% in the 2-week arm (p = 0.004). Interpretation: The two radiation schedules were comparable in terms of acute skin toxicity. The cosmetic outcome was better with the 2-week schedule. The preliminary findings indicate 2-week radiotherapy schedule with 2-D technique was better than the 3-week schedule in patients with breast cancer. However, disease outcomes and late-term toxicities need to be further checked. Funding: This study was funded by Science and Engineering Research Board (SERB), India.

Advanced and Metastatic Gastrointestinal Stromal Tumors Presenting With Surgical Emergencies Managed With Surgical Resection: A Case Series.

Advanced and metastatic gastrointestinal stromal tumors (GISTs) presenting with surgical emergencies are rare. Neoadjuvant imatinib being the treatment of choice for non-metastatic advanced disease with a proven role in downstaging the disease may not be feasible in patients presenting with bleeding and obstruction. We present a case series with retrospective analysis of a prospectively maintained database of patients with advanced and metastatic GISTs presenting with surgical emergencies. Clinical characteristics, imaging and endoscopic findings, surgical procedures, histological findings, and outcomes in these patients were studied. Four patients were included in this case series, with three males and one female (age range: 24-60 years). Two patients presented with melena; one with hemodynamic instability despite multiple blood transfusions underwent urgent exploratory laparotomy for bleeding gastric GIST, while the other underwent surgical exploration after careful evaluation given the recurrent, metastatic disease with a stable metabolic response on six months of imatinib. One patient with metastatic jejunal GIST who presented with an umbilical nodule and intestinal obstruction was given a trial of non-operative management for 72 hours, but due to non-resolution of obstruction, segmental jejunal en bloc resection with the dome of the urinary bladder with reconstruction and metastasectomy was needed. The patient with advanced gastric GIST who presented with gastric outlet obstruction was resuscitated, and an attempt of endoscopic naso-jejunal tube placement was tried, which failed, and exploration was needed. The mean length of hospital stay was 7.5 days. Histopathological examination confirmed GIST in all four patients with microscopic negative resection margins. All patients were started on imatinib with dose escalation to 800 mg in the patient with recurrent and metastatic disease; however, the patient with bleeding gastric GIST experienced severe adverse effects of imatinib and discontinued the drug shortly. All four patients are disease-free on follow-ups of 15 months, 48 months for the patient with advanced non-metastatic disease, and six and 24 months for the patients with metastatic disease. In the era of tyrosine kinase inhibitor (TKI) therapy for advanced and metastatic disease, upfront surgery is usually reserved for surgical emergencies only. Surgical resection, the cornerstone for the treatment of resectable GIST, may also be clinically relevant in metastatic settings, although it requires a careful and individualized approach.

Neuroendocrine tumour of the lesser omentum.

A man in his early 20s presented to us in the outpatient department with a history of diarrhoea for 4 months. Investigations revealed elevated serum chromogranin levels and an intensely avid lesion in the gastrohepatic ligament in Gallium DOTATATE positron emission tomography (PET). The tumour was excised laparoscopically, and no other lesions were seen. The patient improved clinically and had a normal serum chromogranin level postoperatively. He is currently much improved at the 1year follow-up. We did an extensive workup to look for a primary tumour. It was concluded that it was a de novo tumour arising from the lesser sac. The recommended investigations in case of neuroendocrine tumour (NET) with unknown primary include blood investigations to look for the functional status of the tumour, histopathological examination, including immunohistochemistry, and radiological imaging, which must include a Gallium DOTATATE PET. An isolated NET of the lesser sac has not been reported in the literature.

Small bowel obstruction: a rare presentation of systemic mastocytosis.

Mastocytosis (MC) is a heterogeneous group of disorders characterised by abnormal growth, neoplastic proliferation and accumulation of mast cells. Approximately, 80% of patients with MC have evidence of skin involvement, while the rest may involve the gastrointestinal tract, liver, spleen or bone. Isolated gastrointestinal presentation of aggressive MC without bone marrow involvement and cutaneous symptoms is rare. Isolated MC with tumour cells infiltrating the ileum presenting with mechanical obstruction has not been reported in the literature to date. Here, we present a case of a patient in his late 50s who presented with malignant MC of the small bowel with obstruction in the emergency surgical outpatient department. The patient underwent surgical resection of the affected bowel. Histopathological examination along with immunohistochemistry revealed malignant MC. The further evaluation consisted of bone marrow biopsies, PET CT and other mutation analyses. However, the patient succumbed to death during the further course of treatment. The differential of MC must be kept in mind when there is the presence of abnormal hematopoietic cells in gastrointestinal biopsies even in the absence of cutaneous manifestations and bone marrow abnormalities.

Effect of intraperitoneal magnesium sulfate in the prevention of postoperative pain in daycare laparoscopic cholecystectomy - A prospective randomized controlled trial.

This study evaluated the efficacy of intraperitoneal magnesium sulfate (MgSO4) in preventing postoperative pain after elective laparoscopic cholecystectomy (LC). It is a prospective, double-blinded, placebo-controlled, randomized trial which included 64 patients who underwent LC. Patients were equally randomized into Groups A and B. MgSO4 and normal saline were instilled in subdiaphragmatic space in Groups A and B, respectively, after creating pneumoperitoneum and before starting dissection. The Visual analogue Scale (VAS) was used to determine postoperative pain. Patients who received intraperitoneal MgSO4 had lower average VAS scores for the first 6 h postoperatively, and also, the time for the requirement of first analgesic was longer (3.6 ± 0.4 vs. 2.3 ± 1.0 h). The incidence of vomiting and the requirement for rescue antiemetic was also lower in Group A. Intraperitoneal instillation of MgSO4 reduces postoperative pain and vomiting following elective LC without incurring additional side effects.

Visceral Obesity as a Predictor of Postoperative Complications After Pancreaticoduodenectomy.

INTRODUCTION: Obesity is associated with increased morbidity and mortality post surgery. The measurement of visceral obesity can predict postoperative outcomes after pancreaticoduodenectomy. METHODS: This is a prospective observational study. Visceral obesity was calculated by measuring the fat thickness in the retro-renal area by using a computed tomography scan. Visceral obesity was defined as retro-renal fat thickness (RRFT) of ≥ 2 cm. Patients were divided into two groups: Group-A (RRFT < 2 cm, non-obese) and Group-B (RRFT > 2 cm, obese). Demographic, clinical, and intraoperative variables were correlated with postoperative outcomes. RESULTS: Fifty-six patients were included in the study. Thirty-two patients were included in Group-A, and 24 patients were included in Group-B. The two groups had comparable outcomes. A total of 21 patients in Group-A (65.62%) and 17 patients in Group-B (70.83%) had comorbidities, including diabetes mellitus, hypertension, and coronary disease (p=0.680). American Society of Anesthesiologists (ASA) grading was comparable (p=0.927). BMI was also comparable (p=0.354). Type of pancreaticoduodenectomy, pancreatic texture, pancreatic duct diameter, and technique of pancreaticojejunostomy anastomosis were comparable. The mean operative time was longer in Group-B (362 ± 36.2 min vs. 298 ± 45.2 min) (p=0.001). Intraoperative blood loss was more in Group-B (312 ± 36.8 ml vs. 267 ± 23.7 ml) (p=0.001). The rates of postoperative pancreatic fistula and delayed gastric emptying were comparable (p=0.402 and p=0.134, respectively). The length of hospital stay was longer in patients in Group-B (p=0.004). There was one death in Group-B (obese group). CONCLUSION: Visceral obesity is a risk factor for postoperative complications after a pancreaticoduodenectomy.

Diagnostic and Therapeutic Challenges in the Management of Acute Massive Overt Bleeding of Jejunal Gastrointestinal Stromal Tumours: Case Series.

INTRODUCTION: Jejunal gastrointestinal stromal tumours (GIST) are rare mesenchymal tumours. Acute massive overt bleeding from jejunal GIST is very rare and poses both diagnostic and therapeutic challenges in emergent conditions. METHODS: A case series with retrospective analysis of prospectively maintained database of patients presenting with acute massive overt bleeding secondary to histologically proven jejunal GIST was done. Clinical characteristics, endoscopic and imaging diagnostic features, histological findings, surgical procedures and outcomes in these patients were studied. RESULTS: Three patients were included in this case series. Mean age of presentation was 49.0 years with two male and one female patient. All three patients presented with melena and hemodynamic instability, resuscitated with adequate blood transfusions. Routine endoscopic assessment were inconclusive. Multiphasic Computed Tomographic Angiography (CTA) revealed hypodense hypervascular mass in jejunum in all three patients. One patient was unresponsive to blood transfusion and underwent emergency exploratory laparotomy. One patient underwent laparoscopic resection and reconstruction. Mean length of hospital stay was 5.3 days. Histopathological examination confirmed jejunal GIST in all three patients with microscopically negative resection margins. Two patients were disease free till 18-month follow-up and the one patient lost to follow-up after 1 year. CONCLUSION: Multiphasic CTA is a single-step diagnostic tool for localisation of bleed and assessment of tumour characteristics in emergent conditions. Surgical resection is the mainstay of treatment for both control of bleed and to provide oncologically clear resection margins.

Successful management of impaled ice-pick stab injury of the suprarenal aorta - lessons learnt.

Penetrating injury of the aorta is rare and lethal. The reported mortality rate is very high. Few patients survive and present to the hospital. Some of these injuries are salvageable if treated in a timely and aggressive manner. Here we present a case of penetrating injury of the aorta with impalement of the ice-pick, which was successfully managed with laparotomy and primary repair of the aortic injury after adequate resuscitation.

Correlation of Multi-Detector Computed Tomography and Intraoperative Variations of the Celiac Trunk and Hepatic Artery in Resectable Hepatobiliary Pancreatic Cancers.

Introduction Knowledge of celiac artery variations is imperative to perform complex hepato-biliary pancreatic surgical procedures to avoid inadvertent complications. Multi-detector computed tomographic (MDCT) angiography aids in detecting these variations preoperatively. Surgical confirmation is considered the gold standard. Aims and objectives Preoperative assessment of celiac artery variations by MDCT angiography and surgical confirmation intraoperatively in resectable hepato-biliary pancreatic cancers. Patients and methods MDCT angiography was performed in 40 patients with clinical evidence of resectable hepato-biliary-pancreatic cancers. Three dimensional (3D) reconstructions were performed to confirm the celiac artery variations. Surgery was performed as per the institute's protocol in all these patients for resection of tumor and confirmation of celiac artery anatomy. Variations were confirmed surgically that were identified through imaging. Results MDCT angiography identified normal trifurcated celiac artery anatomy in 33 (82.5%) patients and variant anatomy in seven (17.5%) patients. The most common variation was a replaced right hepatic artery (r-RHA) from the superior mesenteric artery (SMA) in four (10%) of patients. A replaced left hepatic artery (r-LHA) from the celiac trunk, a common hepatic artery (CHA) from the abdominal aorta, and an accessory right hepatic artery (ac-RHA) from the proper hepatic artery itself were identified in one (2.5%) patient each, respectively. All these findings were confirmed intraoperatively. There was a 100% statistical correlation between imaging and surgical findings. Conclusion Surgical confirmation of radiological data of celiac artery variations is the gold standard to avoid disastrous complications such as inadvertent vascular bleeds, biliary injuries, and hepatic necrosis. Since the presence of variations warrants the preservation or excision of the arterial system without oncological compromise and minimizing surgical complications.

Plant Ribosomal Proteins, RPL12 and RPL19, Play a Role in Nonhost Disease Resistance against Bacterial Pathogens.

Characterizing the molecular mechanism involved in nonhost disease resistance is important to understand the adaptations of plant-pathogen interactions. In this study, virus-induced gene silencing (VIGS)-based forward genetics screen was utilized to identify genes involved in nonhost resistance in Nicotiana benthamiana. Genes encoding ribosomal proteins, RPL12 and RPL19, were identified in the screening. These genes when silenced in N. benthamiana caused a delay in nonhost bacteria induced hypersensitive response (HR) with concurrent increase in nonhost bacterial multiplication. Arabidopsis mutants of AtRPL12 and AtRPL19 also compromised nonhost resistance. The studies on NbRPL12 and NbRPL19 double silenced plants suggested that both RPL12 and RPL19 act in the same pathway to confer nonhost resistance. Our work suggests a role for RPL12 and RPL19 in nonhost disease resistance in N. benthamiana and Arabidopsis. In addition, we show that these genes also play a minor role in basal resistance against virulent pathogens.

Comparative proteomics of yeast-elicited Medicago truncatula cell suspensions reveals induction of isoflavonoid biosynthesis and cell wall modifications.

The temporal proteome response of Medicago truncatula suspension cell cultures to yeast elicitation (which mimics fungal infection) was investigated using two-dimensional polyacrylamide gel electrophoresis (2-DE) and nanoliquid chromatography coupled to tandem mass spectrometry (nano LC-MS/MS). Reproducibility of 2-DE was assessed using the number of the visualized protein spots and spot volume. Average coefficient of variation was determined to be less than 6% for the number of spots and around 50% for spot volume. About 4% of the total visualized proteins, that is, 34 out of 861, were differentially accumulated in the suspension cells 24 h after yeast elicitation, including isoflavononid biosynthetic enzymes and a putative laccase. The induction of the putative laccase was highly correlated with the polymerization of phenolics such as 4-hydroxybenzoic acid, 4-hydroxybenzaldehyde, and ferulic acid into cell walls. In contrast, lignin was only induced at the later stages of the temporal study, indicating that this specific laccase is primarily involved in cell wall modifications and/or fortifications rather than in lignification in response to yeast elicitation.

A role for chloroplast-localized Thylakoid Formation 1 (THF1) in bacterial speck disease development.

Coronatine (COR), a jasmonate mimic produced by Pseudomonas syringae pv. tomato DC3000 (Pst DC3000) is required for full virulence of Pst DC3000 in tomato and Arabidopsis. COR is shown to induce a range of physiological processes including chlorosis, root growth inhibition and anthocyanin accumulation in tomato. To elucidate the host/signaling genes involved in COR-responses, we utilized a forward genetics approach using Nicotiana benthamiana and virus-induced gene silencing (VIGS) and identified genes that play a role in COR-mediated chlorosis. We designated these genes as altered COR response (ALC). When silenced, one gene designated ALC1 produced a hypersensitive/necrosis-like phenotype after COR application in a coronatine insensitive 1 (COI1)-dependent manner. In pathogenicity assays performed on Arabidopsis thylakoid formation 1 (thf1) knockout lines and SlALC1-silenced tomato plants, Pst DC3000 induced coalescing necrotic lesions in an accelerated manner. Furthermore, we showed that COR affects ALC1 localization in chloroplast in a COI1-dependent manner. In conclusion, our results show the potential of VIGS-based, forward genetic screens to identify new players in COR-mediated signal transduction.

A virus-induced gene silencing screen identifies a role for Thylakoid Formation1 in Pseudomonas syringae pv tomato symptom development in tomato and Arabidopsis.

Pseudomonas syringae pv tomato DC3000 (Pst DC3000), which causes disease in tomato (Solanum lycopersicum) and Arabidopsis (Arabidopsis thaliana), produces coronatine (COR), a non-host-specific phytotoxin. COR, which functions as a jasmonate mimic, is required for full virulence of Pst DC3000 and for the induction of chlorosis in host plants. Previous genetic screens based on insensitivity to COR and/or methyl jasmonate identified several potential targets for COR and methyl jasmonate. In this study, we utilized Nicotiana benthamiana and virus-induced gene silencing to individually reduce the expression of over 4,000 genes. The silenced lines of N. benthamiana were then screened for altered responses to purified COR. Using this forward genetics approach, several genes were identified with altered responses to COR. These were designated as ALC (for altered COR response) genes. When silenced, one of the identified genes, ALC1, produced a hypersensitive/necrosis-like phenotype upon COR application in a Coronatine-Insensitive1 (COI1)-dependent manner. To understand the involvement of ALC1 during the Pst DC3000-host interaction, we used the nucleotide sequence of ALC1 and identified its ortholog in Arabidopsis (Thylakoid Formation1 [THF1]) and tomato (SlALC1). In pathogenicity assays performed on Arabidopsis thf1 mutant and SlALC1-silenced tomato plants, Pst DC3000 induced accelerated coalescing necrotic lesions. Furthermore, we showed that COR affects ALC1 localization in chloroplasts in a COI1-dependent manner. In conclusion, our results show that the virus-induced gene silencing-based forward genetic screen has the potential to identify new players in COR signaling and disease-associated necrotic cell death.

Mutations in Arabidopsis fatty acid amide hydrolase reveal that catalytic activity influences growth but not sensitivity to abscisic acid or pathogens.

Fatty acid amide hydrolase (FAAH) terminates the endocannabinoid signaling pathway that regulates numerous neurobehavioral processes in animals by hydrolyzing N-acylethanolamines (NAEs). Recently, an Arabidopsis FAAH homologue (AtFAAH) was identified, and several studies, especially those using AtFAAH overexpressing and knock-out lines, have suggested an in vivo role for FAAH in the catabolism of NAEs in plants. We previously reported that overexpression of AtFAAH in Arabidopsis resulted in accelerated seedling growth, and in seedlings that were insensitive to exogenous NAEs but hypersensitive to abscisic acid (ABA) and hypersusceptible to nonhost pathogens. Here we show that whereas the enhanced growth and NAE tolerance of the AtFAAH overexpressing seedlings depend on the catalytic activity of AtFAAH, hypersensitivity to ABA and hypersusceptibility to nonhost pathogens are independent of its enzymatic activity. Five amino acids known to be critical for rat FAAH activity are also conserved in AtFAAH (Lys-205, Ser-281, Ser-282, Ser-305, and Arg-307). Site-directed mutation of each of these conserved residues in AtFAAH abolished its hydrolytic activity when expressed in Escherichia coli, supporting a common catalytic mechanism in animal and plant FAAH enzymes. Overexpression of these inactive AtFAAH mutants in Arabidopsis showed no growth enhancement and no NAE tolerance, but still rendered the seedlings hypersensitive to ABA and hypersusceptible to nonhost pathogens to a degree similar to the overexpression of the native AtFAAH. Taken together, our findings suggest that the AtFAAH influences plant growth and interacts with ABA signaling and plant defense through distinctly different mechanisms.

Role of nucleotide binding in septin-septin interactions and septin localization in Saccharomyces cerevisiae.

Septins are a conserved family of eukaryotic GTP-binding, filament-forming proteins. In Saccharomyces cerevisiae, five septins (Cdc3p, Cdc10p, Cdc11p, Cdc12p, and Shs1p) form a complex and colocalize to the incipient bud site and as a collar of filaments at the neck of budded cells. Septins serve as a scaffold to localize septin-associated proteins involved in diverse processes and as a barrier to diffusion of membrane-associated proteins. Little is known about the role of nucleotide binding in septin function. Here, we show that Cdc3p, Cdc10p, Cdc11p, and Cdc12p all bind GTP and that P-loop and G4 motif mutations affect nucleotide binding and result in temperature-sensitive defects in septin localization and function. Two-hybrid, in vitro, and in vivo analyses show that for all four septins nucleotide binding is important in septin-septin interactions and complex formation. In the absence of complete complexes, septins do not localize to the cortex, suggesting septin localization factors interact only with complete complexes. When both complete and partial complexes are present, septins localize to the cortex but do not form a collar, perhaps because of an inability to form filaments. We find no evidence that nucleotide binding is specifically involved in the interaction of septins with septin-associated proteins.

Role of a Cdc42p effector pathway in recruitment of the yeast septins to the presumptive bud site.

The septins are GTP-binding, filament-forming proteins that are involved in cytokinesis and other processes. In the yeast Saccharomyces cerevisiae, the septins are recruited to the presumptive bud site at the cell cortex, where they form a ring through which the bud emerges. We report here that in wild-type cells, the septins typically become detectable in the vicinity of the bud site several minutes before ring formation, but the ring itself is the first distinct structure that forms. Septin recruitment depends on activated Cdc42p but not on the normal pathway for bud-site selection. Recruitment occurs in the absence of F-actin, but ring formation is delayed. Mutant phenotypes and suppression data suggest that the Cdc42p effectors Gic1p and Gic2p, previously implicated in polarization of the actin cytoskeleton, also function in septin recruitment. Two-hybrid, in vitro protein binding, and coimmunoprecipitation data indicate that this role involves a direct interaction of the Gic proteins with the septin Cdc12p.