ESMO recommendations on microsatellite instability testing for immunotherapy in cancer, and its relationship with PD-1/PD-L1 expression and tumour mutational burden: a systematic review-based approach.

BACKGROUND: Cancers with a defective DNA mismatch repair (dMMR) system contain thousands of mutations most frequently located in monomorphic microsatellites and are thereby defined as having microsatellite instability (MSI). Therefore, MSI is a marker of dMMR. MSI/dMMR can be identified using immunohistochemistry to detect loss of MMR proteins and/or molecular tests to show microsatellite alterations. Together with tumour mutational burden (TMB) and PD-1/PD-L1 expression, it plays a role as a predictive biomarker for immunotherapy. METHODS: To define best practices to implement the detection of dMMR tumours in clinical practice, the ESMO Translational Research and Precision Medicine Working Group launched a collaborative project, based on a systematic review-approach, to generate consensus recommendations on the: (i) definitions related to the concept of MSI/dMMR; (ii) methods of MSI/dMMR testing and (iii) relationships between MSI, TMB and PD-1/PD-L1 expression. RESULTS: The MSI-related definitions, for which a consensus frame-work was used to establish definitions, included: 'microsatellites', 'MSI', 'DNA mismatch repair' and 'features of MSI tumour'. This consensus also provides recommendations on MSI testing; immunohistochemistry for the mismatch repair proteins MLH1, MSH2, MSH6 and PMS2 represents the first action to assess MSI/dMMR (consensus with strong agreement); the second method of MSI/dMMR testing is represented by polymerase chain reaction (PCR)-based assessment of microsatellite alterations using five microsatellite markers including at least BAT-25 and BAT-26 (strong agreement). Next-generation sequencing, coupling MSI and TMB analysis, may represent a decisive tool for selecting patients for immunotherapy, for common or rare cancers not belonging to the spectrum of Lynch syndrome (very strong agreement). The relationships between MSI, TMB and PD-1/PD-L1 expression are complex, and differ according to tumour types. CONCLUSIONS: This ESMO initiative is a response to the urgent questions raised by the growing success of immunotherapy and provides also important insights on the relationships between MSI, TMB and PD-1/PD-L1.

Identification and validation of novel microenvironment-based immune molecular subgroups of head and neck squamous cell carcinoma: implications for immunotherapy.

Background: Targeting the immune checkpoint pathway has demonstrated antitumor cytotoxicity in treatment-refractory head and neck squamous cell carcinoma (HNSC). To understand the molecular mechanisms underpinning its antitumor response, we characterized the immune landscape of HNSC by their tumor and stromal compartments to identify novel immune molecular subgroups. Patients and methods: A training cohort of 522 HNSC samples from the Cancer Genome Atlas profiled by RNA sequencing was analyzed. We separated gene expression patterns from tumor, stromal, and immune cell gene using a non-negative matrix factorization algorithm. We correlated the expression patterns with a set of immune-related gene signatures, potential immune biomarkers, and clinicopathological features. Six independent datasets containing 838 HNSC samples were used for validation. Results: Approximately 40% of HNSCs in the cohort (211/522) were identified to show enriched inflammatory response, enhanced cytolytic activity, and active interferon-γ signaling (all, P < 0.001). We named this new molecular class of tumors the Immune Class. Then we found it contained two distinct microenvironment-based subtypes, characterized by markers of active or exhausted immune response. The Exhausted Immune Class was characterized by enrichment of activated stroma and anti-inflammatory M2 macrophage signatures, WNT/transforming growth factor-ß signaling pathway activation and poor survival (all, P < 0.05). An enriched proinflammatory M1 macrophage signature, enhanced cytolytic activity, abundant tumor-infiltrating lymphocytes, high human papillomavirus (HPV) infection, and favorable prognosis were associated with Active Immune Class (all, P < 0.05). The robustness of these immune molecular subgroups was verified in the validation cohorts, and Active Immune Class showed potential response to programmed cell death-1 blockade (P = 0.01). Conclusions: This study revealed a novel Immune Class in HNSC; two subclasses characterized by active or exhausted immune responses were also identified. These findings provide new insights into tailoring immunotherapeutic strategies for different HNSC subgroups.

Acetylcholinesterase inhibitory potential of scorpion venom in Aedes aegypti (Diptera: Culicidae).

Scorpion venom contains a variety of neurotoxins which interact with ion channels and affect their activities. The present study was designed to evaluate the potential of scorpion venom as acetylcholinesterase (AChE) inhibitor by using Aedes aegypti as model organism. Venoms of two species, Hottentota tamulus (Fabricus, 1798) and Androctonus finitimus (Pocock, 1897) were selected for this study. Two peptides (36 kDa from H. tamulus and 54 kDa from A. finitimus) were separated from scorpion venom by using HPLC. Selected peptides caused significantly higher mortality in larvae and adults of Aedes aegypti than control (no mortalities were observed in control groups). Significant acetylcholinesterase (AChE) inhibitory potential of both peptides was recorded by spectrophotometer. The peptide of A. finitimus caused significantly higher mortality (95±1.53% in larvae and 100% in adults) than the peptide of H. tamulus (84.33±2.33% in larvae and 95.37±1.45% in adults). While H. tamulus peptide was more efficient in reducing AChE activity (0.029±0.012 in larvae and 0.03±0.003 in adults) than the peptide of A. finitimus (0.049±0.005 in larvae and 0.047±0.001 in adults). It was concluded that H. tamulus venom peptide was more efficiently reducing AChE activity, thus it could be a potential bio-insecticide which can be synthesized at industrial scale for the control of harmful insects.

Healing of a large avulsed mandibular bony segment: report of a unique case.

This case report puts an emphasis on retaining and re-fixing any avulsed bony segments in the maxillofacial region and maintaining the periosteal layer whenever possible, especially in young patients. Adequate bony fixation and watertight soft tissue closure are vital components for bone healing. The healing potential of facial bones is much higher as compared to the long bones, due to the superior blood supply.

Use of MV and kV imager correlation for maintaining continuous real-time 3D internal marker tracking during beam interruptions.

The integration of onboard kV imaging together with a MV electronic portal imaging device (EPID) on linear accelerators (LINAC) can provide an easy to implement real-time 3D organ position monitoring solution for treatment delivery. Currently, real-time MV-kV tracking has only been demonstrated by simultaneous imagining by both MV and kV imaging devices. However, modalities such as step-and-shoot IMRT (SS-IMRT), which inherently contain MV beam interruptions, can lead to loss of target information necessary for 3D localization. Additionally, continuous kV imaging throughout the treatment delivery can lead to high levels of imaging dose to the patient. This work demonstrates for the first time how full 3D target tracking can be maintained even in the presence of such beam interruption, or MV/kV beam interleave, by use of a relatively simple correlation model together with MV-kV tracking. A moving correlation model was constructed using both present and prior positions of the marker in the available MV or kV image to compute the position of the marker on the interrupted imager. A commercially available radiotherapy system, equipped with both MV and kV imaging devices, was used to deliver typical SS-IMRT lung treatment plans to a 4D phantom containing internally embedded metallic markers. To simulate actual lung tumor motion, previous recorded 4D lung patient motion data were used. Lung tumor motion data of five separate patients were inputted into the 4D phantom, and typical SS-IMRT lung plans were delivered to simulate actual clinical deliveries. Application of the correlation model to SS-IMRT lung treatment deliveries was found to be an effective solution for maintaining continuous 3D tracking during 'step' beam interruptions. For deliveries involving five or more gantry angles with 50 or more fields per plan, the positional errors were found to have < or =1 mm root mean squared error (RMSE) in all three spatial directions. In addition to increasing the robustness of MV-kV tracking against beam interruption, it was also found that use of correlation can be an effective way of lowering kV dose to the patient and for increasing kV image quality by reduction of MV scatter interference.

Lipoxygenase inhibitory tetraketones: potential remedial source for inflammation and asthma.

OBJECTIVE: A series of tetraketones has been synthesized by way of a one pot synthesis and screened for inhibitory activity against the enzyme lipoxygenase. METHOD: An efficient and high yielding one pot synthesis to tetraketones [2-22] has been developed by way of tetraethyl ammonium bromide (Et4N+Br-) mediated condensation of cyclohexane-1, 3-dione [1] with a variety of aldehydes. Lipoxygenase enzyme solution was prepared so that enzyme concentration in reaction mixture was adjusted to give rates of 0.05 absorbance/minute. The test compounds were prepared in methanol of concentrations 50, 25, 12.5, 6.25 and 3.125 microM. The reaction mixture contained 160 microL (100 mM) sodium phosphate buffer (pH 8.0), 10 microL of test-compound solution and 20 microL of lipoxygenase solution. The contents were mixed and incubated for 10 minutes at 25 degrees C. The reaction was then initiated by the addition of 10 microL substrate solution (linoleic acid, 0.5 mM, 0.12% w/v tween 20 in the ratio of 1:2), with the formation of (9Z, 11E)-(13S)-13-hydroperoxyoctadeca-9,11-dienoate, the change of absorbance at 234 nm was followed for 6 minutes. The concentrations of the test compounds that inhibited the lipoxygenase activity by 50% (IC50) were determined by monitoring the effect of increasing concentrations of these compounds in the assays on the degree of inhibition. The IC50 values were calculated by means of the EZ-Fit Enzyme-Kinetics Program (Perrella Scientific Inc., Amherst, U.S.A.). RESULT: The tetraketones [2-22] were synthesized in high yields (91-98%) using mild reaction conditions. Most of these compounds showed significant inhibitory activity against the enzyme lipoxygenase. It was found that the presence of substituents which increase delocalization of electrons enhances the inhibitory activity. CONCLUSION: It is concluded that the study is likely to lead to the discovery of therapeutically efficient agents against important disorders such as inflammation and asthma.

Patterns of facial fractures in children.

Morbidity and mortality among children is usually the result of trauma. Because a child's face is retruded relative to the protecting skull, has a thicker layer of adipose tissue, more elastic bones, flexible sutures lines, the presence of tooth buds within the jaws, and the lack of pneumatisation of the sinuses, the facial bones fracture less commonly than in adults. Our aim was to assess the patterns of such fractures in children who presented to the department of Oral and Maxillofacial Surgery, King Edward Medical University/Mayo Hospital Lahore, Pakistan. All 535 eligible children between the ages of 1-16 years who presented during the two years December 2009 - December 2011 were included in the study. Facial fractures were diagnosed by clinical examination, plain radiographs, and computed tomography, and the pattern of fractures of the facial bones including the frontal bone, orbital bones, maxilla, zygoma, naso-orbito-ethmoidal complex, mandible, and dentoalveolar region was documented. The male:female ratio was 2:1 with 369 male (70%) and 166 female (31%) patients. Fall was the cause in 212 (39%), and in 167 (31%) it was road traffic accidents, while sports were the cause in 135 (25%). The naso-orboto-ethmoid complex was fractured in 37 cases (7%) while 104 children (19%) presented with isolated fractures of the zygomatic bone. The maxilla was fractured in 195 cases (36%), the mandible in 380 (71%), and dentoalveolar trauma was the cause in 256 (50%). The mandible was the bone that was most often fractured (mostly in boys and usually as a result of falls during summer vacations), with the peak occurring in those aged 8-12 years.

Poverty-associated emerging infection of Cystic Echinococcosis in population of Northern Pakistan: A hospital based study.

Cystic Echinococcosis (CE) is one of the most important zoonotic parasitic diseases in human, livestock, and wildlife globally. The prevalence of CE depends upon human behavioral risk factors, the diversity and ecology of animal host interactions and the genetic diversity within Echinococcus species which differ in their zoonotic potential and pathogenicity. It is a neglected, economic and socio-cultural problem in Pakistan. The available data about the incidence of CE is very limited and no extensive study has been reported in Pakistan. The current study was aimed to analyze the hospital reported cases of CE and the associated risk factors related to the incidence of CE. The hospital-based data of CE for the time period of January 2012-December 2017 was collected from Islamabad, Rawalpindi and Peshawar. The data covered demographic characteristics including age, gender, and cyst localization of infected individuals and socioeconomic determinants. The data was analyzed based upon different risk factors along with the different socioeconomic parameters that has an important impact on the distribution of disease. A total of 228 cases were presented in the selected hospitals of different cities during the study period. Out of total 228 patients, 59.21% were males and 40.78% were females (P<0.001). Most infections have been recorded in young adults (>20-30) showing 22.8% of total infected individuals followed by children (0-10) showing 10.5% (n=24), respectively (P<0.001). Liver was the most vulnerable organ (58.77%, n=134) followed by lungs (14.47%, n=33) (P<0.001). The infection was higher among rural communities (84.2%) than urban (12.8%) (P<0.001). Socioeconomic and demographic factors had an important impact on the intensity of disease (P<0.001). The occurrence of cases in children and young adults was an important finding as it indicated an active transmission of the parasite in Pakistan along with the poverty index. Emergence of echinococcosis in Pakistan showed that emerging health issues in Pakistan could bring the disease to limelight for future research. This finding, together with the fact that 1 hospital reported 214 cases over 6 years 325 underlines the need for a program for prevention/control of this disease in Pakistan. The timely measure needs to be taken to hamper the disease development and establishment. In order to control the disease, complete surveillance should be done which in turn weighs down the disease progress.

Detection of Cancer DNA in Early Stage and Metastatic Breast Cancer Patients.

Breast cancer is the leading cause of cancer in women and the second leading cause of cancer-related death. There are many subtypes of breast cancer, which can be identified through the process of molecular and genetic profiling. While the current standard of care utilizes tumor tissue biopsy to subclassify breast cancer, plasma tumor DNA (ptDNA) can be detected through droplet digital PCR (ddPCR) of plasma obtained from a simple blood draw. Tissue biopsy is not only more invasive but because tumors exhibit heterogeneity it can be less accurate. Blood collects DNA shed from normal and cancerous cells alike, thus ddPCR of plasma offers a broader picture of a cancer's genetic makeup. This chapter summarizes how patients with breast cancer can be screened for specific cancerous mutations in both tissue and plasma through the use of ddPCR.

Compromised BRCA1-PALB2 interaction is associated with breast cancer risk.

The major breast cancer suppressor proteins BRCA1 and BRCA2 play essential roles in homologous recombination (HR)-mediated DNA repair, which is thought to be critical for tumor suppression. The two BRCA proteins are linked by a third tumor suppressor, PALB2, in the HR pathway. While truncating mutations in these genes are generally pathogenic, interpretation of missense variants remains a challenge. To date, patient-derived missense variants that disrupt PALB2 binding have been identified in BRCA1 and BRCA2; however, there has not been sufficient evidence to prove their pathogenicity in humans, and no variants in PALB2 that disrupt either its BRCA1 or BRCA2 binding have been reported. Here we report on the identification of a novel PALB2 variant, c.104T>C (p.L35P), that segregates in a family with a strong history of breast cancer. Functional analyses showed that L35P abrogates the PALB2-BRCA1 interaction and completely disables its abilities to promote HR and confer resistance to platinum salts and PARP inhibitors. Whole-exome sequencing of a breast cancer from a c.104T>C carrier revealed a second, somatic, truncating mutation affecting PALB2, and the tumor displays hallmark genomic features of tumors with BRCA mutations and HR defects, cementing the pathogenicity of L35P. Parallel analyses of other germline variants in the PALB2 N-terminal BRCA1-binding domain identified multiple variants that affect HR function to varying degrees, suggesting their possible contribution to cancer development. Our findings establish L35P as the first pathogenic missense mutation in PALB2 and directly demonstrate the requirement of the PALB2-BRCA1 interaction for breast cancer suppression.

Biological Features of Human Papillomavirus-related Head and Neck Cancers Contributing to Improved Response.

Head and neck squamous cell carcinomas (HNSCC) are the sixth most common malignancy globally, and an increasing proportion of oropharyngeal HNSCCs are associated with the human papillomavirus (HPV). Patients with HPV-associated tumours have markedly improved overall and disease-specific survival compared with their HPV-negative counterparts when treated with chemoradiation. Although the difference in outcomes between these two groups is clearly established, the mechanism underlying these differences remains an area of investigation. Data from preclinical, clinical and genomics studies have started to suggest that an increase in radio-sensitivity of HPV-positive HNSCC may be responsible for improved outcomes, the putative mechanisms of which we will review here. The Cancer Genome Atlas and others have recently documented a multitude of molecular differences between HPV-positive and HPV-negative tumours. Preclinical investigations by multiple groups have explored possible mechanisms of increased sensitivity to therapy, including examining differences in DNA repair, hypoxia and the immune response. In addition to differences in the response to therapy, some groups have started to investigate phenotypic differences between the two diseases, such as tumour invasiveness. Finally, we will conclude with a brief review of ongoing clinical trials that are attempting to de-escalate treatment to minimise long-term toxicity while maintaining cure rates. New insights from preclinical and genomic studies may eventually lead to personalised treatment paradigms for HPV-positive patients.

Gallstone ileus: retrospective review of a single centre's experience using two surgical procedures.

INTRODUCTION: Gallstone ileus is responsible for 1-3 percent of cases of mechanical small bowel obstruction. Debate continues regarding choice of optimal surgical procedure. One-stage procedure includes enterolithotomy, cholecystectomy and repair of fistula at the same setting, whereas staged procedure includes enterolithotomy alone, with fistula repair at a later stage. This study aims to determine factors influencing choice of surgical procedure in patients with gallstone ileus. METHODS: Data was collected for patients diagnosed with gallstone ileus between 1990 and 2005. Five patients underwent enterolithotomy alone (Group 1), while the remaining five patients underwent enterolithotomy with cholecystectomy and repair of fistula as a single stage procedure (Group 2). RESULTS: In Group 1, patients presented late with deranged physiological parameters and pre-existing comorbidities accounting for an American Society of Anesthesiologists (ASA) score of 3 or above. In Group 2, patients presented early with preserved physiological status accounting for an ASA score of 2. The mean operative time was 126 +/- 23 minutes in Group 1 and 245 +/- 54.4 minutes in Group 2. There was no mortality, three patients in Group 1 had superficial wound infection, and one patient in Group 2 had injury to the common bile duct necessitating hepaticojejunostomy. The mean follow-up period was 3.5 +/- 1.5 years. None of the patients in both groups had recurrent symptoms requiring further intervention. CONCLUSION: Choice of surgical procedure was largely determined by the clinical status of the patient. Single-stage procedure was performed in haemodynamically-stable patients, while enterolithotomy alone was considered sufficient for unstable patients.

The effects of an intravenous infusion of isosorbide dinitrate during open heart surgery.

Recent awareness of right ventricular dysfunction during open heart surgery has focused attention upon the importance of protection of the right ventricle and control of right ventricular afterload for the maintenance of the cardiac output. Conventional doses of systemic vasodilators, when used to reduce pulmonary vascular resistance, may produce systemic hypotension, reduce coronary arterial perfusion and even lower the cardiac output. A study of the effects of bolus intravenous isosorbide dinitrate during open heart surgery showed that following cardiopulmonary bypass intravenous isosorbide dinitrate produced highly significant falls in pulmonary artery pressure and induced active pulmonary vasodilatation without systemic side-effects or reduced atrial filling procedures. Treatment with intravenous isosorbide dinitrate by low-dose infusion during and after open heart surgery significantly lowered mean pulmonary artery pressure and pulmonary vascular resistance (P less than 0.001) in patients receiving no sympathomimetic drug support. The results suggest that possibly the effect of low-dose isosorbide dinitrate following cardiopulmonary by-pass is exerted predominantly on the right ventricular afterload if systemic arterial pressure is not elevated. This may have applications in the management of pulmonary hypertension and of acute right heart failure following cardiac surgery.

Lipoxygenase inhibitory tetraketones: potential Remedial source for inflammation and asthma / Las tetracetonas inhibidoras de la lipoxigenasa: fuente potencial Remedial para la inflamación y el asma

OBJETIVE: A series of tetraketones has been synthesized by way of a one pot synthesis and screened for inhibitory activity against the enzyme lipoxygenase. METHOD: An efficient and high yielding one pot synthesis to tetraketones [2-22] has been developed by way of tetraethyl ammonium bromide (Et4N+Br-) mediated condensation of cyclohexane-1, 3-dione [1] with a variety of aldehydes. Lipoxygenase enzyme solution was prepared so that enzyme concentration in reaction mixture was adjusted to give rates of 0.05 absorbance/minute. The test compounds were prepared in methanol of concentrations 50, 25, 12.5, 6.25 and 3.125 µM. The reaction mixture contained 160 µL (100 mM) sodium phosphate buffer (pH 8.0), 10µL of test-compound solution and 20µL of lipoxygenase solution. The contents were mixed and incubated for 10 minutes at 25ºC. The reaction was then initiated by the addition of 10µL substrate solution (linoleic acid, 0.5 mM, 0.12% w/v tween 20 in the ratio of 1:2), with the formation of (9Z,11E)-(13S)-13-hydroperoxyoctadeca-9,11-dienoate, the change of absorbance at 234 nm was followed for 6 minutes. The concentrations of the test compounds that inhibited the lipoxygenase activity by 50% (IC50) were determined by monitoring the effect of increasing concentrations of these compounds in the assays on the degree of inhibition. The IC50 values were calculated by means of the EZ-Fit Enzyme-Kinetics Program (Perrella Scientific Inc., Amherst, USA). RESULT: The tetraketones [2-22] were synthesized in high yields (91-98%) using mild reaction conditions. Most of these compounds showed significant inhibitory activity against the enzyme lipoxygenase. It was found that the presence of substituents which increase delocalization of electrons enhances the inhibitory activity. CONCLUSION: It is concluded that the study is likely to lead to the discovery of therapeutically efficient agents against important disorders such as inflammation and asthma.

OBJETIVO: Una serie de tetracetonas han sido sintetizadas mediante síntesis de varios pasos en un solo reactor (one-pot), y examinadas en relación con su actividad inhibitoria frente a la enzima lipoxigenasa. MÉTODO: Una síntesis one-pot de un rendimiento alto y eficiente para la obtención de tetracetonas (2-22) ha sido desarrollada mediante bromuro amónico tetraetílico (Et4N+Br-) - de ciclohexano-1,3-diona, con una variedad de aldehídos. La solución de enzima lipoxigenasa fue preparada de modo que la concentración de la enzima en las mezcla de la reacción fue ajustada para que diera tasas de 0.05 absorbancia/minuto. Los compuestos de la prueba fueron preparados en metanol de concentraciones (50, 25, 12.5, 6.25 y 3.125 µM). La mezcla de reacción contenía 160 µL (100 mM) de un tampón (buffer) de fosfato de sodio (pH 8.0), 10µL de solución de compuesto de prueba, y 20µL de solución de lipoxigenasa. Los contenidos fueron mezclados e incubados por 10 minutos a 25ºC. La reacción fue iniciada entonces por la adición de 10µL de solución substrato (ácido linoleico, 0.5 mM, 0.12% p/v tween 20 en proporción de 1:2), con la formación de (9Z, 11E)-(13S)-13-hidroperoxioctadeca-9,11-dienoato, el cambio de absorbancia a 234 nm fue seguido por 6 minutos. Las concentraciones de los compuestos de prueba que inhibían la actividad de la lipoxigenasa en un 50% (IC50) fueron determinadas monitoreando el efecto del aumento de las concentraciones de estos compuestos en los ensayos sobre el grado de inhibición. Los valores IC50 fueron calculados mediante el Programa Cinética de la Enzima EZ-Fit (Perrella Scientific Inc., Amherst, USA). RESULTADOS: Las tetracetonas (2-22) se sintetizaron con elevados rendimientos (91-98%) usando condiciones de reacción leve. La mayoría de estos compuestos mostraron una actividad inhibitoria significativa frente a la enzima lipoxigenasa. Se halló que la presencia de sustituyentes que aumentan la deslocalización de los electrones contribuye a mejorar la actividad inhibitoria. CONCLUSIÓN: Se concluye que es probable que el estudio conduzca al descubrimiento de agentes terapéuticamente eficientes frente a trastornos importantes tales como la inflamación y el asma.