Adenovirus-mediated expression of MOAP-1, Bax and RASSF1A antagonizes chemo-drug resistance of human breast cancer cells expressing cancer stem cell markers.

Cancer is one of the major leading causes of mortality globally and chemo-drug-resistant cancers pose significant challenges to cancer treatment by reducing patient survival rates and increasing treatment costs. Although the mechanisms of chemoresistance vary among different types of cancer, cancer cells are known to share several hallmarks, such as their resistance to apoptosis as well as the ability of cancer stem cells to produce metastatic daughter cells that are resistant to chemotherapy. To address the issue of chemo-drug resistance in cancer cells, a tetracistronic expression construct, Ad-MBR-GFP, encoding adenovirus-mediated expression of MOAP-1, Bax, RASSSF1A, and GFP, was generated to investigate its potential activity in reducing or inhibiting the chemo-drug resistant activity of the human breast cancer cells, MCF-7-CR and MDA-MB-231. When infected by Ad-MBR-GFP, the cancer cells exhibited round cell morphology and nuclei condensation with positive staining for annexin-V. Furthermore, our results showed that both MCF-7-CR and MDA-MB-231 cells stained positively for CD 44 and negatively for CD 24 (CD44+/CD24-) with high levels of endogenous ALDH activity whereas SNU-1581 breast cancer cells were identified as CD 44-/CD 24- cells with relatively low levels of endogenous ALDH activity and high sensitivity toward chemo-drugs, suggesting that both CD 44 and ALDH activity contribute to chemo-drug resistance. Moreover, both MCF-7-CR and MDA-MB-231 cells showed strong chemo-drug sensitivity to cisplatin when the cells were infected by Ad-MBR-GFP, leading to 9-fold and 2-fold reduction in the IC 50 values when compared to cisplatin treatment alone, respectively. The data were further supported by 3D (soft agar) and spheroid cell models of MCF-7-CR and MDA-MB-231 cells which showed a 2-fold reduction of a number of cell colonies and spheroid size when treated with both Ad-MBR-GFP and cisplatin, and compared to control. Other than chemo-sensitivity, Ad-MBR-GFP-infected cancer cells retarded cell migration. Flow cytometry analysis showed that the mechanism of action of Ad-MBR-GFP involved cell cycle arrest at the G1 phase and inhibition of cellular DNA synthesis. Taken together, our investigation showed that Ad-MBR-GFP mediated chemo-drug sensitization in the infected cancer cells involved the activation of apoptosis signaling, cell cycle arrest, and inhibition of DNA synthesis, suggesting that Ad-MBR-GFP is potentially efficacious for the treatment of chemo-drug resistant cancers.

Cascade testing for hereditary cancer in Singapore: how population genomics help guide clinical policy.

Hereditary Cancer makes up around 5-10% of all cancers. It is important to diagnose hereditary cancer in a timely fashion, as not only do patients require long-term care from a young age, but their relatives also require management. The main approach to capture at-risk relatives is cascade testing. It involves genetic testing of relatives of the first detected carrier of a pathogenic variant in a family i.e. the proband. The current standard of care for cascade testing is a patient-mediated approach. Probands are then advised to inform and encourage family members to undergo genetic testing. In Singapore, cascade testing is inefficient, around 10-15%, lower than the 30% global average. Here, we describe the cascade testing process and its effort to increase testing in Singapore. Precision Health Research, Singapore (PRECISE), was set up to coordinate Singapore's National Precision Medicine strategy and has awarded five clinical implementation pilots, with one of them seeking to identify strategies for how cascade testing for hereditary cancer can be increased in a safe and cost-efficient manner. Achieving this will be done through addressing barriers such as cost, manpower shortages, exploring a digital channel for contacting at-risk relatives, and getting a deeper insight into why genetic testing gets declined. If successful, it will likely result in care pathways that are a cost-effective public health intervention for identifying individuals at risk. Surveillance and management of those unaffected at-risk individuals, if caught early, will result in improved patient outcomes, and further reduce the healthcare burden for the economy.

Impact of UV radiation on Mxene-mediated tubulin dissociation and mitochondrial apoptosis in breast cancer cells.

Breast cancer is a global health concern that requires personalized therapies to prevent relapses, as conventional treatments may develop resistance over time. Photothermal therapy using spectral radiation or intense light emission is a broad-spectrum treatment that induces hyperthermia-mediated cancer cell death. MXene, a two-dimensional material, has been reported to have potential biological applications in photothermal therapy for cancer treatment. In this study, we investigated the apoptotic activity of MXene and UV-irradiated MXene in MCF-7 breast cancer cells by treating them with varying concentrations of MXene. The cytotoxicity of MXene and UV was evaluated by analyzing cellular morphology, nuclei condensation, caspase activation, and apoptotic cell death. We also assessed the effect of the combined treatment on the expression and cellular distribution of Tubulin, a key component of microtubules required for cell division. At low concentrations of MXene (up to 100 µg/ml), the level of cytotoxicity in MCF-7 cells was low. However, the combined treatment of MXene and UV resulted in a synergistic increase in cytotoxicity, causing rounded cellular morphology, condensed nuclei, caspase activation, and apoptotic cell death. Furthermore, the treatment reduced Tubulin protein expression and cellular distribution, indicating a potent inducer of cell death with potential application for cancer treatment. The study demonstrates that the combined treatment of MXene and UVB irradiation is a promising strategy for inducing apoptotic cell death in breast cancer cells, suggesting its potential as a therapeutic intervention for breast cancer.

Controversies in terminology associated with management of BCG-unresponsive NMIBC in Asia-Pacific.

OBJECTIVES: Examine the understanding of terminologies and management patterns of bacillus Calmette-Guérin (BCG)-unresponsive nonmuscle invasive bladder cancer (NMIBC) in six territories in Asia-Pacific. METHODS: This study involved two phases: (1) a survey with 32 urologists and 7 medical oncologists (MOs) and (2) a factorial experiment and in-depth interviews with 23 urologists and 2 MOs. All clinicians had ≥8 years' experience managing NMIBC patients in Australia, Hong Kong, Japan, South Korea, Singapore, and Taiwan. Data from Phase 1 were summarized using descriptive statistics; content and thematic analyses applied in Phase 2. RESULTS: In phase 1, 35% of clinicians defined BCG-unresponsive as BCG-refractory, -relapse and -resistant, 6% defined it as BCG-refractory and -relapse; 22% classified BCG-failure as BCG-refractory, -relapse, -resistant, and when muscle-invasive bladder cancer is detected. If eligible and willing, 50% (interquartile range [IQR], 50%-80%) of BCG-unresponsive patients would undergo radical cystectomy (RC), and 50% (IQR 20%-50%) of RC-eligible patients would receive bladder-sparing treatment or surveillance. In phase 2, we found that 32%, 88%, and 48% of clinicians, respectively, used "BCG-unresponsive," "BCG-refractory," and "BCG-relapse" in clinical practice but with no consistent interpretation of the terms. Compared with EAU definitions, 8%-60% of clinicians appropriately classified 9 tumor types that are persistent or recurrent after adequate BCG. Fifty percent of clinicians mentioned a lack of bladder-preserving treatment that outperforms RC in quality of life as a reason to retreat BCG-unresponsive patients with BCG. CONCLUSIONS: Our study revealed varied understanding and application of BCG-unresponsive terminologies in practice. There is a need for a uniform and simple definition of BCG-unresponsive disease in Asia-Pacific.

Risk stratification and management of non-muscle-invasive bladder cancer: A physician survey in six Asia-Pacific territories.

OBJECTIVES: Multiple clinical practice guidelines, conflicting evidence, and physician perceptions result in variations in risk stratification among patients with non-muscle-invasive bladder cancer (NMIBC). This study aims to describe the extent of this variation and its impact on management approaches in the Asia-Pacific region. METHODS: We conducted a cross-sectional survey involving 32 urologists and seven medical oncologists with ≥8 years of experience managing early-stage bladder cancer patients across Australia, Hong Kong, Japan, South Korea, Singapore, and Taiwan. The physicians completed an anonymous questionnaire that assessed their risk stratification and respective management approaches, based on 19 NMIBC characteristics. For each NMIBC characteristic, they were required to select one risk group, and their most preferred management approach. RESULTS: Our results demonstrated a higher consensus on risk classification versus management approaches. More than 50% of the respondents agreed on the risk classification of all NMIBC characteristics, but 42% or fewer chose the same treatment option as their preferred choice for all but two characteristics-existence of variant histology (55%) and persistent high-grade T1 disease on repeat resection (52%). Across territories, there was the greatest variation in preferred treatment options (i.e., no treatment, intravesical chemotherapy, or Bacillus Calmette-Guérin [BCG] treatment) for intermediate-risk patients and the highest consensus on the treatment of very high-risk patients, namely radical cystectomy. CONCLUSIONS: Our study revealed considerable variation in risk stratification and management of NMIBC in the region. It is critical to develop practical algorithms to facilitate the recognition of NMIBC and standardize the treatment of NMIBC patients.

Clinical decision pathway and management of locally advanced head and neck squamous cell carcinoma: A multidisciplinary consensus in Asia-Pacific.

OBJECTIVES: To develop consensus on patient characteristics and disease-related factors considered in deciding treatment approaches for locally advanced head and neck squamous cell carcinoma (LA-HNSCC) based on real-world treatment patterns in 4 territories in Asia-Pacific. METHODS: A three-round modified Delphi involving a multidisciplinary panel of HN surgeons, medical oncologists, and radiation oncologists was used. Of 41 panelists recruited, responses of 26 from Australia, Japan, Singapore, and Taiwan were analyzed. All panelists had ≥five years' experience managing LA-HNSCC patients and treated ≥15 patients with LA-HNSCC annually. RESULTS: All statements on definitions of LA-HNSCC, treatment intolerance and cisplatin dosing reached consensus. 4 of 7 statements on unresectability, 2 of 4 on adjuvant chemoradiotherapy, 7 of 13 on induction chemotherapy, 1 of 8 on absolute contraindications and 7 of 11 on relative contraindications to high-dose cisplatin did not reach consensus. In all territories except Taiwan, high-dose cisplatin was preferred in definitive and adjuvant settings for patients with no contraindications to cisplatin; weekly cisplatin (40 mg/m2) preferred for patients with relative contraindications to high-dose cisplatin. For Taiwan, the main treatment option was weekly cisplatin. For patients with absolute contraindications to cisplatin, carboplatin ± 5-fluorouracil or radiotherapy alone were preferred alternatives in both definitive and adjuvant settings. CONCLUSION: This multidisciplinary consensus provides insights into management of LA-HNSCC in Asia-Pacific based on patient- and disease-related factors that guide selection of treatment modality and systemic treatment. Despite strong consensus on use of cisplatin-based regimens, areas of non-consensus showed that variability in practice exists where there is limited evidence.

Novel Variants and Clinical Characteristics of 16 Patients from Southeast Asia with Genetic Variants in Neurofibromin-1.

Neurofibromatosis type 1 (NF1) is one of the most common inherited disorders. It is caused by mutations in the neurofibromin-1 gene ( NF1 ) and affects the formation and growth of nerve tissues. More than 3,600 pathogenic variants in the NF1 gene have been identified from patients with most of the germline variants are from the Western populations. We found 16 patients (15 Chinese and 1 Asian Indian) who had heterozygous variants in NF1 through targeted next-generation sequencing. There were 15 different variants: 4 frameshift, 4 nonsense, 5 missense, and 2 splice variants. One nonsense variant and three frameshift variants had never been reported in any population or patient database. Twelve of the 16 patients met the NF1 diagnostic criteria, and each was found to have a pathogenic or likely pathogenic variant. Three different missense variants of unknown significance were discovered in the other four patients who did not meet NF1 diagnostic criteria. Our findings add four novel variants to the list of genetic mutations linked to NF1's various clinical manifestations.

Hepatic glutamine synthetase controls N5-methylglutamine in homeostasis and cancer.

Glutamine synthetase (GS) activity is conserved from prokaryotes to humans, where the ATP-dependent production of glutamine from glutamate and ammonia is essential for neurotransmission and ammonia detoxification. Here, we show that mammalian GS uses glutamate and methylamine to produce a methylated glutamine analog, N5-methylglutamine. Untargeted metabolomics revealed that liver-specific GS deletion and its pharmacological inhibition in mice suppress hepatic and circulating levels of N5-methylglutamine. This alternative activity of GS was confirmed in human recombinant enzyme and cells, where a pathogenic mutation in the active site (R324C) promoted the synthesis of N5-methylglutamine over glutamine. N5-methylglutamine is detected in the circulation, and its levels are sustained by the microbiome, as demonstrated by using germ-free mice. Finally, we show that urine levels of N5-methylglutamine correlate with tumor burden and GS expression in a ß-catenin-driven model of liver cancer, highlighting the translational potential of this uncharacterized metabolite.

Engineered Oncolytic Adenoviruses: An Emerging Approach for Cancer Therapy.

Cancer is among the major leading causes of mortality globally, and chemotherapy is currently one of the most effective cancer therapies. Unfortunately, chemotherapy is invariably accompanied by dose-dependent cytotoxic side effects. Recently, genetically engineered adenoviruses emerged as an alternative gene therapy approach targeting cancers. This review focuses on the characteristics of genetically modified adenovirus and oncology clinical studies using adenovirus-mediated gene therapy strategies. In addition, modulation of the tumor biology and the tumor microenvironment as well as the immunological responses associated with adenovirus-mediate cancer therapy are discussed.

MCC is a centrosomal protein that relocalizes to non-centrosomal apical sites during intestinal cell differentiation.

The gene mutated in colorectal cancer (MCC) encodes a coiled-coil protein implicated, as its name suggests, in the pathogenesis of hereditary human colon cancer. To date, however, the contributions of MCC to intestinal homeostasis and disease remain unclear. Here, we examine the subcellular localization of MCC, both at the mRNA and protein levels, in the adult intestinal epithelium. Our findings reveal that Mcc transcripts are restricted to proliferating crypt cells, including Lgr5+ stem cells, where the Mcc protein is distinctly associated with the centrosome. Upon intestinal cellular differentiation, Mcc is redeployed to the apical domain of polarized villus cells where non-centrosomal microtubule organizing centers (ncMTOCs) are positioned. Using intestinal organoids, we show that the shuttling of the Mcc protein depends on phosphorylation by casein kinases 1δ and ε, which are critical modulators of WNT signaling. Together, our findings support a role for MCC in establishing and maintaining the cellular architecture of the intestinal epithelium as a component of both the centrosome and ncMTOC.

Idiopathic upper extremity deep vein thrombosis: A systematic review.

BACKGROUND: Idiopathic upper extremity deep vein thrombosis (UEDVT) management is controversial and ranges from anticoagulation alone to the addition of further interventions such as thrombolysis and decompressive surgery. OBJECTIVES: The objective of this systematic review was to assess the effects of anticoagulation alone compared to anticoagulation with additional interventions such as thrombolysis or decompressive surgery on the incidence of recurrent UEDVT and post-thrombotic syndrome (PTS) in patients with idiopathic UEDVT (including those associated with the oral contraceptive pill). PATIENTS/METHODS: A systematic search was conducted for studies which focused on acute UEDVT treatment defined as therapies starting within 4 weeks of symptom onset. We limited studies to those that recruited 10 or more subjects and involved at least 6 weeks to 12 months anticoagulation alone or together with additional interventions with at least 6-month follow-up. Primary outcomes were symptomatic recurrent radiologically confirmed UEDVT and PTS. Secondary outcomes were symptomatic venous thromboembolism, bleeding and mortality. RESULTS: We found seven studies which reported recurrent UEDVT rates and five that reported PTS rates. All studies were retrospective or cross-sectional. None compared anticoagulation alone to anticoagulation with additional intervention. Study heterogeneity precluded meta-analysis and risk of bias was moderate to serious. Recurrent UEDVT occurred in 0% to 12% post-anticoagulation alone and 0% to 23% post-additional interventions. PTS rates varied from 4% to 32% without severe PTS. Only limited studies reported on our secondary outcomes. CONCLUSION: There is limited evidence behind idiopathic UEDVT management. Prospective comparative studies in this area are essential.

New synthetic phenylquinazoline derivatives induce apoptosis by targeting the pro-survival members of the BCL-2 family.

Chemo-resistant cancer cells acquire robust growth potential through cell signaling mechanisms such as the down-regulation of tumor suppressors and the up-regulation of pro-survival proteins, respectively. To overcome chemo-resistance of cancer, small molecule drugs that interact with the cell signaling proteins to enhance sensitization of cancer cells toward cancer therapies are likely to be effective for the treatment of chemo-drug resistant cancer. To identify high potency small molecules, a series of ten novel phenylquinazoline derivatives were synthesized to determine their cellular effects in MCF-7 and MCF-7- cisplatin-resistant (CR) human breast cancer cells which led to the identification of two bioactive compounds, SMS-IV-20 and SMS-IV-40, that exhibited an elevated level of cytotoxicity against the human breast cancer cells and spheroid cells. In addition, both compounds enhanced chemo-sensitization of the human breast cancer cells that were genetically engineered to express the tumor suppressor and pro-apoptotic proteins, MOAP-1, Bax, and RASSF1a (MBR), suggesting that the compounds interact with the MBR signaling pathway. Furthermore, when MCF-7-CR cells were treated with SMS-IV-20 and SMS-IV-40 in the presence of ABT-737, a BCL-XL and BCL-2 inhibitor, enhanced chemo-sensitization was observed, suggesting SMS-IV-20 and SMS-IV-40 exert antagonistic activity to regulate the functional activity of BCL-2 and BCL-XL. Western blot analysis showed that both SMS-IV-20 and SMS-IV-40 induced down-regulation of BCL-2 or both BCl-2 and BCL-XL expression, respectively while promoting the release of mitochondrial Cytochrome C. Taken together, the data showed that SMS-IV-20 and SMS-IV-40 are potent activators of apoptosis that enhance chemo-sensitization through their antagonistic actions on the pro-survival activity of the BCl-2 family in human cancer cells.

A 5-FU Precursor Designed to Evade Anabolic and Catabolic Drug Pathways and Activated by Pd Chemistry In Vitro and In Vivo.

5-Fluorouracil (5-FU) is an antineoplastic antimetabolite that is widely administered to cancer patients by bolus injection, especially to those suffering from colorectal and pancreatic cancer. Because of its suboptimal route of administration and dose-limiting toxicities, diverse 5-FU prodrugs have been developed to confer oral bioavailability and increase the safety profile of 5-FU chemotherapy regimens. Our contribution to this goal is presented herein with the development of a novel palladium-activated prodrug designed to evade the metabolic machinery responsible for 5-FU anabolic activation and catabolic processing. The new prodrug is completely innocuous to cells and highly resistant to metabolization by primary hepatocytes and liver S9 fractions (the main metabolic route for 5-FU degradation), whereas it is rapidly converted into 5-FU in the presence of a palladium (Pd) source. In vivo pharmokinetic analysis shows the prodrug is rapidly and completely absorbed after oral administration and exhibits a longer half-life than 5-FU. In vivo efficacy studies in a xenograft colon cancer model served to prove, for the first time, that orally administered prodrugs can be locally converted to active drugs by intratumorally inserted Pd implants.

Alpha-Mangostin Activates MOAP-1 Tumor Suppressor and Mitochondrial Signaling in MCF-7 Human Breast Cancer Cells.

α-Mangostin, one of the major constituents of Garcinia mangostana, has been reported to possess several biological activities, including antioxidant, anti-inflammatory, antibacterial, and cytotoxic activities associated with the inhibition of cell proliferation and activation of apoptosis. However, the cellular signaling pathway mediated by α-mangostin has not been firmly established. To investigate the cellular activities of α-mangostin, human cancer cells, MCF-7 and MCF-7-CR cells, were treated with α-mangostin to measure the cellular responses, including cytotoxicity, protein-protein interaction, and protein expression. Cancer cells stably expressed Myc-BCL-XL and HA-MOAP-1 were also included in the studies to delineate the cell signaling events mediated by α-mangostin. Our results showed that the apoptosis signaling mediated by α-mangostin involves the upregulation of endogenous MOAP-1, which interacts with α-mangostin activated BAX (act-BAX) while downregulating the expression of BCL-XL. Moreover, α-mangostin was found to induce BAX oligomerization, the release of mitochondrial cytochrome C, and activation of caspase in MCF-7 cells. In overexpression studies, MCF-7 cells and spheroids stably expressed HA-MOAP-1 and Myc-BCL-XL exhibited differential chemosensitivity toward α-mangostin in which the stable clones expressing HA-MOAP-1 and MYC-BCL-XL were chemosensitive and chemoresistant to the apoptosis signaling events mediated by α-mangostin, respectively, when compared to untreated cells. Together, the data suggest that the cytotoxicity of α-mangostin involves the activation of MOAP-1 tumor suppressor and its interaction with act-BAX, leading to mitochondria dysfunction and cell death.

Idiopathic Intracranial Hypertension in a Malaysian Preschooler.

A four-year-old girl presented with a three-day history of squint and irritability. Examination showed reduced visual acuity in both eyes, the presence of a false localizing sign, and bilateral optic disc swelling. On investigation, her blood laboratory workup was within the normal range. Imaging of the brain showed no evidence of a space-occupying lesion or cerebral venous sinus thrombosis. The lumbar puncture opening pressure was 27cmH20 and the cerebrospinal fluid workup was normal. The diagnosis of idiopathic intracranial hypertension (IIH) was made based on the diagnostic criteria for pseudotumor cerebri syndrome. She was successfully treated with acetazolamide with resolved symptoms and signs. This highlights the possibility of IIH presenting with inconspicuous symptoms in preschool children, which needs a high index suspicion by clinicians. Hence, solving the challenges in the workup, especially in children, is very crucial.

Implications of Dual Practice on Cataract Surgery Waiting Time and Rescheduling: The Case of Malaysia.

BACKGROUND: Dual practice was implemented in selected Ministry of Health Malaysia hospitals to reduce brain drain and provide an alternative for patients willing to pay higher user fees to seek prompt treatment from the specialist of their choice. This study aimed to assess the implications of dual practice on waiting time and rescheduling for cataract surgery. METHODS: A retrospective study was conducted in a referral hospital. Inpatient medical records of patients who underwent cataract procedures were used to study the waiting times to surgery and rescheduling between private and public groups. RESULTS: Private patients had a considerably shorter waiting time for cataract surgery, seven times shorter compared to public patients where all surgeries were conducted after hours on weekdays or weekends. Additionally, 14.9% of public patients experienced surgery rescheduling, while all private patients had their surgeries as planned. The main reason for surgery rescheduling was the medical factor, primarily due to uncontrolled blood pressure and upper respiratory tract infection. CONCLUSION: Private service provision utilizing out-of-office hours slots for cataract surgery optimizes public hospital resources, allowing shorter waiting times and providing an alternative to meet healthcare needs.

An NKX2-1GFP and TP63tdTomato dual fluorescent reporter for the investigation of human lung basal cell biology.

Basal cells are multipotent stem cells responsible for the repair and regeneration of all the epithelial cell types present in the proximal lung. In mice, the elusive origins of basal cells and their contribution to lung development were recently revealed by high-resolution, lineage tracing studies. It however remains unclear if human basal cells originate and participate in lung development in a similar fashion, particularly with mounting evidence for significant species-specific differences in this process. To address this outstanding question, in the last several years differentiation protocols incorporating human pluripotent stem cells (hPSC) have been developed to produce human basal cells in vitro with varying efficiencies. To facilitate this endeavour, we introduced tdTomato into the human TP63 gene, whose expression specifically labels basal cells, in the background of a previously described hPSC line harbouring an NKX2-1GFP reporter allele. The functionality and specificity of the NKX2-1GFP;TP63tdTomato hPSC line was validated by directed differentiation into lung progenitors as well as more specialised lung epithelial subtypes using an organoid platform. This dual fluorescent reporter hPSC line will be useful for tracking, isolating and expanding basal cells from heterogenous differentiation cultures for further study.

Does estrogen regulate vitellogenin synthesis in corals?

Most broadcast spawner corals have a vitellogenic phase that lasts at least 6 months. It is established that estrogen regulates vitellogenin synthesis in vertebrates. Although some research have been conducted on the physiological role of sex steroids in corals, little is known about their involvement in oocyte development. This study aimed to detect steroid hormones - progesterone, testosterone, and estradiol-17ß (E2) - in Acropora tenuis and study the relationships between vitellogenesis/vitellogenin synthesis and these steroids. This study also investigated the effect of E2 on vitellogenin synthesis in corals and identified steroidogenic enzymes in A. tenuis genome. Branches from tagged coral colonies were collected monthly from March to November. Histological observations showed that oocytes were vitellogenic from March to May (Stage IV and V), but not in June, and that gonads were occupied by immature oocytes in September (Stage I). Real-time qPCR revealed that vitellogenin (vg1 and vg2) transcript levels in coral branches were high in April and May, implying that corals actively underwent vitellogenesis during these months, and spawned before June. Liquid chromatography-mass spectrometry revealed that E2 could be detected in coral branches in March, April, and May, but not in June, whereas testosterone and progesterone did not fluctuate much in the same months. Immersing branches in E2-containing seawater failed to increase vitellogenin transcript levels. The results indicate that E2 is involved in oogenesis but does not positively regulate vitellogenin synthesis. Steroidogenic enzymes (except CYP19A) were identified in A. tenuis, suggesting that corals may endogenously synthesize progestogens and androgens from cholesterol.

Singapore Undiagnosed Disease Program: Genomic Analysis aids Diagnosis and Clinical Management.

OBJECTIVE: Use next-generation sequencing (NGS) technology to improve our diagnostic yield in patients with suspected genetic disorders in the Asian setting. DESIGN: A diagnostic study conducted between 2014 and 2019 (and ongoing) under the Singapore Undiagnosed Disease Program. Date of last analysis was 1 July 2019. SETTING: Inpatient and outpatient genetics service at two large academic centres in Singapore. PATIENTS: Inclusion criteria: patients suspected of genetic disorders, based on abnormal antenatal ultrasound, multiple congenital anomalies and developmental delay. EXCLUSION CRITERIA: patients with known genetic disorders, either after clinical assessment or investigations (such as karyotype or chromosomal microarray). INTERVENTIONS: Use of NGS technology-whole exome sequencing (WES) or whole genome sequencing (WGS). MAIN OUTCOME MEASURES: (1) Diagnostic yield by sequencing type, (2) diagnostic yield by phenotypical categories, (3) reduction in time to diagnosis and (4) change in clinical outcomes and management. RESULTS: We demonstrate a 37.8% diagnostic yield for WES (n=172) and a 33.3% yield for WGS (n=24). The yield was higher when sequencing was conducted on trios (40.2%), as well as for certain phenotypes (neuromuscular, 54%, and skeletal dysplasia, 50%). In addition to aiding genetic counselling in 100% of the families, a positive result led to a change in treatment in 27% of patients. CONCLUSION: Genomic sequencing is an effective method for diagnosing rare disease or previous 'undiagnosed' disease. The clinical utility of WES/WGS is seen in the shortened time to diagnosis and the discovery of novel variants. Additionally, reaching a diagnosis significantly impacts families and leads to alteration in management of these patients.