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Durable remissions occur in ~50% of LSFL patients whereas ASFL is incurable. In keeping with recent IHC CD8 protein data (Los-de Vries, Bld Adv 2022), CD8A gene expression was raised in stage I LSFL vs. 68 ASFL patients treated with immunochemotherapy ( I p i = 0.02). Recently, we demonstrated in limited and advanced stage FL (LSFL, ASFL), that patients with longer remissions had raised clonally expanded intratumoral CD8+ T cells (Tobin, JCO 2019). [Extracted from the article]

Soil compaction impedes plant growth by negatively affecting water infiltration, soil aeration, access to soil water and nutrients, and consequently restricts root development. Previous studies to alleviate such problems in rehabilitated mine soils using a single amendment material did not provide a long lasting solution. The aim of the study was to quantify the role of different amendments in alleviating selected soil hydraulic properties. Five single amendment materials and three different blends of amendments were mixed thoroughly with degraded mine soil in a 1:3 (amendment:soil) ratio and packed in columns. Two additional unamended soils with different bulk densities were included as benchmarks. In general, the application of amendments reduced bulk density (BD) by 4–20%, enhanced infiltration rate by 15–70%, increased porosity by 5–35% and increased plant available water (PAW) by 9–33% compared with the unamended soils. Between amendments, the blends of amendments reduced BD by 9–16%, enhanced infiltration rate by 17–59%, increased porosity by 6–32%, and PAW by 4–28% compared with single amendments. The study showed that a blend of amendments have better soil restoration capacity through improving porosity, infiltration rate and plant available water. A blend of amendments therefore has the potential to be a sustainable and an important component of a rehabilitation strategy for surface coal mined soils. [ABSTRACT FROM AUTHOR]

Background: Immune checkpoint inhibition (ICI) has led to unprecedented outcomes for melanoma patients but is associated with toxicity. ICI resumption after high grade irAEs poses a significant challenge in the clinical management of melanoma patients and there are no biomarkers that can help identify patients that might benefit from resuming treatment. This study aims to determine if circulating tumor DNA (ctDNA) levels at the time of treatment-limiting irAE could guide treatment decisions in this clinical context., Methods: This is a retrospective exploratory biomarker study from 34 patients treated with combination ICI for stage IV melanoma. Patients had a treatment-limiting toxicity and a baseline plasma collection prior to commencing ICI and within 6 weeks of stopping therapy. Blood samples were tested for ctDNA at baseline and cessation therapy., Results: Median progression free survival (PFS) and overall survival (OS) have not been reached (24-month PFS rate 54% and OS rate 72.3%). PD occurred in 47% (16/34) of patients. Median PFS with detectable ctDNA from plasma collected at the time of toxicity was 6.5 months while not reached (NR) with undetectable levels (HR: 4.0, 95% CI 0.95-17.5, p=0.0023). Median OS with detectable ctDNA at cessation for toxicity was 19.4 months and NR for undetectable ctDNA (HR: 3.9, 95%CI 20.8-18.6, p=0.024). Positive ctDNA at the time of cessation was highly specific (specificity 0.94, 95% CI 0.74-0.99, PPV 0.88, 95% CI 0.53-0.99). However, ctDNA negativity has low sensitivity as a predictor of ongoing disease control (sensitivity 0.437, 95% CI 0.23-0.67). Notably, 4/9 (44%) ctDNA negative patients who had disease progression had brain only disease progression., Conclusions: Undetectable ctDNA and CR on imaging after stopping immunotherapy for toxicity results in high rates of long-term durable control. For patients with immunotherapy related toxicity, who have persistent ctDNA at 8 - 12 weeks, the risk of disease progression is significant., Competing Interests: LW is an Advisory Board Member for Bristol Myers Squibb, AstraZeneca, Roche, Merck Sharp and Dohme and Novartis and has received travel/accommodation support from Pierre Fabre, Merck Sharp & Dohme and Bristol Myers Squibb. MM is an Advisory Board Member melanoma or immuno-oncology for Bristol Myers Squibb, AstraZeneca, Roche, and Merck Sharp & Dohme and has received travel support from Merck Sharp & Dohme, AstraZeneca, and Bristol Myers Squibb. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Warburton, Reid, Amanuel, Calapre, Millward and Gray.)

Complete biomarker workup of non-small cell lung cancer (NSCLC) specimens is essential for appropriate and timely clinical management decisions. This can be challenging to achieve from small cytology and histology specimens, with increasing numbers of molecular and immunohistochemical biomarkers required. We conducted a 5 year retrospective audit of cases at our institution to assess the diagnostic and biomarker testing adequacy rates, particularly those specimens obtained with rapid onsite evaluation (ROSE), performed by a cytopathologist and a cytology scientist or pathology trainee, including all endobronchial ultrasound guided transbronchial needle aspirations (EBUS-TBNA), CT guided lung fine needle aspirations (FNA) and CT guided lung core biopsies. A total of 5,354 cases were identified, of which 92.2% had sufficient material for diagnosis. Of the 1506 cases identified with a recorded diagnosis of lung adenocarcinoma or NSCLC, not otherwise specified, 1001 (66.5%) had biomarker testing requested. Sufficient material was available in 89.5% of cases for a complete biomarker workup which included EGFR and KRAS mutational testing (all cases), ALK, ROS1 and PD-L1 immunohistochemistry (all cases), and ALK and ROS1 FISH (as required). For EGFR and KRAS mutational testing across both cytology and histology specimens, 99% of cases were sufficient. Of the samples in which a complete biomarker workup was unable to be performed, approximately half were only insufficient due to inadequate numbers of tumour cells for PD-L1 immunohistochemistry. Excluding PD-L1 IHC, 952 (95.1%) of samples obtained with ROSE were sufficient for the remainder of the testing requirements. Next generation sequencing using a 33 gene custom AmpliSeq panel was achieved in up to 72% of cases. In conclusion, small cytology and histology specimens obtained with ROSE are suitable for predictive biomarker testing in NSCLC, although attention needs to be paid to obtaining sufficient cells (>100) for PD-L1 immunohistochemistry., (Copyright © 2023 Royal College of Pathologists of Australasia. Published by Elsevier B.V. All rights reserved.)

Abstract: Cutaneous malignant melanoma can show a wide range of cytomorphological variability, in particular exhibiting a rhabdoid appearance is not uncommon in melanoma cells; however, the phenomenon of "dedifferentiation" with loss of melanocytic immunohistochemical properties and expression of skeletal muscle immunomarkers is exceedingly rare. Owing to the rarity of such melanomas, their clinicopathological features and molecular profile remain largely unknown. In this report, we describe the clinical, immunomorphological, and molecular features of melanomas with rhabdomyosarcomatous dedifferentiation by presenting a new case and exploring the literature for the previously reported cases., Competing Interests: The authors declare no conflicts of interest., (Copyright © 2023 Wolters Kluwer Health, Inc. All rights reserved.)

Melanoma is the most aggressive form of skin cancer owing to its high propensity to metastasise in distant organs and develop resistance to treatment. The scarce treatment options available for melanoma underscore the need for biomarkers to guide treatment decisions. In this context, an attractive alternative to overcome the limitations of repeated tissue sampling is the analysis of peripheral blood samples, referred to as 'liquid biopsy'. In particular, the analysis of extracellular vesicles (EVs) has emerged as a promising candidate due to their role in orchestrating cancer dissemination, immune modulation, and drug resistance. As we gain insights into the role of EVs in cancer and melanoma their potential for clinical use is becoming apparent. Herein, we critically summarise the current evidence supporting EVs as biomarkers for melanoma diagnosis, prognostication, therapy response prediction, and drug resistance. EVs are proposed as a candidate biomarker for predicting therapeutic response to immune checkpoint inhibition. However, to realise the potential of EV analysis for clinical decision-making strong clinical validation is required, underscoring the need for further research in this area., Competing Interests: Conflict of Interest Statement None to declare., (Copyright © 2023 Elsevier Ltd. All rights reserved.)

Plasma circulating tumour DNA (ctDNA) has been suggested to be a viable biomarker of response to treatment in patients with high grade serous ovarian carcinoma (HGSOC). TP53 mutations are present in more than 90% of HGSOCs but somatic variants are distributed across all exonic regions of the gene, requiring next generation sequencing (NGS) technologies for mutational analysis. In this study, we compared the suitability of the Accel (Swift) and Oncomine (ThermoFisher) panels for identification of TP53 mutations in ctDNA of HGSOC patients (N = 10). Only 6 patients (60%) were found to have TP53 mutations using the ACCEL panel but the addition of molecular tags in the Oncomine panel improved ctDNA detection with at least one mutation detected in all cases (100%). Orthogonal validation of the 14 somatic variants found by Oncomine, using droplet digital PCR, confirmed 79% (11/14) of the identified mutations. Overall, the Oncomine panel with unique molecular identifiers (UMI) appears more useful for ctDNA analysis in HGSOC., (© 2023. The Author(s).)

Abstract: Deep penetrating nevus (DPN) is a pigmented melanocytic tumor which typically displays a wedge-shaped deep penetrating architecture. Some cases show a coexisting component resembling conventional melanocytic nevus. These morphological attributes are correlated with the acquisition of genomic alterations in the Wnt pathway on a background of underlying activating MAPK pathway mutations. Lesions with features of DPN, but displaying expansile architecture, sheet-like arrangement of cells, cytological atypia, and/or more than rare mitotic activity have been described as "atypical deep penetrating nevus" or "deep penetrating melanocytoma." The molecular correlates of these atypical morphological features are not well-established. In this case report, we describe a tumor in an 8-year-old boy with histological features of atypical DPN showing somatic BRAFV600E , beta catenin , and IDH1R132C mutations. The combination of abnormalities in MAPK and Wnt pathways with IDH1 mutations seems to be a reproducible feature in a subset of atypical DPNs. Whether this "three-hit" combination is associated with a significant risk of adverse outcome remains to be established., Competing Interests: The authors declare no conflicts of interest., (Copyright © 2022 Wolters Kluwer Health, Inc. All rights reserved.)

Targeted therapy (BRAF inhibitor plus MEK inhibitor) is now among the possible treatment options for patients with BRAF mutation-positive stage III or stage IV melanoma. This makes prompt BRAF mutation testing an important step in the management of patients diagnosed with stage III or IV melanoma; one that can help better ensure that the optimal choice of systemic treatment is initiated with minimal delay. This article offers guidance about when and how BRAF mutation testing should be conducted when patients are diagnosed with melanoma in Australia. Notably, it recommends that pathologists reflexively order BRAF mutation testing whenever a patient is found to have American Joint Committee on Cancer (AJCC)/Union for International Cancer Control (UICC) stage III or IV melanoma (i.e., any metastatic spread beyond the primary tumour) and that patient's BRAF mutation status is hitherto unknown, even if BRAF mutation testing has not been specifically requested by the treating clinician (in Australia, Medicare-subsidised BRAF V600 mutation testing does not need to be requested by the treating clinician). When performed in centres with appropriate expertise and experience, immunohistochemistry (IHC) using the anti-BRAF V600E monoclonal antibody (VE1) can be a highly sensitive and specific means of detecting BRAF V600E mutations, and may be used as a rapid and relatively inexpensive initial screening test. However, VE1 immunostaining can be technically challenging and difficult to interpret, particularly in heavily pigmented tumours; melanomas with weak, moderate or focal BRAF V600E immunostaining should be regarded as equivocal. It must also be remembered that other activating BRAF V600 mutations (including BRAF V600K ), which account for ∼10-20% of BRAF V600 mutations, are not detected with currently available IHC antibodies. For these reasons, if available and practicable, we recommend that DNA-based BRAF mutation testing always be performed, regardless of whether IHC-based testing is also conducted. Advice about tissue/specimen selection for BRAF mutation testing of patients diagnosed with stage III or IV melanoma is also offered in this article; and potential pitfalls when interpreting BRAF mutation tests are highlighted., (Copyright © 2021 Royal College of Pathologists of Australasia. Published by Elsevier B.V. All rights reserved.)

Introduction: Grades 2 and 3 gliomas (G2/3 gliomas), when combined, are the second largest group of malignant brain tumours in adults. The outcomes for G2/3 gliomas at progression approach the dismal outcomes for glioblastoma (GBM), yet there is a paucity of trials for Australian patients with relapsed G2/3 gliomas compared with patients with GBM. LUMOS will be a pilot umbrella study for patients with relapsed G2/3 gliomas that aims to match patients to targeted therapies based on molecular screening with contemporaneous tumour tissue. Participants in whom no actionable or no druggable mutation is found, or in whom the matching drug is not available, will form a comparator arm and receive standard of care chemotherapy. The objective of the LUMOS trial is to assess the feasibility of this approach in a multicentre study across five sites in Australia, with a view to establishing a national molecular screening platform for patient treatment guided by the mutational analysis of contemporaneous tissue biopsies METHODS AND ANALYSIS: This study will be a multicentre pilot study enrolling patients with recurrent grade 2/3 gliomas that have previously been treated with radiotherapy and chemotherapy at diagnosis or at first relapse. Contemporaneous tumour tissue at the time of first relapse, defined as tissue obtained within 6 months of relapse and without subsequent intervening therapy, will be obtained from patients. Molecular screening will be performed by targeted next-generation sequencing at the reference laboratory (PathWest, Perth, Australia). RNA and DNA will be extracted from representative formalin-fixed paraffin embedded tissue scrolls or microdissected from sections on glass slides tissue sections following a review of the histology by pathologists. Extracted nucleic acid will be quantified by Qubit Fluorometric Quantitation (Thermo Fisher Scientific). Library preparation and targeted capture will be performed using the TruSight Tumor 170 (TST170) kit and samples sequenced on NextSeq 550 (Illumina) using NextSeq V.2.5 hi output reagents, according to the manufacturer's instructions. Data analysis will be performed using the Illumina BaseSpace TST170 app v1.02 and a custom tertiary pipeline, implemented within the Clinical Genomics Workspace software platform from PierianDx (also refer to section 3.2). Primary outcomes for the study will be the number of patients enrolled and the number of patients who complete molecular screening. Secondary outcomes will include the proportion of screened patients enrolled; proportion of patients who complete molecular screening; the turn-around time of molecular screening; and the value of a brain tumour specific multi-disciplinary tumour board, called the molecular tumour advisory panel as measured by the proportion of patients in whom the treatment recommendation was refined compared with the recommendations from the automated bioinformatics platform of the reference laboratory testing., Ethics and Dissemination: The study was approved by the lead Human Research Ethics Committee of the Sydney Local Health District: Protocol No. X19-0383. The study will be conducted in accordance with the principles of the Declaration of Helsinki 2013, guidelines for Good Clinical Practice and the National Health and Medical Research Council National Statement on Ethical Conduct in Human Research (2007, updated 2018 and as amended periodically). Results will be disseminated using a range of media channels including newsletters, social media, scientific conferences and peer-reviewed publications., Trial Registration Number: ACTRN12620000087954; Pre-results., Competing Interests: Competing interests: None declared., (© Author(s) (or their employer(s)) 2021. Re-use permitted under CC BY-NC. No commercial re-use. See rights and permissions. Published by BMJ.)

Abstract: Mesenchymal neoplasms with oncogenic kinase activity due to genomic alterations in receptor tyrosine kinase genes are a morphologically heterogeneous group with a variable biologic potential. A subset of these neoplasms are characterized by immunophenotypic property of dual S100 protein/CD34 expression, histopathological resemblance to lipofibromatosis or peripheral nerve sheath tumors, and often alterations in neurotrophic tropomyosin-related kinase genes. In this article, we present a case of an S100 protein/CD34-positive spindle cell neoplasm harboring a rare BRAF gene rearrangement (KIAA1549-BRAF fusion) and discuss the clinical, histopathological, and molecular variations associated with such neoplasms., Competing Interests: The authors declare no conflicts of interest., (Copyright © 2020 Wolters Kluwer Health, Inc. All rights reserved.)

In this study, we evaluated the predictive value of circulating tumour DNA (ctDNA) to inform therapeutic outcomes in metastatic melanoma patients receiving systemic therapies. We analysed 142 plasma samples from metastatic melanoma patients prior to commencement of systemic therapy: 70 were treated with BRAF/MEK inhibitors and 72 with immunotherapies. Patient-specific droplet digital polymerase chain reaction assays were designed for ctDNA detection. Plasma ctDNA was detected in 56% of patients prior to first-line anti-PD1 and/or anti-CTLA-4 treatment. The detection rate in the immunotherapy cohort was comparably lower than those with BRAF inhibitors (76%, p = 0.0149). Decreasing ctDNA levels within 12 weeks of treatment was strongly concordant with treatment response (Cohen's k = 0.798, p < 0.001) and predictive of longer progression free survival. Notably, a slower kinetic of ctDNA decline was observed in patients treated with immunotherapy compared to those on BRAF/MEK inhibitors. Whole exome sequencing of ctDNA was also conducted in 9 patients commencing anti-PD-1 therapy to derive tumour mutational burden (TMB) and neoepitope load measurements. The results showed a trend of high TMB and neoepitope load in responders compared to non-responders. Overall, our data suggest that changes in ctDNA can serve as an early indicator of outcomes in metastatic melanoma patients treated with systemic therapies and therefore may serve as a tool to guide treatment decisions.

Immunotherapy is an important and established treatment option for patients with advanced melanoma. Initial anti-PD1 trials arbitrarily defined a two-year treatment duration, but a shorter treatment duration may be appropriate. In this study, we retrospectively assessed 70 patients who stopped anti-PD1 therapy in the absence of progressive disease (PD) to determine clinical outcomes. In our cohort, the median time on treatment was 11.8 months. Complete response was attained at time of anti-PD1 discontinuation in 61 (87%). After a median follow up of 34.2 months (range: 2-70.8) post discontinuation, 81% remained disease free. Using ddPCR, we determine the utility of circulating tumour DNA (ctDNA) to predict progressive disease after cessation ( n = 38). There was a significant association between presence of ctDNA at cessation and disease progression ( p = 0.012, Fisher's exact test) and this conferred a negative and positive predictive value of 0.82 (95% CI: 0.645-0.930) and 0.80 (95% CI 0.284-0.995), respectively. Additionally, dichotomised treatment-free survival in patients with or without ctDNA at cessation was significantly longer in the latter group ( p < 0.001, HR: 0.008, 95% CI: 0.001-0.079). Overall, our study confirms that durable disease control can be achieved with cessation of therapy in the absence of disease progression and undetectable ctDNA at cessation was associated with longer treatment-free survival.

Purpose: We evaluated the predictive value of pretreatment ctDNA to inform therapeutic outcomes in patients with metastatic melanoma relative to type and line of treatment., Experimental Design: Plasma circulating tumor DNA (ctDNA) was quantified in 125 samples collected from 110 patients prior to commencing treatment with immune checkpoint inhibitors (ICIs), as first- ( n = 32) or second-line ( n = 27) regimens, or prior to commencing first-line BRAF/MEK inhibitor therapy ( n = 66). An external validation cohort included 128 patients commencing ICI therapies in the first- ( N = 77) or second-line ( N = 51) settings., Results: In the discovery cohort, low ctDNA (≤20 copies/mL) prior to commencing therapy predicted longer progression-free survival (PFS) in patients treated with first-line ICIs [HR, 0.20; 95% confidence interval (CI) 0.07-0.53; P < 0.0001], but not in the second-line setting. An independent cohort validated that ctDNA is predictive of PFS in the first-line setting (HR, 0.42; 95% CI, 0.22-0.83; P = 0.006), but not in the second-line ICI setting. Moreover, ctDNA prior to commencing ICI treatment was not predictive of PFS for patients pretreated with BRAF/MEK inhibitors in either the discovery or validation cohorts. Reduced PFS and overall survival were observed in patients with high ctDNA receiving anti-PD-1 monotherapy, relative to those treated with combination anti-CTLA-4/anti-PD-1 inhibitors., Conclusions: Pretreatment ctDNA is a reliable indicator of patient outcome in the first-line ICI treatment setting, but not in the second-line ICI setting, especially in patients pretreated with BRAF/MEK inhibitors. Preliminary evidence indicated that treatment-naïve patients with high ctDNA may preferentially benefit from combined ICIs., (©2020 American Association for Cancer Research.)