Imaging the elemental distribution within human malignant melanomas using Laser-Induced Breakdown Spectroscopy.

The diagnosis of malignant melanoma, often an inconspicuous but highly aggressive tumor, is most commonly done by histological examination, while additional diagnostic methods on the level of elements and molecules are constantly being developed. Several studies confirmed differences in the chemical composition of healthy and tumor tissue. Our study presents the potential of the LIBS (Laser-Induced-Breakdown Spectroscopy) technique as a diagnostic tool in malignant melanoma (MM) based on the quantitative changes in elemental composition in cancerous tissue. Our patient group included 17 samples of various types of malignant melanoma and one sample of healthy skin tissue as a control. To achieve a clear perception of results, we have selected two biogenic elements (calcium and magnesium), which showed a dissimilar distribution in cancerous tissue from its healthy surroundings. Moreover, we observed indications of different concentrations of these elements in different subtypes of malignant melanoma, a hypothesis that requires confirmation in a more extensive sample set. The information provided by the LIBS Imaging method could potentially be helpful not only in the diagnostics of tumor tissue but also be beneficial in broadening the knowledge about the tumor itself.

Identification of BRAF Inhibitor Resistance-associated lncRNAs Using Genome-scale CRISPR-Cas9 Transcriptional Activation Screening.

BACKGROUND/AIM: Approximately 50% of melanomas harbor the BRAF V600E mutation and targeted therapies using BRAF inhibitors improve patient outcomes. Nonetheless, resistance to BRAF inhibitors develops rapidly and remains a challenge in melanoma treatment. In this study, we attempted to isolate long noncoding RNAs (lncRNAs) involved in BRAF inhibitor resistance using a comprehensive screening method. MATERIALS AND METHODS: We used a CRISPR-Cas9 synergistic activation mediator (SAM) protein complex in a genome-scale transcriptional activation assay to screen for candidate lncRNA genes related to BRAF inhibitor resistance. Correlation analysis was performed between expression levels of isolated lncRNA genes and IC50 of dabrafenib in a BRAF-mutated melanoma cell line. Next, online databases were used to construct the lncRNA-miRNA-mRNA regulatory network. Finally, we evaluated the significance of the expression levels of these lncRNAs and mRNAs as biomarkers using clinical specimens. RESULTS: We isolated three BRAF inhibitor resistance-associated lncRNA genes, namely SNHG16, NDUFV2-AS1, and LINC01502. We constructed a lncRNA-miRNA-mRNA network of 13 nodes consisting of three lncRNAs, six miRNAs, and four mRNAs. The lncRNAs and target mRNAs from each regulatory axis significantly and positively correlated with each other. Finally, Kaplan-Meier analysis showed that higher expression levels of MITF, which was up-regulated by LINC01502, were significantly associated with worse prognosis in BRAF V600E-mutated melanoma. CONCLUSION: The identification of these BRAF inhibitor resistance-associated lncRNA genes at the genomic scale and the establishment of the lncRNA-miRNA-mRNA regulatory network provides new insights into the underlying mechanisms of BRAF inhibitor resistance in melanoma.

Metastasis diagnosis using attenuated total reflection-Fourier transform infra-red (ATR-FTIR) spectroscopy.

The suitability of Fourier transform infrared spectroscopy as a metastasis prognostic tool has not been reported for some cancer types. Our main aim was to show spectroscopic differences between live un-preprocessed cancer cells of different metastatic levels. Spectra of four cancer cell pairs, including colon cancer (SW480, SW620); human melanoma (WM115, WM266.4); murine melanoma (B16F01, B16F10); and breast cancer (MCF7, MDA-MB-231); each pair having the same genetic background, but different metastatic level were analyzed in the regions 1400-1700 cm-1 and 3100-3500 cm-1 using Principal Component Analysis, curve fitting, multifractal dimension and receiver operating characteristic (ROC) curves. The results show spectral markers I1540/I1473, I1652/I1473, [Formula: see text], and multifractal dimension of the spectral images are significantly different for the cells based on their metastatic levels. ROC curve analysis showed good diagnostic performance of the spectral markers in separating cells based on metastatic degree, with areas under the ROC curves having 95% confidence interval lower limits greater than 0.5 for most instances. These spectral features can be important in predicting the probability of metastasis in primary tumors, providing useful guidance for treatment planning. Our markers are effective in differentiating metastatic levels without sample fixation or drying and therefore could be compactible for future use in in-vivo procedures involving spectroscopic cancer diagnosis.

Multimodal MALDI imaging mass spectrometry for improved diagnosis of melanoma.

Imaging mass spectrometry (IMS) provides promising avenues to augment histopathological investigation with rich spatio-molecular information. We have previously developed a classification model to differentiate melanoma from nevi lesions based on IMS protein data, a task that is challenging solely by histopathologic evaluation. Most IMS-focused studies collect microscopy in tandem with IMS data, but this microscopy data is generally omitted in downstream data analysis. Microscopy, nevertheless, forms the basis for traditional histopathology and thus contains invaluable morphological information. In this work, we developed a multimodal classification pipeline that uses deep learning, in the form of a pre-trained artificial neural network, to extract the meaningful morphological features from histopathological images, and combine it with the IMS data. To test whether this deep learning-based classification strategy can improve on our previous results in classification of melanocytic neoplasia, we utilized MALDI IMS data with collected serial H&E stained sections for 331 patients, and compared this multimodal classification pipeline to classifiers using either exclusively microscopy or IMS data. The multimodal pipeline achieved the best performance, with ROC-AUCs of 0.968 vs. 0.938 vs. 0.931 for the multimodal, unimodal microscopy and unimodal IMS pipelines respectively. Due to the use of a pre-trained network to perform the morphological feature extraction, this pipeline does not require any training on large amounts of microscopy data. As such, this framework can be readily applied to improve classification performance in other experimental settings where microscopy data is acquired in tandem with IMS experiments.

[Swelling on the neck]. / Schwellung am Hals.

INTRODUCTION: A 30-year-old female patient presented with a swelling of a cervical left lymph node measuring 1x3 cm, which had been presenting for three weeks. Lymph node excision revealed a metastasis of a malignant melanoma, but the primary tumor was not found. The guidelines recommend neck dissection and adjuvant systemic or immunotherapy. The patient opted for immunotherapy with pembrolizumab and was tumor-free one year later.

Glycyrrhizic acid inhibits DNA damage repair and enhances cisplatin-induced apoptosis of melanoma cells.

This research was designed to prospect the mechanism and impact of glycyrrhizic acid (GA) on DNA damage repair and cisplatin (CP)-induced apoptosis of melanoma cells. First, human melanoma cell SK-MEL-28 was stimulated using GA for 24, 48, and 72 h. Then, the optimal treatment time and dosage were selected. After that, cell counting kit-8 (CCK-8) was employed for testing the cell viability, flow cytometry for the apoptosis, comet assay for the DNA damage of cells, and western blot for the cleaved-Caspase3, Caspase3, Bcl-2, and γH2AX protein expression levels. The experimental outcomes exhibited that as the GA concentration climbed up, the SK-MEL-28 cell viability dropped largely, while the apoptosis level raised significantly, especially at the concentration of 100 µm. In addition, compared with GA or CPtreatment only, CP combined with GA notably suppressed the viability of melanoma cells and promoted cell apoptosis at the cytological level. At the protein level, the combined treatment notably downregulated the Bcl-2 and Caspase3 expression levels, while significantly upregulated the cleaved-Caspase3 and γH2AX expression levels. Besides, CP + GA treatment promoted DNA damage at the DNA molecular level. Collectively, both GA and CP can inhibit DNA damage repair and enhance the apoptosis of SK-MEL-28 cells, and the synergistic treatment of both exhibits better efficacy.

Influence of Anatomic Locations of T1b Melanomas and Patient Age on Outcomes.

BACKGROUND: There are no guidelines on when to more strongly recommend sentinel lymph node biopsy (SLNB) for T1b melanomas. OBJECTIVE: To examine whether anatomic locations of T1b melanomas and patient age influence metastases. METHODS: We conducted a retrospective study using data from two hospitals in Los Angeles County from January 2010 through January 2020. RESULTS: Out of 620 patients with primary melanomas, 566 melanomas were staged based on the American Joint Committee on Cancer 8th edition melanoma staging. Forty-one were T1b, of which 13 were located on the face/ear/scalp and 28 were located elsewhere. T1b melanomas located on the face/ear/scalp had an increased risk of lymph node or distant metastasis compared with other anatomic sites (31% vs 3.6%, P=0.028). For all melanomas, the risk of lymph node or distant metastasis decreased with age of 64 years or greater (P<0.001 and P=0.034). For T1b melanomas, the risk of distant metastasis increased with increasing age (P=0.047). LIMITATIONS: Data were from a single county.  Conclusion: T1b melanomas of the face/ear/scalp demonstrated a higher risk of lymph node or distant metastasis and may help guide the recommendation of SLNB, imaging, and surveillance. Younger patients may be more strongly considered for SLNB and older patients with T1b melanomas may warrant imaging.  J Drugs Dermatol. 2024;23(5):306-310. doi:10.36849/JDD.7667.

Copy number variations in malignant melanoma: genomic regions, biomarkers, and therapeutic targets.

Malignant melanoma is a skin tumor arising from melanocytes, occurring mostly in predisposed individuals. Melanomas are frequently present with copy number variations (CNVs), i.e., gains or losses of specific DNA regions that have provided immense potential for disease diagnosis and classification. The methodology of CNV detection has revolutionized in past decades, and current high throughput technologies enable us to analyze the entire spectrum of CNV alterations at the whole genome scale. Thus, identifying novel CNV biomarkers and evaluating their applicability in biomedicine are becoming increasingly important. The aim of this review was to summarize copy number changes occurring in malignant melanomas. We made an overview of specific genes and chromosomal locations affected in sporadic and familial melanoma and also of known germline alterations in melanoma-prone families. We summarized genomic regions aberrant in malignant melanoma and highlighted those frequently discussed in the literature, suggesting 7q, 11q, 12q, 9p, and 1q, but also others, as the most affected ones.

CD133 Stimulates Cell Proliferation via the Upregulation of Amphiregulin in Melanoma.

CD133, a cancer stem cell (CSC) marker in tumors, including melanoma, is associated with tumor recurrence, chemoresistance, and metastasis. Patient-derived melanoma cell lines were transduced with a Tet-on vector expressing CD133, generating doxycycline (Dox)-inducible cell lines. Cells were exposed to Dox for 24 h to induce CD133 expression, followed by RNA-seq and bioinformatic analyses, revealing genes and pathways that are significantly up- or downregulated by CD133. The most significantly upregulated gene after CD133 was amphiregulin (AREG), validated by qRT-PCR and immunoblot analyses. Induced CD133 expression significantly increased cell growth, percentage of cells in S-phase, BrdU incorporation into nascent DNA, and PCNA levels, indicating that CD133 stimulates cell proliferation. CD133 induction also activated EGFR and the MAPK pathway. Potential mechanisms highlighting the role(s) of CD133 and AREG in melanoma CSC were further delineated using AREG/EGFR inhibitors or siRNA knockdown of AREG mRNA. Treatment with the EGFR inhibitor gefitinib blocked CD133-induced cell growth increase and MAPK pathway activation. Importantly, siRNA knockdown of AREG reversed the stimulatory effects of CD133 on cell growth, indicating that AREG mediates the effects of CD133 on cell proliferation, thus serving as an attractive target for novel combinatorial therapeutics in melanoma and cancers with overexpression of both CD133 and AREG.

Comprehensive analysis of clinical images contributions for melanoma classification using convolutional neural networks.

BACKGROUND: Timely diagnosis plays a critical role in determining melanoma prognosis, prompting the development of deep learning models to aid clinicians. Questions persist regarding the efficacy of clinical images alone or in conjunction with dermoscopy images for model training. This study aims to compare the classification performance for melanoma of three types of CNN models: those trained on clinical images, dermoscopy images, and a combination of paired clinical and dermoscopy images from the same lesion. MATERIALS AND METHODS: We divided 914 image pairs into training, validation, and test sets. Models were built using pre-trained Inception-ResNetV2 convolutional layers for feature extraction, followed by binary classification. Training comprised 20 models per CNN type using sets of random hyperparameters. Best models were chosen based on validation AUC-ROC. RESULTS: Significant AUC-ROC differences were found between clinical versus dermoscopy models (0.661 vs. 0.869, p < 0.001) and clinical versus clinical + dermoscopy models (0.661 vs. 0.822, p = 0.001). Significant sensitivity differences were found between clinical and dermoscopy models (0.513 vs. 0.799, p = 0.01), dermoscopy versus clinical + dermoscopy models (0.799 vs. 1.000, p = 0.02), and clinical versus clinical + dermoscopy models (0.513 vs. 1.000, p < 0.001). Significant specificity differences were found between dermoscopy versus clinical + dermoscopy models (0.800 vs. 0.288, p < 0.001) and clinical versus clinical + dermoscopy models (0.650 vs. 0.288, p < 0.001). CONCLUSION: CNN models trained on dermoscopy images outperformed those relying solely on clinical images under our study conditions. The potential advantages of incorporating paired clinical and dermoscopy images for CNN-based melanoma classification appear less clear based on our findings.

Preferences of physicians for treatment-related toxicity vs. recurrence in melanoma (GERMELATOX-A): the doctors' perspective.

INTRODUCTION: Adjuvant treatment with immune checkpoint inhibitors, such as PD1-antibodies (ICI) ± CTLA4-antibodies (cICI) or targeted therapy with BRAF/MEK inhibitors (TT), has shown a significant improvement in disease-free survival (DFS) for high-risk melanoma patients. However, due to specific side effects, the choice of treatment is often influenced by the risk of toxicity. Therefore, the role of physicians in treatment decisions of patients is crucial. This study investigated for the first time in a multicenter setting the attitudes and preferences of dermatooncologists in Germany and Switzerland regarding adjuvant treatment with (c)ICI and TT. METHODS: In the GERMELATOX-A study, 108 physicians (median age: 32 yrs, 67.6% female) from 11 skin cancer centers were surveyed to rate typical side effect scenarios of (c)ICI and TT treatments and then compared to patients' ratings evaluated in a previous analysis from the same centers. The scenarios described mild-to-moderate or severe toxicity and included melanoma relapse leading to death. The physicians were asked about the level of side effects they would tolerate in exchange for a reduction in melanoma relapse and an increase in survival at 5 years. RESULTS: The preferences of physicians and patients revealed significant differences regarding adjuvant melanoma treatment with (c)ICI and TT (p < 0.05). Compared to patients, physicians tend to value a melanoma relapse less severe, according to a visual analog scale. They were also less threatened by all scenarios of side effects during adjuvant treatment with (c)ICI or TT, compared to patients. Physicians required lower risk reductions for disease-free survival (DFS) and overall survival (OS) for both ICI and TT and their drug-related side effects to accept these treatments. In case of severe side effects, physicians required similar 5-year DFS rates for ICI and TT (60-65%), while patients needed a 15% improvement of 5-year DFS for ICI compared to TT (80%/65%). For survival, physicians expected an OS improvement of + 10% for all three treatment modalities, whereas patients required a higher increase: + 18-22% for ICI and + 15% for TT. CONCLUSION: Our study highlights the importance of understanding the patient's perspective and a potential difference to the doctor's view when making decisions about adjuvant melanoma treatment with (c)ICI and TT, especially as these treatments are increasingly being implemented in earlier stages.

SEnSCA: Identifying possible ligand-receptor interactions and its application in cell-cell communication inference.

Multicellular organisms have dense affinity with the coordination of cellular activities, which severely depend on communication across diverse cell types. Cell-cell communication (CCC) is often mediated via ligand-receptor interactions (LRIs). Existing CCC inference methods are limited to known LRIs. To address this problem, we developed a comprehensive CCC analysis tool SEnSCA by integrating single cell RNA sequencing and proteome data. SEnSCA mainly contains potential LRI acquisition and CCC strength evaluation. For acquiring potential LRIs, it first extracts LRI features and reduces the feature dimension, subsequently constructs negative LRI samples through K-means clustering, finally acquires potential LRIs based on Stacking ensemble comprising support vector machine, 1D-convolutional neural networks and multi-head attention mechanism. During CCC strength evaluation, SEnSCA conducts LRI filtering and then infers CCC by combining the three-point estimation approach and single cell RNA sequencing data. SEnSCA computed better precision, recall, accuracy, F1 score, AUC and AUPR under most of conditions when predicting possible LRIs. To better illustrate the inferred CCC network, SEnSCA provided three visualization options: heatmap, bubble diagram and network diagram. Its application on human melanoma tissue demonstrated its reliability in CCC detection. In summary, SEnSCA offers a useful CCC inference tool and is freely available at https://github.com/plhhnu/SEnSCA.

A lactate-SREBP2 signaling axis drives tolerogenic dendritic cell maturation and promotes cancer progression.

Conventional dendritic cells (DCs) are essential mediators of antitumor immunity. As a result, cancers have developed poorly understood mechanisms to render DCs dysfunctional within the tumor microenvironment (TME). After identification of CD63 as a specific surface marker, we demonstrate that mature regulatory DCs (mregDCs) migrate to tumor-draining lymph node tissues and suppress DC antigen cross-presentation in trans while promoting T helper 2 and regulatory T cell differentiation. Transcriptional and metabolic studies showed that mregDC functionality is dependent on the mevalonate biosynthetic pathway and its master transcription factor, SREBP2. We found that melanoma-derived lactate activates SREBP2 in tumor DCs and drives conventional DC transformation into mregDCs via homeostatic or tolerogenic maturation. DC-specific genetic silencing and pharmacologic inhibition of SREBP2 promoted antitumor CD8+ T cell activation and suppressed melanoma progression. CD63+ mregDCs were found to reside within the lymph nodes of several preclinical tumor models and in the sentinel lymph nodes of patients with melanoma. Collectively, this work suggests that a tumor lactate-stimulated SREBP2-dependent program promotes CD63+ mregDC development and function while serving as a promising therapeutic target for overcoming immune tolerance in the TME.

Heterogeneity and molecular landscape of melanoma: implications for targeted therapy.

Uveal cancer (UM) offers a complex molecular landscape characterized by substantial heterogeneity, both on the genetic and epigenetic levels. This heterogeneity plays a critical position in shaping the behavior and response to therapy for this uncommon ocular malignancy. Targeted treatments with gene-specific therapeutic molecules may prove useful in overcoming radiation resistance, however, the diverse molecular makeups of UM call for a patient-specific approach in therapy procedures. We need to understand the intricate molecular landscape of UM to develop targeted treatments customized to each patient's specific genetic mutations. One of the promising approaches is using liquid biopsies, such as circulating tumor cells (CTCs) and circulating tumor DNA (ctDNA), for detecting and monitoring the disease at the early stages. These non-invasive methods can help us identify the most effective treatment strategies for each patient. Single-cellular is a brand-new analysis platform that gives treasured insights into diagnosis, prognosis, and remedy. The incorporation of this data with known clinical and genomics information will give a better understanding of the complicated molecular mechanisms that UM diseases exploit. In this review, we focused on the heterogeneity and molecular panorama of UM, and to achieve this goal, the authors conducted an exhaustive literature evaluation spanning 1998 to 2023, using keywords like "uveal melanoma, "heterogeneity". "Targeted therapies"," "CTCs," and "single-cellular analysis".

Response of treatment-naive brain metastases to stereotactic radiosurgery.

With improvements in survival for patients with metastatic cancer, long-term local control of brain metastases has become an increasingly important clinical priority. While consensus guidelines recommend surgery followed by stereotactic radiosurgery (SRS) for lesions >3 cm, smaller lesions (≤3 cm) treated with SRS alone elicit variable responses. To determine factors influencing this variable response to SRS, we analyzed outcomes of brain metastases ≤3 cm diameter in patients with no prior systemic therapy treated with frame-based single-fraction SRS. Following SRS, 259 out of 1733 (15%) treated lesions demonstrated MRI findings concerning for local treatment failure (LTF), of which 202 /1733 (12%) demonstrated LTF and 54/1733 (3%) had an adverse radiation effect. Multivariate analysis demonstrated tumor size (>1.5 cm) and melanoma histology were associated with higher LTF rates. Our results demonstrate that brain metastases ≤3 cm are not uniformly responsive to SRS and suggest that prospective studies to evaluate the effect of SRS alone or in combination with surgery on brain metastases ≤3 cm matched by tumor size and histology are warranted. These studies will help establish multi-disciplinary treatment guidelines that improve local control while minimizing radiation necrosis during treatment of brain metastasis ≤3 cm.

[Dramatic response to encorafenib plus binimetinib in a patient with leukemoid reaction from metastatic melanoma.] / Melanoma metastatico e reazione leucemoide: il ruolo della combo target.

Paraneoplastic leukemoid reaction (PLR) is an extremely rare condition in patients with melanoma and it is frequently associated with poor prognosis. BRAF gene mutational analysis represents the gold standard in patients with inoperable or metastatic melanoma as the possible presence of target mutations allows the use of the combination treatment with BRAF and MEK inhibitors. In this article, the case of a young woman with BRAF V600E mutated metastatic melanoma associated with PLR who received encorafenib and binimetinib is presented and discussed, with a focus on the relevant treatment response.

[Advanced melanoma with low-burden and low-risk disease: a clinical case.] / Melanoma avanzato con malattia a basso carico e a basso rischio: un caso clinico.

The management of patients with advanced melanoma is becoming more complex than in past years, due to the recent innovations in the therapeutic scenario. New treatment options, such as the immunotherapeutic combination of ipilimumab and nivolumab, and new BRAF and MEK inhibitors recently introduced, namely encorafenib and binimetinib, were recently approved by the Italian regulatory entities, enriching the systemic therapy armamentarium. In this context, tailoring the therapeutic strategy basing on the patient's characteristics is even more crucial to improve the clinical outcome. Patients with low tumor burden and low-risk disease (i.e., with normal LDH levels), having higher survival rates and probability of good outcome to systemic therapy, represent a challenge for the clinician, deserving the attempt to reach long-term complete responses. The following case report is quite representative of the above-mentioned clinical situation.