Duncan Lambie

A potent Chk1 inhibitor is selectively cytotoxic in melanomas with high levels of replicative stress

There are few effective treatments for metastatic melanoma. Checkpoint kinase 1 (Chk1) inhibitors are being trialled for their efficacy in enhancing conventional chemotherapeutic agents, but their effectiveness as single agents is not known. We have examined the effectiveness of two novel Chk1 selective inhibitors, AR323 and AR678, in a panel of melanoma cell lines and normal cell types. We demonstrate that these drugs display single-agent activity, with IC50s in the low nanomolar range. The drugs produce cytotoxic effects in cell lines that are most sensitive to these drugs, whereas normal cells are only sensitive to these drugs at the higher concentrations where they have cytostatic activity. The cytotoxic effect is the consequence of inhibition of S-phase Chk1, which drives cells prematurely from late S phase into an aberrant mitosis and results in either failure of cytokinesis or cell death through an apoptotic mechanism. The sensitivity to the Chk1 inhibitors was correlated with the level of endogenous DNA damage indicating replicative stress. Chk1 inhibitors are viable single-agent therapies that target melanoma cells with high levels of endogenous DNA damage. This sensitivity suggests that Chk1 is a critical component of an adaptation to replicative stress in these cells. It also suggests that markers of DNA damage may be useful in identifying the melanomas and potentially other tumour types that are more likely to be sensitive to Chk1 inhibitors as single agents.

Current trends in the etiology and diagnosis of HPV-related head and neck cancers

Human papilloma virus (HPV) infection is a major risk factor for a distinct subset of head and neck squamous cell carcinoma (HNSCC). The current review summarizes the epidemiology of HNSCC and the disease burden, the infectious cycle of HPV, the roles of viral oncoproteins, E6 and E7, and the downstream cellular events that lead to malignant transformation. Current techniques for the clinical diagnosis of HPV‐associated HNSCC will also be discussed, that is, the detection of HPV DNA, RNA, and the HPV surrogate marker, p16 in tumor tissues, as well as HPV‐specific antibodies in serum. Such methods do not allow for the early detection of HPV‐associated HNSCC and most cases are at an advanced stage upon diagnosis. Novel noninvasive approaches using oral fluid, a clinically relevant biological fluid, allow for the detection of HPV and cellular alterations in infected cells, which may aid in the early detection and HPV‐typing of HNSCC tumors. Noninvasive diagnostic methods will enable early detection and intervention, leading to a significant reduction in mortality and morbidity associated with HNSCC.

Confocal features of equivocal facial lesions on severely sun-damaged skin: Four case studies with dermatoscopic, confocal, and histopathologic correlation

BACKGROUND Facial skin has a distinct histologic architecture and reveals specific dermatoscopic features. Diagnosis of lentigo maligna on the face is often challenging because of the overlap of clinical and morphologic features with other lesions. OBJECTIVES We aim to show the value of reflectance confocal microscopy (RCM) as a noninvasive diagnostic tool for facial lesions and to increase knowledge of RCM morphologic features among the scientific community. METHODS We describe a series of 4 facial lesions on severely sun-damaged skin that was evaluated via RCM immediately after face-to-face examination, followed by shave biopsy for histopathological analysis. RESULTS Lesions included a lentigo maligna, a pigmented seborrheic keratosis, pigmented basal cell carcinoma, and a pigmented actinic keratosis. In the presented cases, RCM enabled an accurate diagnosis. LIMITATIONS The study describes morphologic features on selected cases, but does not test accuracy of RCM criteria. CONCLUSIONS RCM is a useful adjuvant for the accurate and precise diagnosis of equivocal facial lesions.

BRAF mutation status is an independent prognostic factor for resected stage IIIB and IIIC melanoma: Implications for melanoma staging and adjuvant therapy

BACKGROUND 5-year survival for melanoma metastasis to regional lymph nodes (American Joint Committee on Cancer stage III) is <50%. Knowledge of outcomes following therapeutic lymphadenectomy for stage III melanoma related to BRAF status may guide adjuvant use of BRAF/MEK inhibitors along with established and future therapies. AIMS To determine patterns of melanoma recurrence and survival following therapeutic lymph node dissection (TLND) associated with oncogenic mutations. METHODS DNA was obtained from patients who underwent TLND and had ⩾2 positive nodes, largest node >3cm or extracapsular invasion. Mutations were detected using an extended Sequenom MelaCARTA panel. RESULTS Mutations were most commonly detected in BRAF (57/124 [46%] patients) and NRAS (26/124 [21%] patients). Patients with BRAF mutations had higher 3-year recurrence rate (77%) versus 54% for BRAF wild-type patients (hazard ratio (HR) 1.8, p=0.008). The only prognostically significant mutations occurred in BRAF: median recurrence-free (RFS) and disease-specific survival (DSS) for BRAF mutation patients was 7 months and 16 months, versus 19 months and not reached for BRAF wild-type patients, respectively. Multivariate analysis identified BRAF mutant status and number of positive lymph nodes as the only independent prognostic factors for RFS and DSS. CONCLUSIONS Patients with BRAF mutations experienced rapid progression of metastatic disease with locoregional recurrence rarely seen in isolation, supporting incorporation of BRAF status into melanoma staging and use of BRAF/MEK inhibitors post-TLND.

Evidence for Steroidogenic Potential in Human Prostate Cell Lines and Tissues

Malignant prostate cancer (PCa) is usually treated with androgen deprivation therapies (ADTs). Recurrent PCa is resistant to ADT. This research investigated whether PCa can potentially produce androgens de novo, making them androgen self-sufficient. Steroidogenic enzymes required for androgen synthesis from cholesterol (CYP11A1, CYP17A1, HSD3β, HSD17β3) were investigated in human primary PCa (n = 90), lymph node metastases (LNMs; n = 8), and benign prostatic hyperplasia (BPH; n = 6) with the use of IHC. Six prostate cell lines were investigated for mRNA and protein for steroidogenic enzymes and for endogenous synthesis of testosterone and 5α-dihydrotestosterone. All enzymes were identified in PCa, LNMs, BPH, and cell lines. CYP11A1 (rate-limiting enzyme) was expressed in cancerous and noncancerous prostate glands. CYP11A1, CYP17A1, HSD3β, and HSD17β3 were identified, respectively, in 78%, 52%, 16%, and 82% of human BPH and PCa samples. Approximately 10% of primary PCa, LNMs, and BPH expressed all four enzymes simultaneously. CYP11A1 expression was stable, CYP17A1 increased, and HSD3β and HSD17β3 decreased with disease progression. CYP17A1 expression was significantly correlated with CYP11A1 (P = 0.0009), HSD3β (P = 0.0297), and HSD17β3 (P = 0.0090) in vivo, suggesting CYP17A1 has a key role in prostatic steroidogenesis similar to testis and adrenal roles. In vitro, all cell lines expressed mRNA for all enzymes. Protein was not always detectable; however, all cell lines synthesized androgen from cholesterol. The results indicate that monitoring steroidogenic metabolites in patients with PCa may provide useful information for therapy intervention.

Effectiveness and limitations of reflectance confocal microscopy in detecting persistence of basal cell carcinomas: a preliminary study.

Background/Objectives:  Reflectance confocal microscopy (RCM) can accurately and non‐invasively diagnose basal cell carcinoma (BCC). The use of RCM in assessing responses to saucerization or curettage and cautery of BCC has not been established. The aim of the present study was to expound the usefulness of RCM in assessing treatment responses of BCC to saucerization or curettage and cautery 8–12 weeks after treatment.

BRAFV600E Mutation Status of Involuting and Stable Nevi in Dabrafenib Therapy With or Without Trametinib

IMPORTANCE Recent advances in targeting BRAFV600E mutations, which occur in roughly 50% of melanomas and 70% of benign nevi, have improved response rates and survival in patients with melanoma. With increased survival, the importance of other comorbidities increases and requires consideration in long-term management. This case report discusses dynamic dermoscopic nevus changes that occur during dabrafenib therapy and offers some conclusions regarding BRAF mutations and the changes. OBSERVATIONS A man in his 30s had been monitored with whole-body dermoscopy at roughly 7-month intervals as part of a nevus surveillance study. Fourteen months after his initial visit, metastases were found, and the patient entered a clinical trial of dabrafenib with or without trametinib therapy. Continued dermoscopic monitoring for the next 12 months revealed that approximately 50% of the existing acquired melanocytic nevi involuted, while the remaining nevi did not change. Biopsy findings from 1 unchanged and 1 involuted nevus showed BRAF wild type in the unchanged nevus, BRAFV600E mutation in the involuting nevus, and no malignant histopathologic characteristics in either one. CONCLUSIONS AND RELEVANCE Our observations indicate that a previously suggested hypothesis regarding involuting nevi in BRAF inhibitor therapy is correct: Nevi that involute while a patient is undergoing BRAF V600E inhibitor therapy possess the BRAF V600E mutation, while others that grow or remain unchanged are wild type. However larger-scale trials are required to gather conclusive data and create a more complete clinical picture.

Histopathological features of clinical perineural invasion of cutaneous squamous cell carcinoma of the head and neck and the potential implications for treatment.

Nonmelanoma skin cancer (NMSC) with perineural invasion (PNI) is most commonly seen in cutaneous squamous cell carcinoma of the head and neck (SCCHN). The cranial nerves are a conduit for skin cancer to reach the brainstem.

Microscopic colitis with giant cells: a clinico-pathological review of 11 cases and comparison with microscopic colitis without giant cells

Aim: To document clinical and pathological features of microscopic colitis with giant cells (MCGC) which is one of a number of atypical variants of microscopic colitis. Methods: Cases of microscopic colitis were assessed for giant cells during routine reporting and retrieved from the slide file at a private laboratory. The histological features and clinical data were assessed. Histochemistry (trichome and haematoxylin van Gieson) and immunohistochemistry (CD68) was performed to characterise the nature of the giant cells. Results: Giant cells were identified in 11 cases of microscopic colitis. The histological features of MCGC are not significantly different from usual MC except for the presence of multinucleated giant cells in the superficial lamina propria. Apart from the common but not unexpected association with autoimmune disease, no unique clinical features of the MCGC group were identified versus those described in the literature for ordinary MC. Immune disorders included gluten‐sensitive enteropathy, systemic lupus erythematosus and raised titres of antinuclear antibodies. Conclusions: The giant cells have the same immunohistochemical characteristics as histiocytes and appear to form through histiocyte fusion. The presence of giant cells does not appear to confer any further clinical significance and remains a histological curiosity.

A blueprint for staging of murine melanocytic lesions based on the Cdk4 R24C/R24C ::Tyr- NRAS Q 61K model

It has been shown that gene mutations which drive the development of malignant melanoma (MM) in humans also lead to emergence of MM when engineered mice. However, little attention has been paid to the clinical and histopathological features of melanocytic lesions and their natural history in a given mouse model. This knowledge is crucial to enable us to understand how engineered mutations influence the initiation and evolution of melanocytic lesions, and/or for the use of mice as a preclinical model to test specific treatments. We recently reported the development of melanocytic proliferations along the spectrum of naevi to MM in a Cdk4 R24C/R24C ::Tyr‐ NRAS Q 61K mouse model. In this study, we followed the development of lesions over time using digital photography and dermoscopy with the aim to correlate the clinical and histopathological features of lesions developing in this model. We identified two types of lesions. The first are slow‐growing dermal MMs that emanate from dermal naevi. The second did not emanate from naevi, grew rapidly, and appeared to be solely confined to the subcutaneous fat. We present a simple staging system for the MMs that progress from naevi, based on depth of extension into the dermis and subcutis. This represents a blueprint for documentation and follow‐up of MMs in the live animal, which is critical for the proper use of murine melanoma models.