Pathological response and tumour bed histopathological features correlate with survival following neoadjuvant immunotherapy in stage III melanoma.

BACKGROUND: Guidelines for pathological evaluation of neoadjuvant specimens and pathological response categories have been developed by the International Neoadjuvant Melanoma Consortium (INMC). As part of the Optimal Neo-adjuvant Combination Scheme of Ipilimumab and Nivolumab (OpACIN-neo) clinical trial of neoadjuvant combination anti-programmed cell death protein 1/anti-cytotoxic T-lymphocyte-associated protein 4 immunotherapy for stage III melanoma, we sought to determine interobserver reproducibility of INMC histopathological assessment principles, identify specific tumour bed histopathological features of immunotherapeutic response that correlated with recurrence and relapse-free survival (RFS) and evaluate proposed INMC pathological response categories for predicting recurrence and RFS. PATIENTS AND METHODS: Clinicopathological characteristics of lymph node dissection specimens of 83 patients enrolled in the OpACIN-neo clinical trial were evaluated. Two methods of assessing histological features of immunotherapeutic response were evaluated: the previously described immune-related pathologic response (irPR) score and our novel immunotherapeutic response score (ITRS). For a subset of cases (n = 29), cellular composition of the tumour bed was analysed by flow cytometry. RESULTS: There was strong interobserver reproducibility in assessment of pathological response (κ = 0.879) and percentage residual viable melanoma (intraclass correlation coefficient = 0.965). The immunotherapeutic response subtype with high fibrosis had the strongest association with lack of recurrence (P = 0.008) and prolonged RFS (P = 0.019). Amongst patients with criteria for pathological non-response (pNR, >50% viable tumour), all who recurred had ≥70% viable melanoma. Higher ITRS and irPR scores correlated with lack of recurrence in the entire cohort (P = 0.002 and P ≤ 0.0001). The number of B lymphocytes was significantly increased in patients with a high fibrosis subtype of treatment response (P = 0.046). CONCLUSIONS: There is strong reproducibility for assessment of pathological response using INMC criteria. Immunotherapeutic response of fibrosis subtype correlated with improved RFS, and may represent a biomarker. Potential B-cell contribution to fibrosis development warrants further study. Reclassification of pNR to a threshold of ≥70% viable melanoma and incorporating additional criteria of <10% fibrosis subtype of response may identify those at highest risk of recurrence, but requires validation.

Histopathological features of complete pathological response predict recurrence-free survival following neoadjuvant targeted therapy for metastatic melanoma.

BACKGROUND: Recent clinical trials demonstrated the safety and efficacy of neoadjuvant dabrafenib and trametinib (DT) among patients with surgically resectable clinical stage III BRAFV600E/K mutant melanoma. Although patients achieving a complete pathological response (pCR) exhibited superior recurrence-free survival (RFS) versus those who did not, 30% of pCR patients relapsed. We sought to identify whether histopathological features of the pathological response further delineated risk of relapse. METHODS: Surgical resection specimens from DT-treated patients in two phase 2 clinical trials were reviewed. Histopathological features, including relative amounts of viable tumour, necrosis, melanosis, and fibrosis (hyalinized or immature/proliferative) were assessed for associations with patient outcomes. RESULTS: Fifty-nine patients underwent surgical resection following neoadjuvant DT. Patients achieving pCR (49%) had longer RFS compared with patients who did not (P = 0.005). Patients whose treated tumour showed any hyalinized fibrosis had longer RFS versus those without (P = 0.014), whereas necrosis (P = 0.012) and/or immature/proliferative fibrosis (P = 0.026) correlated with shorter RFS. Multivariable analyses showed absence of pCR or presence of immature fibrosis independently predicted shorter RFS. Among pCR patients, mature/hyalinized-type fibrosis correlated with improved RFS (P = 0.035). CONCLUSIONS: The extent and composition of the pathological response following neoadjuvant DT in BRAFV600E/K mutant melanoma correlates with RFS, including pCR patients. These findings support the need for detailed histological analysis of specimens collected after neoadjuvant therapy.

Spitzoid melanoma with histopathological features of ALK gene rearrangement exhibiting ALK copy number gain: a novel mechanism of ALK activation in spitzoid neoplasia.

Spitzoid neoplasms pose diagnostic difficulties because their morphology is not consistently predictive of their biological potential. Recent advances in the molecular characterization of these tumours provides a framework by which they can now begin to be categorized. In particular, spitzoid lesions with ALK rearrangement have been specifically associated with a characteristic plexiform growth pattern of intersecting fascicles of amelanotic spindled melanocytes. We report the case of an 87-year-old man with a 3-cm nodule on his mid-upper back comprised of an intradermal proliferation of fusiform amelanotic melanocytes arranged in intersecting fascicles with occasional peritumoral clefts. Immunohistochemical studies demonstrated diffuse, strong expression of SOX10 and S100 by the tumour cells and diffuse, weak-to-moderate cytoplasmic positivity for anaplastic lymphoma kinase (ALK), suggestive of ALK rearrangement. Fluorescence in situ hybridization revealed no ALK rearrangements but instead revealed at least three intact ALK signals in 36% of the tumour cells, confirming ALK copy number gain. To our knowledge, this is the first reported case of a plexiform spitzoid neoplasm exhibiting ALK copy number gain instead of ALK rearrangement. This case suggests that ALK copy number gain is a novel mechanism of ALK activation but with the same characteristic histopathological growth pattern seen among ALK-rearranged spitzoid neoplasms.

Pathological assessment of resection specimens after neoadjuvant therapy for metastatic melanoma.

Background: Clinical trials have recently evaluated safety and efficacy of neoadjuvant therapy among patients with surgically resectable regional melanoma metastases. To capture informative prognostic data connected to pathological response in such trials, it is critical to standardize pathologic assessment and reporting of tumor response after this treatment. Methods: The International Neoadjuvant Melanoma Consortium meetings in 2016 and 2017 assembled pathologists from academic centers to develop consensus guidelines for pathologic examination and reporting of surgical specimens from AJCC (8th edition) stage IIIB/C/D or oligometastatic stage IV melanoma patients treated with neoadjuvant-targeted or immune therapy. Patterns of pathologic response are provided context to inform these guidelines. Results: Based on our collective experience and guided by efforts in well-established neoadjuvant settings like breast cancer, procedures directing handling of pre- and post-neoadjuvant therapy-treated melanoma specimens are provided to facilitate comparison of findings across different trials and centers. Definitions of pathologic response are provided together with guidelines for reporting and quantifying the extent of pathologic response. Finally, the spectrum of histopathologic responses observed following neoadjuvant-targeted and immune-checkpoint therapy is described and illustrated. Conclusions: Standardizing pathologic evaluation of resected melanoma metastases following neoadjuvant-targeted or immune-checkpoint therapy allows more robust stratification of patient outcomes. This includes recognizing the spectrum of histopathologic response patterns to neoadjuvant therapy and a standard approach to grading pathologic responses. Such an approach will facilitate comparison of results across clinical trials and inform ongoing correlative studies into the mechanisms of response and resistance to agents applied in the neoadjuvant setting.

Pruritic arthropod bite-like papules in T-cell large granular lymphocytic leukaemia and chronic myelomonocytic leukaemia.

T-cell large granular lymphocytic leukaemia (T-LGLL) is a clinically indolent mature T-cell neoplasm characterized by a monoclonal population of CD3+ CD8+ cytotoxic T cells, which usually presents as neutropenia, anaemia and thrombocytopenia. Chronic myelomonocytic leukaemia (CMML) is a clonal haematopoietic disorder with features of both a myeloproliferative neoplasm and myelodysplastic syndrome (MDS). Patients with CMML exhibit a persistent peripheral blood monocytosis in addition to myelodysplastic features. Because of the rarity of T-LGLL, its cutaneous manifestations are poorly documented, but include vasculitis, vasculopathy, persistent ulcerations, generalized pruritus and disseminated granuloma annulare. Various types of skin lesions have been observed in patients with CMML and reportedly occur in approximately 10% of cases. We report the extraordinary case of a patient with MDS who developed T-LGLL, and subsequently the MDS progressed to CMML. The patient then developed diffuse arthropod bite-like papules and intractable pruritus.

Use of New Techniques in Addition to IHC Applied to the Diagnosis of Melanocytic Lesions, With Emphasis on CGH, FISH, and Mass Spectrometry.

Melanoma remains one of the most aggressive forms of cutaneous malignancies. While its diagnosis based on histologic parameters is usually straight forward in most cases, distinguishing a melanoma from a melanocytic nevus can be challenging in some instances, especially when there are overlapping clinical and histopathologic features. Occasionally, melanomas can histologically mimic other tumors and even demonstration of melanocytic origin can be challenging. Thus, several ancillary tests may be employed to arrive at the correct diagnosis. The objective of this review is to summarize these tests, including the well-established and commonly used ones such as immunohistochemistry, with specific emphasis on emerging techniques such as comparative genomic hybridization, fluorescence in situ hybridization and imaging mass spectrometry.

Suppression of food intake and growth by amino acids in Drosophila: the role of pumpless, a fat body expressed gene with homology to vertebrate glycine cleavage system.

We have isolated a Drosophila mutant, named pumpless, which is defective in food intake and growth at the larval stage. pumpless larvae can initially feed normally upon hatching. However, during late first instar stage, they fail to pump the food from the pharynx into the esophagus and concurrently begin moving away from the food source. Although pumpless larvae do not feed, they do not show the typical physiologic response of starving animals, such as upregulating genes involved in gluconeogenesis or lipid breakdown. The pumpless gene is expressed specifically in the fat body and encodes a protein with homology to a vertebrate enzyme involved in glycine catabolism. Feeding wild-type larvae high levels of amino acids could phenocopy the feeding and growth defects of pumpless mutants. Our data suggest the existence of an amino acid-dependent signal arising from the fat body that induces cessation of feeding in the larva. This signaling system may also mediate growth transition from larval to the pupal stage during Drosophila development.

Lack of Drosophila cytoskeletal tropomyosin affects head morphogenesis and the accumulation of oskar mRNA required for germ cell formation.

Drosophila encodes five muscle and one cytoskeletal isoform of the actin-binding protein tropomyosin. We have identified a lack-of-function mutation in the cytoskeletal isoform (cTmII). Zygotic mutant embryos show a defect in head morphogenesis, while embryos lacking maternal cTmII are defective in germ cell formation but otherwise give rise to viable adults. oskar mRNA, which is required for both germ cell formation and abdominal segmentation, fails to accumulate at the posterior pole in these embryos. nanos mRNA, however, which is required exclusively for abdominal segmentation, is localized at wild-type levels. These results indicate that head morphogenesis and the accumulation of high levels of oskar mRNA necessary for germ cell formation require tropomyosin-dependent cytoskeleton.

Identification of an isoform of the estrogen receptor messenger RNA lacking exon four and present in the brain.

An isoform of the estrogen receptor messenger RNA (ER-mRNA) was identified in RNA from the brain of lizards and rats. Poly(A)+ RNA from brain and uteri was reverse transcribed using gene-specific primer for the ER. The resulting complementary DNA was amplified in a polymerase chain reaction followed by cloning and sequencing of the amplified products. This isoform lacks exon four and is designated delta 4 ER-mRNA. Although several isoforms of the ER have been described from cancerous cells, to our knowledge, none has been identified previously in the brain. Furthermore, the delta 4 isoform is the only isoform detected in normal tissue. The delta 4 isoform appeared most abundant in RNA from brain tissue, whereas uterine RNA contained only trace amounts of the isoform. Apparently, tissue-specific alternative splicing accounts for these differences in abundance. Because exon four encodes a part of the steroid-binding domain, we predict that the corresponding protein encoded by the isoform will not bind estradiol and may therefore belong to a growing subclass of the steroid/thyroid/vitamin superfamily known as orphan receptors. We predict that the putative delta 4 protein may function as a ligand-independent transcription factor that acts on the same DNA response elements as the conventional ER. The abundance of this isoform in the brain relative to the uterus raises fundamental questions regarding the regulation of estrogen-responsive genes in different tissues.