Clinical Impact of Immune Checkpoint Inhibitor (ICI) Response, DNA Damage Repair (DDR) Gene Mutations and Immune-Cell Infiltration in Metastatic Melanoma Subtypes.

Molecular and histopathological analysis of melanoma subtypes has revealed distinct epidemiological, genetic, and clinical features. However, immunotherapy for advanced metastatic melanoma patients does not differ based on subtype. Response to immune checkpoint inhibitors (ICI) has been shown to vary, therefore, predictive biomarkers are needed in the design of precision treatments. Targeted sequencing and histopathological analysis (CD8 and CD20 immunohistochemistry) were performed on subtypes of metastatic melanoma (cutaneous melanoma (CM, n = 10); head and neck melanoma (HNM, n = 7); uveal melanoma (UM, n = 4); acral lentiginous melanoma (AM, n = 1) and mucosal melanoma (MM, n = 1) treated with ICI). Progression-free survival (PFS) was significantly associated with high CD8 expression (p = 0.025) and mutations in DNA damage repair (DDR) pathway genes (p = 0.012) in all subtypes but not with CD20 expression. Our study identified that immune cell infiltration and DDR gene mutations may have an impact in response to ICI treatment in metastatic melanoma but differs among subtypes. Therefore, a comprehensive understanding of the immune infiltration cells' role and DDR gene mutations in metastatic melanoma may identify prognostic biomarkers.

PRDX3 is associated with metastasis and poor survival in uveal melanoma.

AIMS: Uveal melanoma (UM) is the most common primary intraocular malignancy in adults, and 40% develop fatal metastatic disease. Overexpression of thioredoxin-dependent peroxidase reductase (PRDX3) has been implicated in several cancers, including prostate, breast, colorectal and lung cancer. The aim of this study was to compare the immunohistochemical expression of PRDX3 in formalin-fixed, paraffin-embedded (FFPE) primary UM tissues of patients who did and did not develop metastatic disease. METHODS: Immunohistochemical staining of PRDX3 was performed on FFPE tissue microarray samples of 92 primary UM tumours from patients who did and did not develop metastatic disease. The immunohistochemical staining was assessed by two observers who were blinded to all clinicopathological and cytogenetic details including metastatic/non-metastatic information. Based on a scoring system, expression of PRDX3 was graded as high or low. RESULTS: There were 55 tumours (59.8%) from patients who developed metastatic disease, while 37 (40.2%) were from patients who did not develop metastasis. A statistically significant difference in PRDX3 expression was observed in patients who did and did not develop metastasis (p=0.001). A significant positive correlation between high PRDX3 expression and metastasis was observed (p=0.001). A significant negative correlation between PRDX3 expression and survival was found (p=0.005). Kaplan-Meier survival analysis showed a statistically significant difference in overall survival between tumours that demonstrated low and high expression of PRDX3 (67.61 vs 130.64 months, respectively, p=0.013). CONCLUSIONS: High immunohistochemical expression of PRDX3 in primary UM tissue is associated with metastasis and poor survival.

Current and future biomarkers for risk-stratification and treatment personalisation in multiple myeloma.

Multiple myeloma, an incurable malignancy of the plasma cells in the bone marrow, has a complex pathogenesis due to clonal heterogeneity. Over the years, many clinical trials and researches have led to the development of effective myeloma treatments, resulting in survival prolongation. Molecular prognostic markers for risk-stratification to predict survival, and predictive markers for treatment response are being extensively explored. This review discusses the current risk-adaptive strategies based on genetic and molecular risk signatures that are in practice to predict survival and describes the future prognostic and predictive biomarkers across the fields of genomics, proteomics, and glycomics in myeloma. Gene expression profiling and next generation sequencing are coming to the forefront of risk-stratification and therapeutic-response prediction. Similarly, proteomic and glycomic-based platforms are gaining momentum in biomarker discovery to predict drug resistance and disease progression.

Polyclonal Immunoglobulin G N-Glycosylation in the Pathogenesis of Plasma Cell Disorders.

The pathological progression from benign monoclonal gammopathy of undetermined significance (MGUS) to smoldering myeloma (SMM) and finally to active myeloma (MM) is poorly understood. Abnormal immunoglobulin G (IgG) glycosylation in myeloma has been reported. Using a glycomic platform composed of hydrophilic interaction UPLC, exoglycosidase digestions, weak anion-exchange chromatography, and mass spectrometry, polyclonal IgG N-glycosylation profiles from 35 patients [MGUS (n = 8), SMM (n = 5), MM (n = 8), complete-response (CR) post-treatment (n = 5), relapse (n = 4), healthy age-matched control (n = 5)] were characterized to map glycan structures in distinct disease phases of multiple myeloma. N-Glycan profiles from MGUS resembled normal control. The abundance of neutral glycans containing terminal galactose was highest in SMM, while agalactosylated glycans and fucosylated glycans were lowest in MM. Three afucosyl-biantennary-digalactosylated-sialylated species (A2G2S1, A2BG2S1, and A2BG2S2) decreased 2.38-, 2.4-, and 4.25-fold, respectively, from benign to active myeloma. Increased light chain sialylation was observed in a longitudinal case of transformation from MGUS to MM. Bisecting N-acetylglucosamine was lowest in the CR group, while highest in relapsed disease. Gene expression levels of FUT 8, ST6GAL1, B4GALT1, RECK, and BACH2 identified from publicly available GEP data supported the glycomic changes seen in MM compared to control. The observed differential glycosylation underlined the heterogeneity of the myeloma spectrum. This study demonstrates the feasibility of mapping glycan modifications on the IgG molecule and provides proof of principle that differential IgG glycosylation patterns can be successfully identified in plasma cell disorders.

Elevated levels of 14-3-3 proteins, serotonin, gamma enolase and pyruvate kinase identified in clinical samples from patients diagnosed with colorectal cancer.

BACKGROUND: Colorectal cancer (CRC), a heterogeneous disease that is common in both men and women, continues to be one of the predominant cancers worldwide. Lifestyle, diet, environmental factors and gene defects all contribute towards CRC development risk. Therefore, the identification of novel biomarkers to aid in the management of CRC is crucial. The aim of the present study was to identify candidate biomarkers for CRC, and to develop a better understanding of their role in tumourogenesis. METHODS: In this study, both plasma and tissue samples from patients diagnosed with CRC, together with non-malignant and normal controls were examined using mass spectrometry based proteomics and metabolomics approaches. RESULTS: It was established that the level of several biomolecules, including serotonin, gamma enolase, pyruvate kinase and members of the 14-3-3 family of proteins, showed statistically significant changes when comparing malignant versus non-malignant patient samples, with a distinct pattern emerging mirroring cancer cell energy production. CONCLUSION: The diagnosis and management of CRC could be enhanced by the discovery and validation of new candidate biomarkers, as found in this study, aimed at facilitating early detection and/or patient stratification together with providing information on the complex behaviour of cancer cells.

Predictive biomarkers for dasatinib treatment in melanoma.

Dasatinib has anti-proliferative and anti-invasive effects in melanoma cell lines. However clinical trials have shown modest activity for dasatinib in metastatic melanoma. Although dasatinib targets SRC kinase, neither expression nor phosphorylation of SRC appears to predict response to dasatinib. Identification of predictive biomarkers for dasatinib may facilitate selection of melanoma patients who are more likely to respond to dasatinib. We correlated the anti-proliferative effects of dasatinib in 8 melanoma cell lines with expression of a previously identified 6-gene biomarker panel. We examined the relationship between response to dasatinib and expression of each gene at both the mRNA and protein level. Dasatinib inhibited growth in 3 of the 8 cell lines tested. mRNA expression of the panel of 6 biomarkers did not correlate with response, whilst elevated protein expression of ANXA1, CAV-1 and EphA2 correlated significantly with response to dasatinib in the panel of cell lines. Expression of ANXA1, CAV-1 and EphA2 were analysed in 124 melanoma samples by immunohistochemistry. ANXA1 protein was detected in 81 % (97/120) of tumours, CAV-1 in 44 % (54/122) of tumours and EphA2 in 74 % (90/121) of tumours. Thirty one % (35/113) of tumours tested expressed all three markers and 19 % (21/112) had moderate or strong expression of ANXA1, CAV-1 and EphA2. Seventeen percent (19/112) of melanoma samples were positive for SRC kinase expression, combined with high expression of ANXA1, CAV-1 and EphA2. This subgroup may represent a population of melanoma patients who would be more likely to derive clinical benefit from dasatinib treatment.

Potential of skin fibroblasts for application to anterior cruciate ligament tissue engineering.

Fibroblasts isolated from skin and from anterior cruciate ligament (ACL) secrete type I and type III collagens in vivo and in vitro. However, it is much easier and practical to obtain a small skin biopsy than an ACL sample to isolate fibroblasts for tissue engineering applications. Various tissue engineering strategies have been proposed for torn ACL replacement. We report here the results of the implantation of bioengineered ACLs (bACLs), reconstructed in vitro using a type I collagen scaffold, anchored with two porous bone plugs to allow bone-ligament-bone surgical engraftment. The bACLs were seeded with autologous living dermal fibroblasts, and grafted for 6 months in goat knee joints. Histological and ultrastructural observations ex vivo demonstrated a highly organized ligamentous structure, rich in type I collagen fibers and cells. Grafts' vascularization and innervation were observed in all bACLs that were entirely reconstructed in vitro. Organized Sharpey's fibers and fibrocartilage, including chondrocytes, were present at the osseous insertion sites of the grafts. They showed remodeling and matrix synthesis postimplantation. Our tissue engineering approach may eventually provide a new solution to replace torn ACL in humans.

MDR-1, but not MDR-3 gene expression, is associated with unmutated IgVH genes and poor prognosis chromosomal aberrations in chronic lymphocytic leukemia.

Two P-glycoprotein (P-gp) genes, MDR-1 (ABCB1) and MDR-3 (ABCB4), have been identified in humans. This study was designed to investigate whether associations exist between expression of MDR-1 and MDR-3 P-gp and other markers of poor prognosis and/or prior exposure to therapeutic agents in chronic lymphocytic leukemia (CLL). IgVH mutational status, gene usage, CD38 positivity, FISH analysis and clinical information were available on all patients. Twenty-one of 101 patients tested showed MDR-3 P-gp positivity. Associations with markers of poor prognosis or prior chemotherapy did not reach statistical significance, but MDR-3 P-gp positive patients had significantly shorter survivals than MDR-3 P-gp negative patients. MDR-1 P-gp expression (18/25) showed a strong association with unmutated IgVH genes and adverse prognosis cytogenetics (p = 0.015, p = 0.014, respectively), but was independent of prior exposure to chemotherapeutic agents. These results suggest a role for MDR-1 and MDR-3 in chemoresistant disease. This study highlights the value of determining MDR phenotype in CLL patients prior to treatment, to allow the design of novel drug regimens containing agents that reverse MDR function.