An observational study of dose dense chemotherapy with lipegfilgrastim support in early breast cancer.

PURPOSE: Breast cancer is one of the most prevalent malignant diseases in women. The development of dose dense chemotherapy regimens has improved clinical outcomes but has been associated with increased hematological toxicity. Currently there is a paucity of data on the use of lipegfilgrastim in dose dense AC treatment in early breast cancer. The purpose of this study was to assess the use of lipegfilgrastim in the treatment of early breast cancer and to examine the incidence of treatment-related neutropenia during the dose dense AC phase and subsequent paclitaxel treatment. METHODS: This was a single arm, non-interventional, prospective study. The primary endpoint was to determine the rate of neutropenia defined as ANC of < 1.0 × 109/L, during four cycles of dose dense AC with lipegfilgrastim support. The secondary endpoints were the incidence of febrile neutropenia, (temperature > 38 °C and ANC < 1.0 × 109/L), treatment delays, premature treatment cessation and toxicity. RESULTS: Forty-one participants were included in the study. Of the 160 planned dose dense AC treatments, 157 were administered, and 95% (152/160) of these were given on time. The rate of treatment delay was 5% (95% CI 2.2 to 9.9%) due to infection (4) and mucositis (1). Four (10%) patients developed febrile neutropenia. The most frequently occurring adverse event was grade 1 bone pain. CONCLUSION: Lipegfilgrastim is an effective option in the prophylaxis of chemotherapy-induced neutropenia, and its use in everyday anti-cancer treatment can be considered.

New Roles for Vitamin D Superagonists: From COVID to Cancer.

Vitamin D is a potent steroid hormone that induces widespread changes in gene expression and controls key biological pathways. Here we review pathophysiology of vitamin D with particular reference to COVID-19 and pancreatic cancer. Utility as a therapeutic agent is limited by hypercalcemic effects and attempts to circumvent this problem have used vitamin D superagonists, with increased efficacy and reduced calcemic effect. A further caveat is that vitamin D mediates multiple diverse effects. Some of these (anti-fibrosis) are likely beneficial in patients with COVID-19 and pancreatic cancer, whereas others (reduced immunity), may be beneficial through attenuation of the cytokine storm in patients with advanced COVID-19, but detrimental in pancreatic cancer. Vitamin D superagonists represent an untapped resource for development of effective therapeutic agents. However, to be successful this approach will require agonists with high cell-tissue specificity.

Receptor tyrosine kinases and their activation in melanoma.

Receptor tyrosine kinases (RTKs) and their downstream signalling pathways have long been hypothesized to play key roles in melanoma development. A decade ago, evidence was derived largely from animal models, RTK expression studies and detection of activated RAS isoforms in a small fraction of melanomas. Predictions that overexpression of specific RTKs implied increased kinase activity and that some RTKs would show activating mutations in melanoma were largely untested. However, technological advances including rapid gene sequencing, siRNA methods and phospho-RTK arrays now give a more complete picture. Mutated forms of RTK genes including KIT, ERBB4, the EPH and FGFR families and others are known in melanoma. Additional over- or underexpressed RTKs and also protein tyrosine phosphatases (PTPs) have been reported, and activities measured. Complex interactions between RTKs and PTPs are implicated in the abnormal signalling driving aberrant growth and survival in malignant melanocytes, and indeed in normal melanocytic signalling including the response to ultraviolet radiation. Kinases are considered druggable targets, so characterization of global RTK activity in melanoma should assist the rational development of tyrosine kinase inhibitors for clinical use.

Functional neurons and melanocytes induced from immortal lines of postnatal neural crest-like stem cells.

Stem cells, that is, cells that can both reproduce themselves and differentiate into functional cell types, attract much interest as potential aids to healing and disease therapy. Embryonic neural crest is pluripotent and generates the peripheral nervous system, melanocytes, and some connective tissues. Neural-crest-related stem cells have been reported previously in postnatal skin: committed melanocytic stem cells in the hair follicle, and pluripotent cell types from the hair follicle and papilla that can produce various sets of lineages. Here we describe novel pluripotent neural crest-like stem cells from neonatal mouse epidermis, with different potencies, isolated as 3 independent immortal lines. Using alternative regulatory factors, they could be converted to large numbers of either Schwann precursor cells, pigmented melanocytes, chondrocytes, or functional sensory neurons showing voltage-gated sodium channels. Some of the neurons displayed abundant active TRPV1 and TRPA1 receptors. Such functional neurons have previously been obtained in culture only with difficulty, by explantation. The system was also used to generate comparative gene expression data for the stem cells, melanocytes, and melanoblasts that sufficiently explain the lack of pigment in melanoblasts and provide a rationale for some genes expressed apparently ectopically in melanomas, such as ephrin receptors.

Real-time quantitative reverse transcriptase-polymerase chain reaction analysis of melanoma progression-associated genes.

BACKGROUND: Melanoma is an aggressive disease that spreads quickly and is resistant to most therapeutic agents. In an effort to provide insight into the molecular basis of melanoma progression, the expression of 94 genes in 20 metastatic melanomas using a high-throughput real-time quantitative RT-PCR assay was analysed. MATERIALS AND METHODS: A TaqMan low density array (LDA) was designed containing probes/primers directed towards a cohort of genes previously found to be differentially expressed in an isogenic cell line model of melanoma progression. For each sample, cDNA was prepared and added to the quantitative assay. The resulting data were then analysed for correlations with clinical data. RESULTS: Clustering analysis divided the melanomas into two major subgroups based on gene expression patterns. When analysed individually, several genes were associated with overall survival, depth and type of the primary tumour. CONCLUSION: We have identified a selection of genes linked to melanoma progression and patient outcome.

Multiple markers for melanoma progression regulated by DNA methylation: insights from transcriptomic studies.

The incidence of melanoma is increasing rapidly, with advanced lesions generally failing to respond to conventional chemotherapy. Here, we utilized DNA microarray-based gene expression profiling techniques to identify molecular determinants of melanoma progression within a unique panel of isogenic human melanoma cell lines. When a poorly tumorigenic cell line, derived from an early melanoma, was compared with two increasingly aggressive derivative cell lines, the expression of 66 genes was significantly changed. A similar pattern of differential gene expression was found with an independently derived metastatic cell line. We further examined these melanoma progression-associated genes via use of a tailored TaqMan Low Density Array (LDA), representing the majority of genes within our cohort of interest. Considerable concordance was seen between the transcriptomic profiles determined by DNA microarray and TaqMan LDA approaches. A range of novel markers were identified that correlated here with melanoma progression. Most notable was TSPY, a Y chromosome-specific gene that displayed extensive down-regulation in expression between the parental and derivative cell lines. Examination of a putative CpG island within the TSPY gene demonstrated that this region was hypermethylated in the derivative cell lines, as well as metastatic melanomas from male patients. Moreover, treatment of the derivative cell lines with the DNA methyltransferase inhibitor, 2'-deoxy-5-azacytidine (DAC), restored expression of the TSPY gene to levels comparable with that found in the parental cells. Additional DNA microarray studies uncovered a subset of 13 genes from the above-mentioned 66 gene cohort that displayed re-activation of expression following DAC treatment, including TSPY, CYBA and MT2A. DAC suppressed tumor cell growth in vitro. Moreover, systemic treatment of mice with DAC attenuated growth of melanoma xenografts, with consequent re-expression of TSPY mRNA. Overall, our data support the hypothesis that multiple genes are targeted, either directly or indirectly, by DNA hypermethylation during melanoma progression.

p16(Ink4a) in melanocyte senescence and differentiation.

BACKGROUND: The Ink4a-Arf tumor suppressor locus encodes two growth inhibitors, p16 and Arf, both of which are also implicated as effectors in cellular senescence. Because human germline defects in the INK4A-ARF locus are associated with familial melanoma, melanocytes may have unusual INK4A-ARF functions or controls of cell senescence. Because senescence is believed to be an anticancer mechanism, we investigated the role of Ink4a-Arf and its individual components in melanocyte senescence. METHODS: Melanocytes were cultured from littermate mice with zero, one, or two functional copies of the Ink4a-Arf locus. Senescence was evaluated by cumulative population doubling curves and by the assessment of acidic beta-galactosidase (an indicator of senescence) expression. Pigmentation and cell size were evaluated by spectrophotometry and microscopy. p16 and Arf expression in primary and spontaneously immortalized melanocyte or melanocyte precursor cell lines were evaluated by immunoblotting. Retroviral vectors containing normal p16 and Arf complementary DNAs were used to restore expression of these genes in Ink4a-Arf(-/-) melanocytes. RESULTS: Wild-type melanocytes (i.e., Ink4a-Arf(+/+)) senesced within 4-5 weeks of culture. Ink4a-Arf(-/-) melanocytes did not senesce and readily became immortal. Ink4a-Arf(+/-) melanocytes showed defective senescence. Senescent Ink4a-Arf(+/+) melanocytes were heavily pigmented, but Ink4a-Arf(+/-) and Ink4a-Arf(-/-) melanocytes were less pigmented. All of six spontaneously immortalized melanocyte or melanocyte precursor lines from Ink4a-Arf(+/+) mice lacked p16 protein expression, although most retained Arf protein expression. After restoration of p16 but not Arf expression, Ink4a-Arf(-/-) melanocytes stopped growing, became highly melanized, and expressed acidic beta-galactosidase. By contrast, restoration of Arf but not p16 expression led to cell death without evidence of senescence. CONCLUSION: Normal mouse melanocyte senescence and associated pigmentation require both copies of Ink4a-Arf and appear to depend more on p16 than on Arf function. Mutations of the INK4A-ARF locus may favor tumorigenesis from melanocytes by impairing senescence, cell differentiation, and (where ARF is disrupted) cell death.