Parthenolide as Cooperating Agent for Anti-Cancer Treatment of Various Malignancies

Primary and acquired resistance of cancer to therapy is often associated with activation of nuclear factor kappa B (NF-κB). Parthenolide (PN) has been shown to inhibit NF-κB signaling and other pro-survival signaling pathways, induce apoptosis and reduce a subpopulation of cancer stem-like cells in several cancers. Multimodal therapies that include PN or its derivatives seem to be promising approaches enhancing sensitivity of cancer cells to therapy and diminishing development of resistance. A number of studies have demonstrated that several drugs with various targets and mechanisms of action can cooperate with PN to eliminate cancer cells or inhibit their proliferation. This review summarizes the current state of knowledge on PN activity and its potential utility as complementary therapy against different cancers

Dissecting Mechanisms of Melanoma Resistance to BRAF and MEK Inhibitors Revealed Genetic and Non-Genetic Patient- and Drug-Specific Alterations and Remarkable Phenotypic Plasticity

The clinical benefit of MAPK pathway inhibition in BRAF-mutant melanoma patients is limited by the development of acquired resistance. Using drug-naïve cell lines derived from tumor specimens, we established a preclinical model of melanoma resistance to vemurafenib or trametinib to provide insight into resistance mechanisms. Dissecting the mechanisms accompanying the development of resistance, we have shown that (i) most of genetic and non-genetic alterations are triggered in a cell line- and/or drug-specific manner; (ii) several changes previously assigned to the development of resistance are induced as the immediate response to the extent measurable at the bulk levels; (iii) reprogramming observed in cross-resistance experiments and growth factor-dependence restricted by the drug presence indicate that phenotypic plasticity of melanoma cells largely contributes to the sustained resistance. Whole-exome sequencing revealed novel genetic alterations, including a frameshift variant of RBMX found exclusively in phospho-AKThigh resistant cell lines. There was no similar pattern of phenotypic alterations among eleven resistant cell lines, including expression/activity of crucial regulators, such as MITF, AXL, SOX, and NGFR, which suggests that patient-to-patient variability is richer and more nuanced than previously described. This diversity should be considered during the development of new strategies to circumvent the acquired resistance to targeted therapies

Natural Compounds' Activity against Cancer Stem-Like or Fast-Cycling Melanoma Cells

<div><p>Background</p><p>Accumulating evidence supports the concept that melanoma is highly heterogeneous and sustained by a small subpopulation of melanoma stem-like cells. Those cells are considered as responsible for tumor resistance to therapies. Moreover, melanoma cells are characterized by their high phenotypic plasticity. Consequently, both melanoma stem-like cells and their more differentiated progeny must be eradicated to achieve durable cure. By reevaluating compounds in heterogeneous melanoma populations, it might be possible to select compounds with activity not only against fast-cycling cells but also against cancer stem-like cells. Natural compounds were the focus of the present study.</p><p>Methods</p><p>We analyzed 120 compounds from The Natural Products Set II to identify compounds active against melanoma populations grown in an anchorage-independent manner and enriched with cells exerting self-renewing capacity. Cell viability, cell cycle arrest, apoptosis, gene expression, clonogenic survival and label-retention were analyzed.</p><p>Findings</p><p>Several compounds efficiently eradicated cells with clonogenic capacity and nanaomycin A, streptonigrin and toyocamycin were effective at 0.1 µM. Other anti-clonogenic but not highly cytotoxic compounds such as bryostatin 1, siomycin A, illudin M, michellamine B and pentoxifylline markedly reduced the frequency of ABCB5 (ATP-binding cassette, sub-family B, member 5)-positive cells. On the contrary, treatment with maytansine and colchicine selected for cells expressing this transporter. Maytansine, streptonigrin, toyocamycin and colchicine, even if highly cytotoxic, left a small subpopulation of slow-dividing cells unaffected. Compounds selected in the present study differentially altered the expression of melanocyte/melanoma specific microphthalmia-associated transcription factor (MITF) and proto-oncogene c-MYC.</p><p>Conclusion</p><p>Selected anti-clonogenic compounds might be further investigated as potential adjuvants targeting melanoma stem-like cells in the combined anti-melanoma therapy, whereas selected cytotoxic but not anti-clonogenic compounds, which increased the frequency of ABCB5-positive cells and remained slow-cycling cells unaffected, might be considered as a tool to enrich cultures with cells exhibiting melanoma stem cell characteristics.</p></div

The summary of natural compound activities at 5 µM.

<p>After initial screening, compounds were grouped based on their activities. A compound was defined as anti-clonogenic when it reduced the percentage of clones formed in soft agar to less than 20% of control treated with vehicle (0.05% DMSO). A compound was named cytostatic when it reduced the viable cell number to less than 50% of control using viable cell number counting, and cytotoxic using the flow cytometry after PI-staining. Numbers corresponding to compounds that accumulated melanoma cells in subG₁ are underlined. Compounds that caused cell cycle arrest are marked in green for G₀/G₁ phase, blue for S phase and red for G₂/M phase. Several compounds (46) were not active in any assay. Few compounds were cytotoxic but not markedly influenced the numbers of colonies formed in agar. Few other compounds exerted their effects only on clonogenic cells or reduced clonogenicity below 20% of control, caused cytostatic/cytostatic effect without inducing substantial cell death. Several compounds were cytostatic and cytotoxic and in addition they efficiently eradicated cells with clonogenic potential. They were in the focus of the further study. See Table S1 in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0090783#pone.0090783.s001" target="_blank">File SI</a> for the names of the compounds corresponding to numbers shown in the Figure.</p

Comparison of cytotoxic and anti-clonogenic activities of highly potent natural compounds from Natural Products Set II.

<p>Comparison of cytotoxic and anti-clonogenic activities of highly potent natural compounds from Natural Products Set II.</p

The compound screening procedure.

<p>First, compounds from The Natural Products Set II were tested in two melanoma cell lines obtained from pathologically distinct specimens, DMBC11 and DMBC12, at a single concentration of 5 µM. Changes in viability (APA assay, volumetric assay, PI staining and subG₁ assay) were measured as short-term effects and changes in clonogenic potential as long-term effects of the tested compounds. All viability tests were compared for potential inconsistencies. Two criteria were used to select compounds for further analysis: compound should reduce cell viability (PI-staining) to ≤ 50% of control and clonogenicity to ≤ 20% of control. Next, the selected compounds were used at lower concentrations for dose-response curves. The most potent compounds were then investigated for their ability to induce apoptosis, to influence the frequency of ABCB5-positive cells and label-retaining slow-cycling cells, and to alter gene expression.</p

Selected natural compounds induced diverse effects on the expression of <i>MITF</i> and <i>c-MYC</i>.

<p>(A) Cell viability was measured for highly cytotoxic compounds to choose a concentration not reducing cell viability below 40% of control after 24 h of treatment. (B) qRT-PCR was used to assess the fold change in the expression of <i>MITF</i> and <i>c-MYC</i> after treatment with selected compounds at indicated concentrations and time of exposure. (*P<0.05; **P<0.01; ***P<0.001).</p