Evaluation of anticancer activity of novel platinum(II) bis(thiosemicarbazone) complex against breast cancer.

The study aims to synthesize a novel bis(thiosemicarbazone) derivative based on platinum (thioPt) and evaluate its anticancer properties against MFC-7 and MDA-MB-231 breast cancer cells. A new platinum complex was synthesised by reacting K2PtCl4 with 2,2'-(1,2-diphenylethane-1,2-diylidene)bis(hydrazine-1-carbothioamide) in ethanol in the presence of K2CO3. In the obtained complex, the platinum atom is coordinated by a conjugated system = N-NC-S-The structures of the new compound were characterised using NMR spectroscopy, HR MS, IR, and X-ray structural analysis. The obtained results of the cytotoxicity assay indicate that compound thioPt had potent anticancer activity (MCF-7: 61.03 ± 3.57 µM, MDA-MB-231: 60.05 ± 5.40 µM) with less toxicity against normal MCF-10A breast epithelial cells, even compared to the reference compound (cisplatin). In addition, subsequent experiments found that thioPt induces apoptosis through both an extrinsic (↑caspase 8 activity) and intrinsic (↓ΔΨm) pathway, which ultimately leads to an increase in active caspase 3/7 levels. The induction of autophagy and levels of proteins involved in this process (LC3A/B and Beclin-1) were examined in MCF-7 and MDA-MB-231 breast cancer cells exposed to tested compounds (thio, thioPt, cisPt) at a concentration of 50 µM for 24 h. Based on these results, it can be concluded that thio and thioPt do not significantly affect the autophagy process. This demonstrates their superiority over cisplatin, which can stimulate cancer cell survival through its effect on stimulation of autophagy.

The activity of pyrazolo[4,3-e][1,2,4]triazine and pyrazolo[4,3-e]tetrazolo[1,5-b][1,2,4]triazine sulphonamide derivatives in monolayer and spheroid breast cancer cell cultures.

In the last decade, an increasing interest in compounds containing pyrazolo[4,3-e][1,2,4]triazine moiety is observed. Therefore, the aim of the research was to synthesise a novel sulphonyl pyrazolo[4,3-e][1,2,4]triazines (2a, 2b) and pyrazolo[4,3-e]tetrazolo[1,5-b][1,2,4]triazine sulphonamide derivatives (3a, 3b) to assess their anticancer activity. The MTT assay showed that 2a, 2b, 3a, 3b have stronger cytotoxic activity than cisplatin in both breast cancer cells (MCF-7 and MDA-MB-231) and exhibited weaker effect on normal breast cells (MCF-10A). The obtained results showed that the most active compound 3b increased apoptosis via caspase 9, caspase 8, and caspase 3/7. It is worth to note that compound 3b suppressed NF-κB expression and promoted p53, Bax, and ROS which play important role in activation of apoptosis. Moreover, our results confirmed that compound 3b triggers autophagy through increased formation of autophagosomes, expression of beclin-1 and mTOR inhibition. Thus, our study defines a possible mechanism underlying 3b-induced anti-cancer activity against breast cancer cell lines.

Short Communication: Novel Di- and Triselenoesters as Effective Therapeutic Agents Inhibiting Multidrug Resistance Proteins in Breast Cancer Cells.

Breast cancer has the highest incidence rate among all malignancies worldwide. Its high mortality is mainly related to the occurrence of multidrug resistance, which significantly limits therapeutic options. In this regard, there is an urgent need to develop compounds that would overcome this phenomenon. There are few reports in the literature that selenium compounds can modulate the activity of P-glycoprotein (MDR1). Therefore, we performed in silico studies and evaluated the effects of the novel selenoesters EDAG-1 and EDAG-8 on BCRP, MDR1, and MRP1 resistance proteins in MCF-7 and MDA-MB-231 breast cancer cells. The cytometric analysis showed that the tested compounds (especially EDAG-8) are inhibitors of BCRP, MDR1, and MRP1 efflux pumps (more potent than the reference compounds-novobiocin, verapamil, and MK-571). An in silico study correlates with these results, suggesting that the compound with the lowest binding energy to these transporters (EDAG-8) has a more favorable spatial structure affecting its anticancer activity, making it a promising candidate in the development of a novel anticancer agent for future breast cancer therapy.

Di- and Triselenoesters-Promising Drug Candidates for the Future Therapy of Triple-Negative Breast Cancer.

Breast cancer is a major malignancy among women, characterized by a high mortality rate. The available literature evidence indicates that selenium, as a trace element, has chemopreventive properties against many types of cancer; as such, compounds containing it in their structure may potentially exhibit anticancer activity. Accordingly, we have undertaken a study to evaluate the effects of novel selenoesters (EDAG-1, -7, -8, -10) on MCF-7 and MDA-MB-231 breast cancer cells. Our analysis included investigations of cell proliferation and viability as well as cytometric determinations of apoptosis/autophagy induction, changes in mitochondrial membrane polarity (ΔΨm), caspase 3/7, 8, and 9 activities, and Bax, Bcl-2, p53, Akt, AMPK, and LC3A/B proteins. The obtained data revealed that the tested derivatives are highly cytotoxic and inhibit cell proliferation even at nanomolar doses (0.41-0.79 µM). Importantly, their strong proapoptotic properties (↑ caspase 3/7) are attributable to the effects on both the extrinsic (↑ caspase 8) and intrinsic (↓ ΔΨm and Bcl-2, ↑ Bax, p53, and caspase 9) pathways of apoptosis. Moreover, the tested compounds are autophagy activators (↓ Akt, ↑ autophagosomes and autolysosomes, AMPK, LC3A/B). In summary, the potent anticancer activity suggests that the tested compounds may be promising drug candidates for future breast cancer therapy.

Recent Development of Fluoroquinolone Derivatives as Anticancer Agents.

Cancer is the second leading cause of death in the world following cardiovascular disease. Its treatment, including radiation therapy and surgical removal of the tumour, is based on pharmacotherapy, which prompts a constant search for new and more effective drugs. There are high costs associated with designing, synthesising, and marketing new substances. Drug repositioning is an attractive solution. Fluoroquinolones make up a group of synthetic antibiotics with a broad spectrum of activity in bacterial diseases. Moreover, those compounds are of particular interest to researchers as a result of reports of their antiproliferative effects on the cells of the most lethal cancers. This article presents the current progress in the development of new fluoroquinolone derivatives with potential anticancer and cytotoxic activity, as well as structure-activity relationships, along with possible directions for further development.

Thiosemicarbazide Derivatives Targeting Human TopoIIα and IDO-1 as Small-Molecule Drug Candidates for Breast Cancer Treatment.

In 2020, breast cancer became the most frequently diagnosed type of cancer, with nearly 2.3 million new cases diagnosed. However, with early diagnosis and proper treatment, breast cancer has a good prognosis. Here, we investigated the effect of thiosemicarbazide derivatives, previously identified as dual inhibitors targeting topoisomerase IIα and indoleamine-2,3-dioxygenase 1 (IDO 1), on two distinct types of breast cancer cells (MCF-7 and MDA-MB-231). The investigated compounds (1-3) selectively suppressed the growth of breast cancer cells and promoted apoptosis via caspase-8- and caspase-9-related pathways. Moreover, these compounds caused S-phase cell cycle arrest and dose-dependently inhibited the activity of ATP-binding cassette transporters (MDR1, MRP1/2 and BCRP) in MCF-7 and MDA-MB-231 cells. Additionally, following incubation with compound 1, an increased number of autophagic cells within both types of the investigated breast cancer cells was observed. During preliminary testing of ADME-Tox properties, the possible hemolytic activities of compounds 1-3 and their effects on specific cytochrome P450 enzymes were evaluated.

Tamoxifen Modulates the Immune Landscape of the Tumour Microenvironment: The Paired Siglec-5/14 Checkpoint in Anti-Tumour Immunity in an In Vitro Model of Breast Cancer.

Since the role of sialome-Siglec axis has been described as a regulatory checkpoint of immune homeostasis, the promotion of stimulatory or inhibitory Siglec-related mechanisms is crucial in cancer progression and therapy. Here, we investigated the effect of tamoxifen on the sialic acid-Siglec interplay and its significance in immune conversion in breast cancer. To mimic the tumour microenvironment, we used oestrogen-dependent or oestrogen-independent breast cancer cells/THP-1 monocytes transwell co-cultures exposed to tamoxifen and/or ß-estradiol. We found changes in the cytokine profiles accompanied by immune phenotype switching, as measured by the expression of arginase-1. The immunomodulatory effects of tamoxifen in THP-1 cells occurred with the altered SIGLEC5 and SIGLEC14 genes and the expression of their products, as confirmed by RT-PCR and flow cytometry. Additionally, exposure to tamoxifen increased the binding of Siglec-5 and Siglec-14 fusion proteins to breast cancer cells; however, these effects appeared to be unassociated with oestrogen dependency. Our results suggest that tamoxifen-induced alterations in the immune activity of breast cancer reflect a crosstalk between the Siglec-expressing cells and the tumour's sialome. Given the distribution of Siglec-5/14, the expression profile of inhibitory and activatory Siglecs in breast cancer patients may be useful in the verification of therapeutic strategies and predicting the tumour's behaviour and the patient's overall survival.

Multi-Targeting Anticancer Activity of a New 4-Thiazolidinone Derivative with Anti-HER2 Antibodies in Human AGS Gastric Cancer Cells.

Combining chemotherapy with immunotherapy still remains a regimen in anticancer therapy. Novel 4-thiazolidinone-bearing hybrid molecules possess well-documented anticancer activity, and together with anti-HER2 antibodies, may represent a promising strategy in treating patients with gastric cancer with confirmed human epidermal growth factor receptor 2 (HER2) expression. The aim of the study was to synthesize a new 4-thiazolidinone derivative (Les-4367) and investigate its molecular mechanism of action in combination with trastuzumab or pertuzumab in human AGS gastric cancer cells. AGS cell viability and antiproliferative potential were examined. The effect of the tested combinations as well as monotherapy on apoptosis and autophagy was also determined. Metalloproteinase-2 (MMP-2), intercellular adhesion molecule 1 (ICAM-1), pro-inflammatory and anti-inflammatory cytokine concentrations were also demonstrated by the ELISA technique. We proved that pertuzumab and trastuzumab were very effective in increasing the sensitivity of AGS gastric cancer cells to novel Les-4367. The molecular mechanism of action of the tested combination is connected with the induction of apoptosis. Additionally, the anticancer activity is not associated with the autophagy process. Decreased concentrations of pro-inflammatory cytokines, MMP-2 and ICAM-1-were observed. The novel combination of drugs based on anti-HER2 antibodies with Les-4367 is a promising strategy against AGS gastric cancer cells.

An Overview of the Importance of Transition-Metal Nanoparticles in Cancer Research.

Several authorities have implied that nanotechnology has a significant future in the development of advanced cancer therapies. Nanotechnology makes it possible to simultaneously administer drug combinations and engage the immune system to fight cancer. Nanoparticles can locate metastases in different organs and deliver medications to them. Using them allows for the effective reduction of tumors with minimal toxicity to healthy tissue. Transition-metal nanoparticles, through Fenton-type or Haber-Weiss-type reactions, generate reactive oxygen species. Through oxidative stress, the particles induce cell death via different pathways. The main limitation of the particles is their toxicity. Certain factors can control toxicity, such as route of administration, size, aggregation state, surface functionalization, or oxidation state. In this review, we attempt to discuss the effects and toxicity of transition-metal nanoparticles.

Chitosan-Enriched Solution Blow Spun Poly(Ethylene Oxide) Nanofibers with Poly(Dimethylsiloxane) Hydrophobic Outer Layer for Skin Healing and Regeneration.

Chitosan (CS)/poly(ethylene oxide) (PEO)-based nanofiber mats have attracted particular attention as advanced materials for medical and pharmaceutical applications. In the scope of present studies, solution blow spinning was applied to produce nanofibers from PEO and CS and physicochemical and biopharmaceutical studies were carried out to investigate their potential as wound nanomaterial for skin healing and regeneration. Additional coating with hydrophobic poly(dimethylsiloxane) was applied to favor removal of nanofibers from the wound surface. Unmodified nanofibers displayed highly porous structure with the presence of uniform, randomly aligned nanofibers, in contrast to coated materials in which almost all the free spaces were filled in with poly(dimethylsiloxane). Infrared spectroscopy indicated that solution blow technique did not influence the molecular nature of native polymers. Obtained nanofibers exhibited sufficient wound exudate absorbency, which appears beneficial to moisturize the wound bed during the healing process. Formulations displayed greater tensile strength as compared to commercial hydrofiber-like dressing materials comprised of carboxymethylcellulose sodium or calcium alginate, which points toward their protective function against mechanical stress. Coating with hydrophobic poly(dimethylsiloxane) (applied to favor nanofiber removal from the wound surface) impacted porosity and decreased both mechanical properties and adherence to excised human skin, though the obtained values were comparable to those attained for commercial hydrofiber-like materials. In vitro cytotoxicity and irritancy studies showed biocompatibility and no skin irritant response of nanofibers in contact with a reconstituted three-dimensional human skin model, while scratch assay using human fibroblast cell line HDFa revealed the valuable potential of CS/PEO nanofibers to promote cell migration at an early stage of injury.

LC-PDA-MS and GC-MS Analysis of Scorzonera hispanica Seeds and Their Effects on Human Breast Cancer Cell Lines.

Scorzonera hispanica is an herbaceous perennial cultivated in Central and Southern Europe. This study aimed to qualitatively and quantitatively evaluate the composition of oil, extracts, and fractions (SH1-SH12) obtained from S. hispanica seeds. Furthermore, an evaluation of biological activities in breast cancer cell lines was also performed. GC-MS analysis revealed that the primary components of the seed oil (SH12) were fatty acids and ß-sitosterol. In the evaluation of extracts (SH1-SH3, SH8-SH10) and fractions (SH4-SH7, SH11) composition, the presence of apigenin, derivatives of p-coumaric and caffeic acids, was reported. In the biological assays, methanolic extract (SH1), diethyl ether (SH4), and chloroform (SH11) fractions exhibited cytotoxicity toward cells. The highest activity was observed for fatty acids- and 3,4-dimethoxycinnamate-rich SH11 (IC50: 399.18 µg/mL for MCF-7, 781.26 µg/mL for MDA-MB-231). SH11 was also observed to induce apoptosis in MCF-7 cells (52.4%). SH1, SH4, and SH11 attenuate signaling pathways and affect the expression of apoptosis-, autophagy-, and inflammation-related proteins. SH12 was non-toxic toward either cancer or normal cell lines in concentrations up to 1 mg/mL. The results suggest that S. hispanica seeds exhibit a wide range of potential uses as a source of oil and bioactive compounds for complementary therapy of breast cancer.

2-{5-[(Z,2Z)-2-Chloro-3-(4-nitrophenyl)-2-propenylidene]-4-oxo-2-thioxothiazolidin-3-yl}-3-methylbutanoic Acid as a Potential Anti-Breast Cancer Molecule.

It was established that the synthesis of hybrid molecules containing a thiazolidinone and a (2Z)-2-chloro-3-(4-nitrophenyl)prop-2-ene structural fragments is an effective approach for the design of potential anticancer agents. Given the results of the previous SAR-analysis, the aim of the study was to synthesize a novel 4-thiazolidinone derivative Les-3331 and investigate its molecular mechanism of action in MCF-7 and MDA-MB-231 breast cancer cells. The cytotoxic properties and antiproliferative potential of Les-3331 were determined. The effect of the tested compound on apoptosis induction and mitochondrial membrane potential was checked by flow cytometry. ELISA was used to determine caspase-8 and caspase-9, LC3A, LC3B, Beclin-1, and topoisomerase II concentration. Additionally, PAMPA, in silico or in vitro prediction of metabolism, CYP3A4/2D6 inhibition, and an Ames test were performed. Les-3331 possesses high cytotoxic and antiproliferative activity in MCF-7 and MDA-MB-231 breast cancer cells. Its molecular mechanism of action is associated with apoptosis induction, decreased mitochondrial membrane potential, and increased caspase-9 and caspase-8 concentrations. Les-3331 decreased LC3A, LC3B, and Beclin-1 concentration in tested cell lines. Topoisomerase II concentration was also lowered. The most probable metabolic pathways and no DDIs risk of Les-3331 were confirmed in in vitro assays. Our studies confirmed that a novel 4-thiazolidinone derivative represents promising anti-breast cancer activity.

Synthesis and Anticancer Activity of 1,3,4-Thiadiazoles with 3-Methoxyphenyl Substituent.

Based on the results of previous work, we designed and synthesized 1,3,4-thiadiazole derivatives. The cytotoxic activity of the obtained compounds was then determined in biological studies using MCF-7 and MDA-MB-231 breast cancer cells and a normal cell line (fibroblasts). The results showed that all compounds displayed weak anticancer activity towards two breast cancer lines: an estrogen-dependent cell line (MCF-7) and an estrogen-independent cell line (MDA-MB-231). The compound most active towards MCF-7 breast cancer cells was SCT-4, which decreased DNA biosynthesis to 70% ± 3 at 100 µM. The mechanism of the anticancer action of 1,3,4-thiadiazole was also investigated. We choose a set of the most investigated proteins, which are attractive anticancer targets. In silico studies demonstrated a possible multitarget mode of action for the synthesized compounds but the most likely mechanism of action for the new compounds is connected with the activity of caspase 8.

Development of Novel Pyridine-Thiazole Hybrid Molecules as Potential Anticancer Agents.

Novel pyridine-thiazole hybrid molecules were synthesized and subjected to physico-chemical characterization and screening of their cytotoxic action towards a panel of cell lines derived from different types of tumors (carcinomas of colon, breast, and lung, glioblastoma and leukemia), and normal human keratinocytes, for comparison. High antiproliferative activity of the 3-(2-fluorophenyl)-1-[4-methyl-2-(pyridin-2-ylamino)-thiazol-5-yl]-propenone 3 and 4-(2-{1-(2-fluorophenyl)-3-[4-methyl-2-(pyridin-2-ylamino)-thiazol-5-yl]-3-oxopropylsulfanyl}-acetylamino)-benzoic acid ethyl ester 4 was revealed. The IC50 of the compound 3 in HL-60 cells of the acute human promyelocytic leukemia was 0.57 µM, while in the pseudo-normal human cell lines, the IC50 of this compound was >50 µM, which suggests that the compounds 3 and 4 might be perspective anticancer agents. The detected selectivity of the derivatives 3 and 4 for cancer cell lines inspired us to study the mechanisms of their cytotoxic action. It was shown that preincubation of tumor cells with Fluzaparib (inhibitor of PARP1) reduced the cytotoxic activity of the derivatives 3 and 4 by more than twice. The ability of these compounds to affect DNA nativity and cause changes in nucleus morphology allows for the suggestion that the mechanism of action of the novel pyridine-thiazole derivatives might be related to inducing the genetic instability in tumor cells.

Exploration of novel heterofused 1,2,4-triazine derivative in colorectal cancer.

Colorectal cancer (CRC) is the third leading cause of cancer-related deaths in men and in women. The impact of the new pyrazolo[4,3-e]tetrazolo[1,5-b][1,2,4]triazine sulphonamide (MM-129) was evaluated against human colon cancer in vitro and in zebrafish xenografts. Our results show that this new synthesised compound effectively inhibits cell survival in BTK-dependent mechanism. Its effectiveness is much higher at a relatively low concentration as compared with the standard chemotherapy used for CRC, i.e. 5-fluorouracil (5-FU). Flow cytometry analysis after annexin V-FITC and propidium iodide staining revealed that apoptosis was the main response of CRC cells to MM-129 treatment. We also found that MM-129 effectively inhibits tumour development in zebrafish embryo xenograft model, where it showed a markedly synergistic anticancer effect when used in combination with 5-FU. The above results suggest that this novel heterofused 1,2,4-triazine derivative may be a promising candidate for further evaluation as chemotherapeutic agent against CRC.

Selenium Compounds as Novel Potential Anticancer Agents.

The high number of new cancer incidences and the associated mortality continue to be alarming, leading to the search for new therapies that would be more effective and less burdensome for patients. As there is evidence that Se compounds can have chemopreventive activity, studies have begun to establish whether these compounds can also affect already existing cancers. This review aims to discuss the different classes of Se-containing compounds, both organic and inorganic, natural and synthetic, and the mechanisms and molecular targets of their anticancer activity. The chemical classes discussed in this paper include inorganic (selenite, selenate) and organic compounds, such as diselenides, selenides, selenoesters, methylseleninic acid, 1,2-benzisoselenazole-3[2H]-one and selenophene-based derivatives, as well as selenoamino acids and Selol.

Mechanism of Anticancer Action of Novel Imidazole Platinum(II) Complex Conjugated with G2 PAMAM-OH Dendrimer in Breast Cancer Cells.

Transition metal coordination compounds play an important role in the treatment of neoplastic diseases. However, due to their low selectivity and bioavailability, as well as the frequently occurring phenomenon of drug resistance, new chemical compounds that could overcome these phenomena are still being sought. The solution seems to be the synthesis of new metal complexes conjugated with drug carriers, e.g., dendrimers. Numerous literature data have shown that dendrimers improve the bioavailability of the obtained metal complexes, solving the problem of their poor solubility and stability in an aqueous environment and also breaking down inborn and acquired drug resistance. Therefore, the aim of this study was to synthesize a novel imidazole platinum(II) complex conjugated with and without the second-generation PAMAM dendrimer (PtMet2-PAMAM and PtMet2, respectively) and to evaluate its antitumor activity. Cell viability studies indicated that PtMet2-PAMAM exhibited higher cytotoxic activity than PtMet2 in MCF-7 and MDA-MB-231 breast cancer cells at relatively low concentrations. Moreover, our results indicated that PtMet2-PAMAM exerted antiproliferative effects in a zebrafish embryo model. Treatment with PtMet2-PAMAM substantially increased apoptosis in a dose-dependent manner via caspase-9 (intrinsic pathway) and caspase-8 (extrinsic pathway) activation along with pro-apoptotic protein expression modulation. Additionally, we showed that apoptosis can be induced by activating POX, which induces ROS production. Furthermore, our results also clearly showed that the tested compounds trigger autophagy through p38 pathway activation and increase Beclin-1, LC3, AMPK, and mTOR inhibition. The high pro-apoptotic activity and the ability to activate autophagy by the imidazole platinum(II) complex conjugated with a dendrimer may be due to its demonstrated ability to reverse multidrug resistance (MDR) and thereby increase cellular accumulation in breast cancer cells.

Platinum and Palladium Complexes as Promising Sources for Antitumor Treatments.

There is a need for new, safer, and more effective agents to treat cancer. Cytostatics that have transition metals at their core have attracted renewed interest from scientists. Researchers are attempting to use chemotherapeutics, such as cisplatin, in combination therapy (i.e., in order to enhance their effectiveness). Moreover, studies are being carried out to modify molecules, by developing them into multinuclear structures, linking different compounds to commonly used drugs, or encapsulating them in nanoparticles to improve pharmacokinetic parameters, and increase the selectivity of these drugs. Therefore, we attempted to organize recent drug findings that contain palladium and platinum atoms in their structures.

The Paired Siglecs in Brain Tumours Therapy: The Immunomodulatory Effect of Dexamethasone and Temozolomide in Human Glioma In Vitro Model.

The paired sialic acid-binding immunoglobulin like lectins (Siglecs) are characterized by similar cellular distribution and ligand recognition but opposing signalling functions attributed to different intracellular sequences. Since sialic acid-Siglec axis are known to control immune homeostasis, the imbalance between activatory and inhibitory mechanisms of glycan-dependent immune control is considered to promote pathology. The role of sialylation in cancer is described, however, its importance in immune regulation in gliomas is not fully understood. The experimental and clinical observation suggest that dexamethasone (Dex) and temozolomide (TMZ), used in the glioma management, alter the immunity within the tumour microenvironment. Using glioma-microglia/monocytes transwell co-cultures, we investigated modulatory action of Dex/TMZ on paired Siglecs. Based on real-time PCR and flow cytometry, we found changes in SIGLEC genes and their products. These effects were accompanied by altered cytokine profile and immune cells phenotype switching measured by arginases expression. Additionally, the exposure to Dex or TMZ increased the binding of inhibitory Siglec-5 and Siglec-11 fusion proteins to glioma cells. Our study suggests that the therapy-induced modulation of the interplay between sialoglycans and paired Siglecs, dependently on patient's phenotype, is of particular signification in the immune surveillance in the glioma management and may be useful in glioma patient's therapy plan verification.

In Vitro Anticancer Potential of Jasione montana and Its Main Components against Human Amelanotic Melanoma Cells.

Jasione montana L. (Campanulaceae) is used in traditional Belarusian herbal medicine for sleep disorders in children, but the chemical composition and biological activity have not been investigated. In this study, the activities of J. montana extracts, their fractions and main compounds were evaluated in amelanotic melanoma C32 (CRL-1585) cells and normal fibroblasts (PCS-201-012). The extracts and fractions were analyzed using liquid chromatography-photodiode array detection-electrospray ionization-mass spectrometry (LC-PDA-ESI-MS/TOF) to characterize 25 compounds. Further, three major and known constituents, luteolin (22) and its derivatives such as 7-O-glucoside (12) and 7-O-sambubioside (9) were isolated and identified. The cytotoxic activities against fibroblasts and the amelanotic melanoma cell line were determined using the fixable viability stain (FVS) assay. The influence of diethyl ether (Et2O) fraction (JM4) and 22 on apoptosis induction was investigated using an annexin V binding assay. The obtained results showed significant cytotoxicity of JM4 and 22 with IC50 values of 119.7 ± 3.2 and 95.1 ± 7.2 µg/mL, respectively. The proapoptotic potential after 22 treatment in the C32 human amelanotic melanoma cell line was comparable to that of vinblastine sulfate (VLB), detecting 29.2 ± 3.0% apoptotic cells. Moreover, 22 displayed less necrotic potential against melanoma cells than VLB. In addition, the influences of JM4 and 22 on the dysfunction of the mitochondrial membrane potential (MMP), cell cycle and activity of caspases 3, 8, 9, and 10 were established. The effects of JM4 on MMP change (74.5 ± 3.0% of the cells showed a reduced MMP) corresponded to the results obtained from the annexin V binding assay and activation of caspase-9. JM4 and 22 displayed a significant impact on caspase-9 (40.9 ± 2.4% of the cells contained active caspase-9 after JM4 treatment and 16.6 ± 0.8% after incubation with 22) and the intrinsic (mitochondrial) apoptotic pathway. Moreover, studies have shown that JM4 and 22 affect the activation of external apoptosis pathways by inducing the caspase-8 and caspase-10 cascades. Thus, activation of caspase-3 and DNA damage via external and internal apoptotic pathways were observed after treatment with JM4 and 22. The obtained results suggest that J. montana extracts could be developed as new topical preparations with potential anticancer properties due to their promising cytotoxic and proapoptotic potential.