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PMC11085211
Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors
Interestingly, 2A-35 and 2A-36 subfractions were also active at 6 µg/mL, thus representing the most active ones (Figure 8b).
PMC11085211
Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors
To make reading easier, the figures only include the nine active subfractions delimitated by the first available adjacent weakly or non-active subfractions tested (i.e., with available material).
PMC11085211
Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors
Interestingly, groups of numerically adjacent subfractions had similar activity, suggesting that they could harbor the same bioactive compound(s).
PMC11085211
Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors
Based on this assumption, we combined active contiguous subfractions into four clusters (C.2A-29-31, C.2A-35-36, C.2A-48-49, and C.2A-60-61).
PMC11085211
Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors
The toxicity of FR2-A-derived clusters was evaluated in peripheral blood lymphocytes (PBMCs).
PMC11085211
Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors
In particular, PBMCs and GIST-T1 were treated with the established four clusters at 30 µg/mL, while the 2A-20 subfraction, previously evaluated as inactive in GIST cells, was used as a negative control (Figure 9a).
PMC11085211
Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors
All clusters significantly reduced the viability of both GIST-T1 and PBMCs.
PMC11085211
Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors
In particular, C.2A-29-31 and C.2A-35-36 reduced viability by approximately 99% (p < 0.0001) in both PBMCs and GIST-T1 cells.
PMC11085211
Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors
C.2A-48-49 and C.2A-60-61 showed a lower viability reduction compared to the previous ones and more prominent cytotoxicity in PBMCs than in GIST cells.
PMC11085211
Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors
Altogether, the observed cytotoxicity on PBMCs highlighted a non-specific mechanism of action.
PMC11085211
Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors
In this regard, doxorubicin, a chemotherapeutic drug successfully used to treat numerous solid tumors, is frequently associated with myelosuppressive side effects.
PMC11085211
Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors
In agreement with this, doxorubicin significantly impaired PBMC viability, similarly to FR2A clusters (Figure 9b), supporting the hypothesis that phytochemicals in the FR2A clusters could have chemotherapeutic properties.
PMC11085211
Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors
The nine FR2-A active subfractions obtained from SEC were individually analyzed through H NMR to obtain an overview of the main phytochemical constituents.
PMC11085211
Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors
Additionally, UHPLC-MS was performed both in the support of NMR analysis and in an attempt to identify the molecules present at the lowest concentrations.
PMC11085211
Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors
The adjacent weakly or non-active subfractions were also phytochemically investigated.
PMC11085211
Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors
Interestingly, the analysis revealed the presence of pyrogallol-bearing compounds in all the active subfractions (Figure 10 and Supplementary Figure S6a).
PMC11085211
Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors
In particular, the active subfractions forming the cluster C.2A-29-31 harbor a pyrogallol derivative, whose structure remains to be not yet fully elucidated (Supplementary Figure S6b), together with a high quantity of fumaric acid; both were not found in the non-active subfractions.
PMC11085211
Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors
Regarding the subfractions forming the most potent cluster, C.2A-35-36, they predominantly contained gallic acid and the hexoside of the flavonoid myricetin, both bearing a pyrogallol moiety (Supplementary Figure S6c).
PMC11085211
Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors
Interestingly, the adjacent, weakly active subfraction—2A-33—also contained gallic acid, although at a lower concentration compared to the active cluster, supporting the involvement of this molecule in the weaker biological activity.
PMC11085211
Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors
The other adjacent subfraction—2A-39—mainly contained galloyl-arbutin, a pyrogallol derivative, and its weak activity could be attributed again to the pyrogallol moiety.
PMC11085211
Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors
Indeed, arbutin has been tested on GIST cells with negative results, and its non-anticancer activity was also confirmed by the absence of activity in the 2A-20 subfraction, which predominantly contained arbutin (Supplementary Figure S6a).
PMC11085211
Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors
Regarding the subfraction forming the cluster C.2A-48-49, the predominant molecule was gallocatechin, which also bears a pyrogallol moiety in its structure (Supplementary Figure S6d).
PMC11085211
Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors
The predominant phytoconstituent of the adjacent non-active subfraction 2A-47 was catechin (structurally similar to gallocatechin without the pyrogallol substituent).
PMC11085211
Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors
Hence, it could be hypothesized that the presence of the pyrogallol moiety is essential for the anticancer activity in GIST, a hypothesis supported once again by the analysis of the active cluster C.2A-60-61.
PMC11085211
Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors
Indeed, this cluster also presented a pyrogallol-bearing compound, trigalloyl-glucose, as the main phytoconstituent (Supplementary Figure S6e).
PMC11085211
Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors
In contrast, neither trigalloyl-glucose nor additional pyrogallol-bearing compounds were identifiable in the adjacent non-active 2A-58 subfraction.
PMC11085211
Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors
In view of these results, gallic acid, bearing the pyrogallol moiety and already associated with anticancer activity , was tested in both GIST-T1 and GIST-882 cells, similarly to what was previously performed for β-arbutin; the 2A-35 subfraction was used as a positive control.
PMC11085211
Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors
Treatment with 30 µg/mL of gallic acid significantly impaired cell viability (p < 0.0001) similarly to the 2A-35 subfraction (Supplementary Figure S7).
PMC11085211
Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors
Differently, the treatment with 6 µg/mL had no significant effect, while, as expected, 2A-35 significantly impaired the viability in both GIST cell lines (p < 0.0001) (Supplementary Figure S7), suggesting that gallic acid is not sufficient, although it may promote an effect in concert with other phytochemicals.
PMC11085211
Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors
Arbutus unedo L. (Ericaceae family), known as the strawberry tree, is an evergreen shrub growing in circum-Mediterranean regions.
PMC11085211
Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors
Leaves are used in the traditional medicine of the Iberian Peninsula and Sardinia (Italy) to treat many illnesses , demonstrating a number of health-promoting properties associated with this plant.
PMC11085211
Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors
In this work, we showed that an extract from Arbutus unedo L. leaves (indicated as AUN) induces early apoptosis in GIST cells, exerting chemotherapeutic properties.
PMC11085211
Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors
This result is of great importance, considering chemotherapy is generally ineffective in advanced GISTs, and no options are available.
PMC11085211
Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors
Patients’ treatment first relies on imatinib as the first-line therapy.
PMC11085211
Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors
Indeed, primary GISTs are commonly associated with gain-of-function mutations in KIT or PDGFRα (about 90% of cases), the disease drivers which, once mutated, show a ligand-independent dysregulated activity .
PMC11085211
Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors
Moreover, given that secondary mutations in both KIT or PDGFRα have been found in imatinib-resistant GISTs, multi-target TKIs, such as sunitinib, regorafenib, and ripretinib, are administered as further treatment lines after the onset of resistance .
PMC11085211
Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors
However, multi-target TKIs have not kept their promises, only modestly improving the outcomes of imatinib-resistant GIST patients.
PMC11085211
Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors
Indeed, imatinib-resistant subclones escape through alternative pathways that are not targetable from multi-target TKIs, suggesting that a TKI-based approach alone may not represent the only or best option for GIST patients .
PMC11085211
Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors
Therefore, GIST patients urgently require the identification of novel therapies other than TKIs.
PMC11085211
Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors
In this context, we hypothesized that plant extracts could represent an important source of novel NPs to be studied in GIST.
PMC11085211
Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors
Among these, AUN showed antiproliferative activity in osteosarcoma (U2OS) cells , representing a promising unexplored therapeutic strategy in GIST.
PMC11085211
Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors
In view of this consideration, we tested the anticancer activity of AUN in GIST cellular models.
PMC11085211
Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors
AUN significantly impaired GIST cell viability, triggering a significant Annexin (+)/7-AAD (−) cell population, a marker of early apoptosis.
PMC11085211
Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors
AUN was further explored by bio-guided fractionation to deepen what could be the most active part of AUN.
PMC11085211
Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors
Hence, AUN was first fractionated by a liquid/liquid partition, separating roughly AUN phytochemicals on the basis of their polarity.
PMC11085211
Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors
This first step led to the identification of the FR2 fraction as the most active, able to impair cell viability and promote early apoptosis in different models of imatinib-sensitive GISTs (GIST-882 and GIST-T1), i.e., characterized by different primary kit mutations.
PMC11085211
Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors
In view of this consideration, we performed a second step of fractionation, subjecting the FR2 fraction to a reverse-phase MPLC, which led to the identification of the FR2-A fraction as the most active one in imatinib-sensitive cellular models (GIST-882 and GIST-T1), regardless of primary KIT mutations.
PMC11085211
Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors
Moreover, it resulted in being even more potent than FR2, as it significantly impaired cell viability at lower concentrations (12.5 μg/mL vs. 50 μg/mL).
PMC11085211
Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors
Afterward, FR2-A was tested in imatinib-resistant cell lines (GIST48 and GIST-48b), once again proving effective in impairing cell viability, providing new insight into the FR2-A mechanism of action.
PMC11085211
Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors
Indeed, the cell line GIST-48b is characterized by the absence of KIT expression, supporting the hypothesis that the mechanism is independent from KIT, possibly paving the way for developing therapeutic strategies other than TKIs in imatinib resistance.
PMC11085211
Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors
Additionally, FR2-A was more efficient with respect to imatinib in sensitive cells, inducing a higher percentage of Annexin V (+)/7-AAD (−) cells.
PMC11085211
Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors
This result confirmed the capacity of AUN to efficiently induce early apoptosis, prompting us to deepen the mechanism.
PMC11085211
Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors
Nevertheless, we did not observe the cleavage of caspase 3, caspase 9, and PARP-1, suggesting a cell death mechanism other than classical apoptosis.
PMC11085211
Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors
Interestingly, although not cleaved, a remarkable and rapid PARP-1 downregulation after FR2-A treatment was observed, possibly suggesting an epigenetic mechanism.
PMC11085211
Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors
Aware of the limitation, mainly due to the limited number of GIST cell models analyzed—though it depends on the availability of commercial cell lines—we are conscious that our results are speculative.
PMC11085211
Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors
However, recent findings in the literature seem to support our hypothesis of an epigenetic mechanism of action.
PMC11085211
Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors
Indeed, a study has identified PARP-1 as a target of miR-7-5p in a model of lung cancer cells, demonstrating that sponging miR-7-5p promotes the homologous repair (HR) path through upregulating PARP1 expression .
PMC11085211
Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors
In another study, Garmutin-A (GA), an NP isolated from Garcinia multiflora, induced apoptosis via the upregulation of miR-17-5p, triggering PARP-1 downregulation, in leukemic CB3 cells .
PMC11085211
Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors
Overall, this finding is in line with the evidence that many NPs could exert their anticancer activity by affecting miRNAs expression .
PMC11085211
Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors
Therefore, a possible speculation is that FR2-A could upregulate different miRNAs, thereby altering the PARP-1-mediated repair capability.
PMC11085211
Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors
Another thought offered by PARP-1 downregulation is the possibility of re-thinking imatinib treatment in GIST, considering synergistic/combinatory approaches.
PMC11085211
Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors
Indeed, most GIST patients commonly achieve partial response or stable disease, suggesting that GIST cells could survive the treatment by activating adaptive responses.
PMC11085211
Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors
In this context, our study corroborates that imatinib mainly stabilizes GIST cells in a non-proliferative state rather than inducing cell death.
PMC11085211
Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors
Therefore, a combination of imatinib and PARP-1 inhibitors could represent a promising strategy that has not yet been investigated in GIST.
PMC11085211
Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors
Regarding the possible bio-active compound characterizing FR2-A, we initially focused on β-arbutin.
PMC11085211
Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors
Indeed, this phytochemical, already identified as one of the primary metabolites in AUN , was identified as the most representative compound in FR2-A. Strengthened by this result, together with the knowledge that β-arbutin was already associated with proapoptotic activity in melanoma cells , we tested pure β-arbutin, unfortunately with negative results.
PMC11085211
Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors
This prompted us to perform an additional fractionation step, this time with a size-exclusion approach, to increase the concentrations of active phytochemicals.
PMC11085211
Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors
This approach led to sixty-one different subfractions, of which nine, forming four different clusters, exhibited a significant biological activity in GIST cells.
PMC11085211
Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors
Interestingly, all clusters contained phytochemicals characterized by the pyrogallol moiety, a polyphenolic structure already associated with proapoptotic activity.
PMC11085211
Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors
A study conducted on As4.1 juxtaglomerular cells, a model of benign kidney tumors, demonstrated that pyrogallol treatment could efficiently induce apoptosis .
PMC11085211
Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors
Interestingly, the authors observed caspase 3 and PARP-1 cleavage; however, treatment with caspase 3 inhibitors did not prevent apoptosis, representing an intriguing result.
PMC11085211
Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors
Indeed, the authors did not evaluate the PARP-1 status in the presence of a caspase 3 inhibitor.
PMC11085211
Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors
Therefore, other mechanisms besides caspase activation could not be excluded.
PMC11085211
Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors
Another study demonstrated that the pyrogallol moiety in polyphenols had an important role to play in apoptosis induction in human embryonic kidney cells (HEK293T) and the chronic myelogenous leukemia cell line (K562), even though the mechanism was not investigated .
PMC11085211
Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors
In the same study, the authors highlighted that the pyrogallol moiety was important for cytotoxic activity.
PMC11085211
Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors
In agreement with this, all the FR2-A active subfractions showed cytotoxicity in PBMCs, similar to those observed with doxorubicin, a traditional chemotherapeutic.
PMC11085211
Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors
These findings suggest a non-targeted but chemotherapy-like mechanism of action, which should not be underestimated considering the lack of effective chemotherapeutic agents for GIST patients.
PMC11085211
Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors
Indeed, doxorubicin, approved in several solid tumor treatments, has been tested in clinical trials with poor response in GISTs, results that led to its non-approval .
PMC11085211
Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors
In confirmation of this, unlike FR2-A clusters, doxorubicin does not impair the viability of imatinib-resistant cells .
PMC11085211
Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors
Therefore, deepening FR2-A composition could be of great interest in GIST, leading to the identification of novel chemotherapeutic agents to be used as alternatives or in combination with the approved TKI.
PMC11085211
Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors
In view of these considerations, the most active FR2-A subfractions (2A-35 and 2A-36) were mainly characterized by the presence of gallic acid, a phenolic acid which has been identified as a promising health-promoting agent in many conditions, including cancer .
PMC11085211
Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors
Despite the expectations, our finding ascribed an activity to gallic acid, which, however, was weaker than 2A-35.
PMC11085211
Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors
Therefore, gallic acid cannot be considered a solely bioactive compound but rather one that acts in concert with other components, highlighting the importance of the phytocomplex.
PMC11085211
Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors
In conclusion, although bio-guided fractionation did not lead to the identification of a single compound active in GIST cellular models, it was confirmed as a feasible tool for circumscribing phytochemicals of interest.
PMC11085211
Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors
In particular, in the present study, we identified pyrogallol-bearing compounds as the main players in GIST cell cytotoxicity, harboring chemotherapeutic properties in imatinib-sensitive and resistant cells.
PMC11085211
Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors
These data may fuel further studies searching for novel treatments in GIST, including additional strategies that augment the existing treatments, resulting in better patient outcomes.
PMC11085211
Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors
Therefore, the chemotherapeutic-like activity should not discourage further preclinical studies on pyrogallol-bearing compounds in GISTs to conclude whether they could represent an alternative strategy for treating GISTs.
PMC11085211
Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors
GIST-882 and GIST-T1 are defined as primary mutated and imatinib-sensitive cellular models.
PMC11085211
Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors
They harbor exon 9 (K642E) homozygous mutations and exon 13 heterozygous (V560-Y579) in the KIT gene, respectively.
PMC11085211
Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors
GIST-882 and GIST-T1 were grown in RPMI-1640 supplemented with 15% FBS.
PMC11085211
Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors
GIST-48 is instead reported as an imatinib- and sunitinib-resistant cell line harboring a primary homozygous mutation on KIT exon 11 (V560D) and an additional secondary heterozygous mutation in exon 17 (D820A).
PMC11085211
Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors
GIST-48b was established in vitro, starting from GIST-48 after HSP-90 inhibitor (17-AAG) drug pressure selection, resulting in a subline characterized by nearly undetectable KIT transcript and protein.
PMC11085211
Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors
GIST-48 and GIST-48b were grown in Iscove’s Modified Dulbecco’s Medium (IMDM) supplemented with 15% FBS.
PMC11085211
Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors
All the indicated cell lines were routinely tested to avoid mycoplasma contamination with MycoBlue Mycoplasma Detector (Vazyme, Nanjing, China).
PMC11085211
Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors
GIST-T1 was purchased from CosmoBio (Tokyo, Japan), while GIST-48 and GIST-882 cells were kindly provided by Fletcher JA, MD (Harvard Medical School).
PMC11085211
Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors
The leaves of Arbutus unedo L. were harvested in Sardinia in 2018 (voucher specimen CAG 878/v, deposited at the General Herbarium of the Department of Life and Environmental Sciences, University of Cagliari).
PMC11085211
Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors
To ensure reproducibility, the plant material was analyzed by H NMR profiling .
PMC11085211
Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors
10 GIST cells were seeded in a 24-well cell culture plate the day before treatment.
PMC11085211
Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors
Cells were treated with AUN or AUN subfractions at the indicated final concentrations in the culture medium for the indicated time points.
PMC11085211
Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors
The analysis of cell viability and the presence of apoptotic cells were evaluated on the entire cellular population, including cells that were in suspension due to the treatment.