Research ArticleExperimental Studies
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CD9-positive Exosomes Derived from Cancer-associated Fibroblasts Might Inhibit the Proliferation of Malignant Melanoma Cells

NAHO FUJII, MASAKAZU YASHIRO, TAKAHARU HATANO, HEISHIRO FUJIKAWA and HISASHI MOTOMURA
Anticancer Research January 2023, 43 (1) 25-33; DOI: https://doi.org/10.21873/anticanres.16130

Abstract

Background/Aim: Exosomes secreted by various cells in the tumour microenvironment have been reported to be mediators of intercellular communication that play an important role in cancer progression. In this study, we aimed to investigate the effects of exosomes derived from cancer-associated fibroblasts (CAFs) on the proliferation of malignant melanoma (MM) cells and evaluated their clinicopathological significance. Materials and Methods: Three malignant melanoma cell lines, A375, MMAc, and COLO679, and three CAFs established from malignant melanomas at stages 1a, 2b, and 3b, were used. The expression of CD9, CD63, and CD81 in CAF-derived exosomes was examined using western blotting. The effect of exosomes on the proliferative potential of cancer cells was analysed using cell counting and MTT assays. The expression of CD9, CD63, and CD81 was also immunohistochemically analysed in 90 malignant melanoma specimens. Results: CAF-derived exosomes were positive for CD9 and CD63 and remarkably inhibited the proliferative capacity of A375 and MMAc cells. The five-year disease-free survival was significantly better in patients with CAF-derived CD9-positive exosomes than in CD9-negative patients. Conclusion: CAF-derived exosomes, especially CD9-positive exosomes, have an inhibitory effect on the proliferation of malignant melanoma cells. These findings suggest that CD9 expression in CAFs is a promising prognostic marker for patients with malignant melanoma.

Key Words:

Malignant melanoma (MM), a malignant transformation of melanocytes, is one of the most aggressive skin cancers, and its incidence rate is increasing rapidly worldwide (1). While most early-stage MMs are considered curable by surgical resection, some have been shown to occasionally develop distant metastasis (2). Moreover, pathologic factors, such as tumour thickness, mitotic rate, and the presence of ulcerations, are also considered important prognostic indicators in MM (3). However, patients with these pathologic factors do not always have a similar clinical course (2). Therefore, discovering useful markers for more accurate assessments of malignant potential in MM is necessary for developing appropriate therapeutic approaches to treat MM.

The crosstalk between stromal and cancer cells has been shown to play key roles in orchestrating the cancer progression through various mechanisms (4-7). Especially cancer-associated fibroblasts (CAFs), which are the major stromal cells in the tumour microenvironment (TME), have been implicated in the progression of cancer cells, including MM (8-12). Exosomes, the membranous nanovesicles or extracellular vesicles (EVs) of 30-150 nm in diameter secreted by various cells, mediate intercellular communication (13). In particular, exosomes secreted by CAFs have been reported to play an important role in cancer progression in several cancers, including gastric and pancreatic cancers (14-16). However, the effects of CAF-derived exosomes on the progression of MM cells remain elusive. Therefore, in this study, we aimed to investigate the effect of CAF-derived exosomes on the proliferation and invasion activity of MM cells and evaluate their clinicopathologic significance.

Materials and Methods

Cell culture and cell lines. Three MM cell lines, A375 (ATCC, Manassas, VA, USA, RRID: CVCL_B3PJ), MMAc (RIKEN cell bank, Ibaragi, Japan, RRID: CVCL_5951), and COLO679 (RIKEN cell bank, RRID: CVCL_1130), and three CAF cell lines, CAF-MM1, CAF-MM2, and CAF-MM3 derived from MM tissues were used. These three CAFs were established from MM tissues at Osaka Metropolitan University: CAF-MM1, CAF-MM2, and CAF-MM3 were established from MMs at stages 1a, 2b, and 3b, respectively, following the protocol described in a previous study (17). Briefly, the MM specimen was excised under aseptic conditions and minced using forceps and scissors. The culture medium comprised Dulbecco’s modified Eagle medium (DMEM; Wako, Osaka, Japan) supplemented with 10% fetal bovine serum (FBS; Nichirei Biosciences Inc., Tokyo, Japan), 100 IU/ml penicillin (Wako), 100 mg/ml streptomycin (Wako), and 0.5 mM sodium pyruvate (Wako). Cells were cultured in 21% O2 at 37°C. Fibroblasts were used from the third to seventh passage in culture, mainly in the fifth passage.

Isolation and characterization of exosomes. Exosomes were obtained from the supernatant of cells as previously described with some modifications (18, 19). In brief, after being grown to semi-confluence in DMEM containing 10% FCS, cells were washed with phosphate-buffered solution (PBS; Wako) and incubated for an additional 48 h in 10 ml serum-free DMEM. The supernatant was collected and centrifuged at 2,000 × g for 10 min to discard cellular debris. Subsequently, the supernatant was filtered using a 0.8 μm pore filter (Millipore, Billerica, MA, USA). The collected filtrate was then ultracentrifuged at 1×105 × g for 1.5 h at 4°C. The exosome-containing pellet was washed with 25 ml PBS, followed by a second step of ultracentrifugation at 1×105 × g for 1.5 h at 4°C. The supernatant was discarded, and the pellet was stored at −80°C. The morphologic characteristics of exosomes were observed by transmission electron microscopy (HT7700; HITACHI, Tokyo, Japan). The amount and number of exosomes were quantified using the Micro BCA™ Protein Assay Kit (Thermo Fisher Scientific, Waltham, MA, USA).

Proliferation assay. The proliferation and viability of A375, MMAc, and COLO679 cells were determined using a 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay or by counting the number of cancer cells. Briefly, A375 and MMAc cells at 6×103 per well and COLO679 cells at 8×103 per well were seeded in 96-well plates with culture medium and incubated for 72 h. Cancer cells were cultured in DMEM supplemented with 1.5% FBS in the presence of CAF-derived exosomes (2 μg/ml) or PBS as control. Subsequently, 100 μl culture medium and 20 μl MTT solution (Promega, Tokyo, Japan) were added to each well. The absorbance at 570 nm was analysed using a microplate reader (Bio-Rad 550; Bio-Rad, Tokyo, Japan). The number of cells was calculated using Coulter Z2 (Beckman Coulter, Fullerton, CA, USA) after culture in DMEM supplemented with 1.5% FBS in the presence of CAF-derived exosomes (2 μg/ml) or PBS as control. A375 and MMAc cells were seeded at 3.6×104 per well, whereas COLO679 cells were seeded at 4.8×104 per well in 24-well plates with culture medium and incubated for 48 h.

Western blotting of exosomes. Exosomes (1 μg protein) were subjected to 10% sodium dodecyl-sulfate polyacrylamide gel electrophoresis, and the separated proteins were transferred onto polyvinylidene difluoride membranes using the Trans-Blo Turbo Transfer System (BIO-RAD, Hercules, CA, USA) according to the manufacturer’s instructions. Membranes were then placed in PBS-T solution containing respective primary antibodies: anti-CD9, anti-CD63, and anti-CD81 (1:200; all from Life Technologies, Carlsbad, CA, USA). Subsequently, membranes were incubated with anti-mouse HRP-conjugated secondary antibody (1:300; Cell Signaling, Danvers, MA, USA). The bands were detected using enhanced chemiluminescence using the ECL prime (GE Health Care, Little Chalfont, UK). Western blots were analysed using the luminescent image analyser LAS 4000-plus (Fuji, Tokyo, Japan).

Patients. Ninety patients who had undergone resection of primary MM at the Hospital of Osaka Metropolitan Medical University were enrolled in this study. Pathologic diagnoses and classifications were performed according to the AJCC Cancer Staging Manual, 8th edition (2017). The study protocol conformed to the ethical guidelines of the Declaration of Helsinki and was approved by the Osaka Metropolitan University ethics committee (approved number: 3422). Written informed consent was obtained from all patients.

Immunohistochemical determination of CD9, CD63, and CD81. Immunohistochemical staining was performed on 90 melanoma specimens. Sides were deparaffinized and then heated with Target Retrieval Solution (Dako, Carpinteria, CA, USA) for 10 min at 105°C in an autoclave. After blocking endogenous peroxidase activity, specimens were incubated with CD9, CD63, and CD81 antibodies for 1 h at 25°C, followed by incubation with biotinylated mouse anti-rabbit IgG for 10 min. Slides were treated with streptavidin-peroxidase reagent (Nichirei Biosciences, Tokyo, Japan), followed by counterstaining with Giemsa stain (Merck, Darmstadt, Germany). The expression of CD9 and CD63 was evaluated by the intensity of staining and percentage of stained cancer and stromal cells, respectively. In particular, the intensity was given a score of 0-3 (0=no, 1=weak, 2=moderate, 3=intense), while the percentage of immune-positive cells was given a score of 0-3 (0=0%, 1=10%, 2=20-30%, 3=40-100%). The two scores were multiplied to obtain a final score of 0-9. Expression was considered positive in tumour cells when scores were 3 or higher and negative when scores were 0-2. The expression of CD81 was evaluated as positive if scores were 1-9, as the number of cases expressing CD81 was small. Scoring was performed by two double-blinded independent observers unaware of clinical data and outcomes.

Statistical analysis. Associations between the expression of CD9, CD63, and CD81 and clinicopathological findings were analyzed using the chi-square test. For smaller sample sizes, Fisher’s exact test was used. Disease-free survival (DFS) rate was defined as the time from surgery to recurrence from any cause. DFS curves were estimated using the Kaplan–Meier method and compared using the log-rank test. Multivariate analysis was performed using the Cox proportional hazard model. For proliferation assays, the t-test was used to determine the significance of differences between groups. All statistical analyses were performed using the R (version 4.2.1, Vienna, Austria). A two-sided probability (p) value of <0.05 was considered statistically significant.

Results

Expression level of CD9, CD63, and CD81 in exosomes from CAFs. Electron microscope findings showed that the diameters of exosomes from the three CAFs, CAF-MM1, CAF-MM2, and CAF-MM3, ranged from 20 to 100 nm (Figure 1A). Western blotting indicated that CD9 and CD63 were expressed in all CAF-derived exosomes, whereas CD81 was not expressed in any CAF-derived exosomes. CD9 and CD63 were most strongly expressed in CAF-MM2 exosomes among the 3 CAF-derived exosomes (Figure 1B).

Figure 1.
Figure 1.

Detection of exosomes derived from cancer-associated fibroblasts (CAFs). (A) Identification of CAF-derived exosomes. Exosomes were observed under a transmission electron microscope and characterized as bound vesicles. Scale bar=100 nm. (B) Western blot analysis showing the expression of CD9, CD63, and CD81 in CAF-derived exosomes. CD9 and CD63 were expressed in exosomes from CAF-MM1, CAF-MM2, and CAF-MM3. CD81 was not expressed in any of the CAF-derived exosomes.

Effect of CAF-derived exosomes on the growth of melanoma cells. MTT assay (Figure 2A) showed that the proliferative ability of MMAc cells was significantly inhibited following the addition of the three CAF-derived exosomes compared to that of the control cells (p<0.05 vs. control). The proliferation of A375 cells was significantly inhibited by the addition of CAF-MM2 and CAF-MM3 exosomes (p<0.05). In contrast, the proliferation of COLO679 cells was not significantly inhibited by any CAF-derived exosomes. As shown in Figure 2B, the growth of MMAc cells was significantly inhibited following the addition of CAF-MM2 exosomes compared to that of the control; wherein all three CAFs had significant inhibitory effects on the growth of A375 cells (p<0.05 in both cells). On the contrary, CAF-derived exosomes did not significantly affect the growth of COLO679 cells.

Figure 2.
Figure 2.

Effect of cancer-associated fibroblast (CAF)-derived exosomes on the proliferation of malignant melanoma (MM) cells. (A) MTT assay. The proliferation of MMAc cells was decreased by the addition of exosomes from CAF-MM1, CAF-MM2, and CAF-MM3. The proliferation of A375 cells was decreased by the addition of exosomes from CAF-MM1 and CAF-MM2. COLO679 cells were not affected by any of the three CAF-derived exosomes. (B) Cell counting. The number of MMAc cells was significantly decreased by addition of exosomes from CAF-MM2. The number of A375 cells was decreased by addition of exosomes from CAF-MM1, CAF-MM2, and CAF-MM3. COLO679 cells were not affected by any of the three CAF-derived exosomes. *p<0.05, **p<0.01. N.S.: not significant vs. control.

Relationship between expression CD9, CD63, and CD81, and clinicopathological factors in 90 patients with malignant melanoma. Immunohistochemical study of MM tissues indicated that the three exosome markers, CD9, CD63, and CD81, were expressed in both cancer and stromal cells (Figure 3). Next, we evaluated the expression of these markers in stromal and cancer cells of 90 patients with MM to investigate the clinicopathological association of these markers with MM. Correlation analysis between the expression of CD9, CD63, and CD81 in CAFs and the clinicopathological factors revealed a significant correlation between CD9 expression in CAFs and T classification (p=0.00189), N classification (p=0.00978), stage (p=0.000536), and presence of skin ulcers (p=0.00104). In contrast, the expression of CD63 and CD81 in CAFs did not correlate with any clinicopathologic factors (Table I). Furthermore, the expression of CD9 in melanoma cells was significantly correlated with T classification (p=0.00273) and stage (p=0.0355), but that of CD63 and CD81 was not correlated with any of the clinicopathological factors (Table II).

Figure 3.
Figure 3.

Representative pictures of CD9, CD63, and CD81 expression. Expression of CD9, CD63, and CD81 in fibroblasts (arrows) and cancer cells (asterisks). CD63 did not express in intestinal-type MM cells (asterisks) even when CD9 was found in the fibroblasts (arrows) from the same patient.

View this table:
Table I.

Correlation between CD9, CD63, and CD81 expression in CAFs and clinicopathologic factors in melanomas.

View this table:
Table II.

Correlation between CD9, CD63, and CD81 expression in cancer cells and clinicopathologic factors in melanomas.

Disease-free survival of 90 patients with malignant melanoma. Patients with CD9 expression in CAFs had a significantly better prognosis than those without CD9 (p=0.0222). In contrast, no significant difference was found in five-year disease-free survival (DFS) rates with respect to the expression of CD63 and CD81 in CAFs (Figure 4A). Moreover, the expression of CD9, CD63, and CD81 in cancer cells was not correlated with the five-year DFS rates of patients with MM (Figure 4B). As shown in Table III, univariate analysis revealed that the DFS of patients was significantly correlated with T-classification, N-classification, presence of ulcers, and CD9 expression of CAFs, wherein multivariate analysis revealed that T and N classifications were independent predictive parameters.

Figure 4.
Figure 4.

Kaplan–Meier survival curve showing disease-free survival (DFS) of patients with malignant melanoma. (A) Survival of patients with malignant melanoma expressing CD9, CD63, and CD81 in cancer cells. Kaplan–Meier survival curve indicated that the disease-free survival (DFS) of patients with CD9-positive CAFs was significantly better than that of patients with CD9-negative CAFs (p=0.022). The DFS of patients with CD63-positive and CD81-positive CAFs was not significantly different from that of those with negative expression. (B) The DFS of patients with CD9-, CD63-, and CD81-positive expression in malignant melanoma cells was not significantly different from that of the patients with negative expression.

View this table:
Table III.

Univariate and multivariate analyses with respect to disease-free survival after surgery in 90 patients with malignant melanoma.

Discussion

In this study, CAF-derived exosomes significantly inhibited the growth of MM cells. This is the first study that suggests CAF-derived exosomes may inhibit the proliferation of MM. We used three types of CAF-derived exosomes from three MM specimens of individual patients. Although all three types of CAF-derived exosomes inhibited the growth of MM, CAF-MM2-derived exosomes showed higher inhibitory effects on MM cells compared to the other CAF-derived exosomes. CD9, CD63, and CD81 are well-known surface markers of exosomes (20). All three CAF-MM-derived exosomes used in this study expressed CD9 and CD63 but not CD81. CD9 and CD63 expression was high in CAF-MM2-derived exosomes compared to that in CAF-MM1-derived and CAF-MM3-derived exosomes.

Immunohistochemical analysis of MM tissues indicated that CD9, CD63, and CD81, were expressed in stromal and cancer cells. The expression of CD9 in CAFs was significantly correlated with T classification, N classification, stage, and presence of skin ulcers. In contrast, the expression of CD63 and CD81 in CAFs was not correlated with any clinicopathological factors. These findings suggest that CD9-positive exosomes are significantly involved in the progression of MM, whereas CD63 and CD81 are not involved in melanoma grade. Although it has been reported that both CD9 and CD63 are important regulators of MM (21, 22), CD9-positive exosomes may be one of the important key factors for the progression of MM.

In cancer cells, the expression of CD9 was significantly correlated with T classification and stage, whereas CD63 and CD81 did not correlate with any clinicopathological factors. In terms of DFS, patients with CD9-expressing CAFs had a significantly better prognosis than those not expressing CD9. In univariate analysis, the DFS rate was significantly correlated with T-classification, N-classification, stage, and the CD9 expression of CAFs. In multivariate analysis, T and N classifications correlated significantly with the DFS. It has been reported that CD9 is widely expressed in exosomes released by various cancer cells and associated with cancer cell proliferation, survival, and metastasis (23, 24). These findings suggest CD9 in CAFs is a useful prognostic marker in MM.

In conclusion, CAF-derived exosomes, especially CD9-positive exosomes, have an inhibitory effect on the proliferation of MM cells. Overall, CD9 expression in CAFs may be a promising prognostic marker for patients with MM.

Acknowledgements

The Authors thank Akiko Tsuda, Yurie Yamamoto (Osaka Metropolitan University Graduate School of Medicine) for their technical assistance. This study was partially founded by JSPS KAKENHI Grant Number 19K10011.

Footnotes

  • Authors’ Contributions

    N.F. performed the experiments of this study, interpreted the data and wrote the manuscript; M.Y. designed the experiments of this study and edited the manuscript; T.H. and H.F. collected the CAFs of the malignant melanoma specimen; H.M. helped draft the manuscript.

  • Conflicts of Interest

    There are no financial or other interests concerning the submitted manuscript that might be construed as conflicts of interest.

  • Received November 16, 2022.
  • Revision received November 29, 2022.
  • Accepted November 30, 2022.

This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY-NC-ND) 4.0 international license (https://creativecommons.org/licenses/by-nc-nd/4.0).

References

  1. ↵
    1. Leiter U,
    2. Eigentler T and
    3. Garbe C
    : Epidemiology of skin cancer. Adv Exp Med Biol 810: 120-140, 2014. PMID: 25207363. DOI: 10.1007/978-1-4939-0437-2_7
    OpenUrlCrossRefPubMed
  2. ↵
    1. Abbas O,
    2. Miller DD and
    3. Bhawan J
    : Cutaneous malignant melanoma: update on diagnostic and prognostic biomarkers. Am J Dermatopathol 36(5): 363-379, 2014. PMID: 24803061. DOI: 10.1097/DAD.0b013e31828a2ec5
    OpenUrlCrossRefPubMed
  3. ↵
    1. Balch CM,
    2. Soong SJ,
    3. Gershenwald JE,
    4. Thompson JF,
    5. Reintgen DS,
    6. Cascinelli N,
    7. Urist M,
    8. McMasters KM,
    9. Ross MI,
    10. Kirkwood JM,
    11. Atkins MB,
    12. Thompson JA,
    13. Coit DG,
    14. Byrd D,
    15. Desmond R,
    16. Zhang Y,
    17. Liu PY,
    18. Lyman GH and
    19. Morabito A
    : Prognostic factors analysis of 17,600 melanoma patients: validation of the American Joint Committee on Cancer melanoma staging system. J Clin Oncol 19(16): 3622-3634, 2001. PMID: 11504744. DOI: 10.1200/JCO.2001.19.16.3622
    OpenUrlAbstract/FREE Full Text
  4. ↵
    1. Yang X,
    2. Li Y,
    3. Zou L and
    4. Zhu Z
    : Role of exosomes in crosstalk between cancer-associated fibroblasts and cancer cells. Front Oncol 9: 356, 2019. PMID: 31131261. DOI: 10.3389/fonc.2019.00356
    OpenUrlCrossRefPubMed
    1. Sun LP,
    2. Xu K,
    3. Cui J,
    4. Yuan DY,
    5. Zou B,
    6. Li J,
    7. Liu JL,
    8. Li KY,
    9. Meng Z and
    10. Zhang B
    : Cancer associated fibroblast derived exosomal miR 382 5p promotes the migration and invasion of oral squamous cell carcinoma. Oncol Rep 42(4): 1319-1328, 2019. PMID: 31364748. DOI: 10.3892/or.2019.7255
    OpenUrlCrossRefPubMed
    1. Li C,
    2. Teixeira AF,
    3. Zhu HJ and
    4. Ten Dijke P
    : Cancer associated-fibroblast-derived exosomes in cancer progression. Mol Cancer 20(1): 154, 2021. PMID: 34852849. DOI: 10.1186/s12943-021-01463-y
    OpenUrlCrossRefPubMed
  7. ↵
    1. Goulet CR,
    2. Champagne A,
    3. Bernard G,
    4. Vandal D,
    5. Chabaud S,
    6. Pouliot F and
    7. Bolduc S
    : Cancer-associated fibroblasts induce epithelial-mesenchymal transition of bladder cancer cells through paracrine IL-6 signalling. BMC Cancer 19(1): 137, 2019. PMID: 30744595. DOI: 10.1186/s12885-019-5353-6
    OpenUrlCrossRefPubMed
  8. ↵
    1. Fuyuhiro Y,
    2. Yashiro M,
    3. Noda S,
    4. Matsuoka J,
    5. Hasegawa T,
    6. Kato Y,
    7. Sawada T and
    8. Hirakawa K
    : Cancer-associated orthotopic myofibroblasts stimulates the motility of gastric carcinoma cells. Cancer Sci 103(4): 797-805, 2012. PMID: 22320235. DOI: 10.1111/j.1349-7006.2012.02209.x
    OpenUrlCrossRefPubMed
    1. Hasegawa T,
    2. Yashiro M,
    3. Nishii T,
    4. Matsuoka J,
    5. Fuyuhiro Y,
    6. Morisaki T,
    7. Fukuoka T,
    8. Shimizu K,
    9. Shimizu T,
    10. Miwa A and
    11. Hirakawa K
    : Cancer-associated fibroblasts might sustain the stemness of scirrhous gastric cancer cells via transforming growth factor-β signaling. Int J Cancer 134(8): 1785-1795, 2014. PMID: 24155219. DOI: 10.1002/ijc.28520
    OpenUrlCrossRefPubMed
    1. Kasashima H,
    2. Yashiro M,
    3. Kinoshita H,
    4. Fukuoka T,
    5. Morisaki T,
    6. Masuda G,
    7. Sakurai K,
    8. Kubo N,
    9. Ohira M and
    10. Hirakawa K
    : Lysyl oxidase-like 2 (LOXL2) from stromal fibroblasts stimulates the progression of gastric cancer. Cancer Lett 354(2): 438-446, 2014. PMID: 25128648. DOI: 10.1016/j.canlet.2014.08.014
    OpenUrlCrossRefPubMed
    1. Hatano T,
    2. Yashiro M,
    3. Fujikawa H and
    4. Motomura H
    : C-X-C Motif Ligand 1 (CXCL1) from melanoma cells down-regulates the invasion of their metastatic melanoma cells. Oncotarget 9(57): 31090-31097, 2018. PMID: 30123429. DOI: 10.18632/oncotarget.25783
    OpenUrlCrossRefPubMed
  12. ↵
    1. Shelton M,
    2. Anene CA,
    3. Nsengimana J,
    4. Roberts W,
    5. Newton-Bishop J and
    6. Boyne JR
    : The role of CAF derived exosomal microRNAs in the tumour microenvironment of melanoma. Biochim Biophys Acta Rev Cancer 1875(1): 188456, 2021. PMID: 33153973. DOI: 10.1016/j.bbcan.2020.188456
    OpenUrlCrossRefPubMed
  13. ↵
    1. Mathivanan S,
    2. Fahner CJ,
    3. Reid GE and
    4. Simpson RJ
    : ExoCarta 2012: database of exosomal proteins, RNA and lipids. Nucleic Acids Res 40(Database issue): D1241-D1244, 2012. PMID: 21989406. DOI: 10.1093/nar/gkr828
    OpenUrlCrossRefPubMed
  14. ↵
    1. Yuana Y,
    2. Sturk A and
    3. Nieuwland R
    : Extracellular vesicles in physiological and pathological conditions. Blood Rev 27(1): 31-39, 2013. PMID: 23261067. DOI: 10.1016/j.blre.2012.12.002
    OpenUrlCrossRefPubMed
    1. Costa-Silva B,
    2. Aiello NM,
    3. Ocean AJ,
    4. Singh S,
    5. Zhang H,
    6. Thakur BK,
    7. Becker A,
    8. Hoshino A,
    9. Mark MT,
    10. Molina H,
    11. Xiang J,
    12. Zhang T,
    13. Theilen TM,
    14. García-Santos G,
    15. Williams C,
    16. Ararso Y,
    17. Huang Y,
    18. Rodrigues G,
    19. Shen TL,
    20. Labori KJ,
    21. Lothe IM,
    22. Kure EH,
    23. Hernandez J,
    24. Doussot A,
    25. Ebbesen SH,
    26. Grandgenett PM,
    27. Hollingsworth MA,
    28. Jain M,
    29. Mallya K,
    30. Batra SK,
    31. Jarnagin WR,
    32. Schwartz RE,
    33. Matei I,
    34. Peinado H,
    35. Stanger BZ,
    36. Bromberg J and
    37. Lyden D
    : Pancreatic cancer exosomes initiate pre-metastatic niche formation in the liver. Nat Cell Biol 17(6): 816-826, 2015. PMID: 25985394. DOI: 10.1038/ncb3169
    OpenUrlCrossRefPubMed
  16. ↵
    1. Miki Y,
    2. Yashiro M,
    3. Okuno T,
    4. Kitayama K,
    5. Masuda G,
    6. Hirakawa K and
    7. Ohira M
    : CD9-positive exosomes from cancer-associated fibroblasts stimulate the migration ability of scirrhous-type gastric cancer cells. Br J Cancer 118(6): 867-877, 2018. PMID: 29438363. DOI: 10.1038/bjc.2017.487
    OpenUrlCrossRefPubMed
  17. ↵
    1. Kinoshita H,
    2. Yashiro M,
    3. Fukuoka T,
    4. Hasegawa T,
    5. Morisaki T,
    6. Kasashima H,
    7. Masuda G,
    8. Noda S and
    9. Hirakawa K
    : Diffuse-type gastric cancer cells switch their driver pathways from FGFR2 signaling to SDF1/CXCR4 axis in hypoxic tumor microenvironments. Carcinogenesis 36(12): 1511-1520, 2015. PMID: 26385890. DOI: 10.1093/carcin/bgv134
    OpenUrlCrossRefPubMed
  18. ↵
    1. Hoshino A,
    2. Costa-Silva B,
    3. Shen TL,
    4. Rodrigues G,
    5. Hashimoto A,
    6. Tesic Mark M,
    7. Molina H,
    8. Kohsaka S,
    9. Di Giannatale A,
    10. Ceder S,
    11. Singh S,
    12. Williams C,
    13. Soplop N,
    14. Uryu K,
    15. Pharmer L,
    16. King T,
    17. Bojmar L,
    18. Davies AE,
    19. Ararso Y,
    20. Zhang T,
    21. Zhang H,
    22. Hernandez J,
    23. Weiss JM,
    24. Dumont-Cole VD,
    25. Kramer K,
    26. Wexler LH,
    27. Narendran A,
    28. Schwartz GK,
    29. Healey JH,
    30. Sandstrom P,
    31. Labori KJ,
    32. Kure EH,
    33. Grandgenett PM,
    34. Hollingsworth MA,
    35. de Sousa M,
    36. Kaur S,
    37. Jain M,
    38. Mallya K,
    39. Batra SK,
    40. Jarnagin WR,
    41. Brady MS,
    42. Fodstad O,
    43. Muller V,
    44. Pantel K,
    45. Minn AJ,
    46. Bissell MJ,
    47. Garcia BA,
    48. Kang Y,
    49. Rajasekhar VK,
    50. Ghajar CM,
    51. Matei I,
    52. Peinado H,
    53. Bromberg J and
    54. Lyden D
    : Tumour exosome integrins determine organotropic metastasis. Nature 527(7578): 329-335, 2015. PMID: 26524530. DOI: 10.1038/nature15756
    OpenUrlCrossRefPubMed
  19. ↵
    1. Yoshioka Y,
    2. Kosaka N,
    3. Konishi Y,
    4. Ohta H,
    5. Okamoto H,
    6. Sonoda H,
    7. Nonaka R,
    8. Yamamoto H,
    9. Ishii H,
    10. Mori M,
    11. Furuta K,
    12. Nakajima T,
    13. Hayashi H,
    14. Sugisaki H,
    15. Higashimoto H,
    16. Kato T,
    17. Takeshita F and
    18. Ochiya T
    : Ultra-sensitive liquid biopsy of circulating extracellular vesicles using ExoScreen. Nat Commun 5: 3591, 2014. PMID: 24710016. DOI: 10.1038/ncomms4591
    OpenUrlCrossRefPubMed
  20. ↵
    1. Yoshioka Y,
    2. Konishi Y,
    3. Kosaka N,
    4. Katsuda T,
    5. Kato T and
    6. Ochiya T
    : Comparative marker analysis of extracellular vesicles in different human cancer types. J Extracell Vesicles 2: 2013. PMID: 24009892. DOI: 10.3402/jev.v2i0.20424
    OpenUrlCrossRefPubMed
  21. ↵
    1. Fan J,
    2. Zhu GZ and
    3. Niles RM
    : Expression and function of CD9 in melanoma cells. Mol Carcinog 49(1): 85-93, 2010. PMID: 19777564. DOI: 10.1002/mc.20580
    OpenUrlCrossRefPubMed
  22. ↵
    1. Si Z and
    2. Hersey P
    : Expression of the neuroglandular antigen and analogues in melanoma. CD9 expression appears inversely related to metastatic potential of melanoma. Int J Cancer 54(1): 37-43, 1993. PMID: 8478146. DOI: 10.1002/ijc.2910540107
    OpenUrlCrossRefPubMed
  23. ↵
    1. Murayama Y,
    2. Oritani K and
    3. Tsutsui S
    : Novel CD9-targeted therapies in gastric cancer. World J Gastroenterol 21(11): 3206-3213, 2015. PMID: 25805926. DOI: 10.3748/wjg.v21.i11.3206
    OpenUrlCrossRefPubMed
  24. ↵
    1. Brosseau C,
    2. Colas L,
    3. Magnan A and
    4. Brouard S
    : CD9 Tetraspanin: A new pathway for the regulation of inflammation? Front Immunol 9: 2316, 2018. PMID: 30356731. DOI: 10.3389/fimmu.2018.02316
    OpenUrlCrossRefPubMed
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CD9-positive Exosomes Derived from Cancer-associated Fibroblasts Might Inhibit the Proliferation of Malignant Melanoma Cells
NAHO FUJII, MASAKAZU YASHIRO, TAKAHARU HATANO, HEISHIRO FUJIKAWA, HISASHI MOTOMURA
Anticancer Research Jan 2023, 43 (1) 25-33; DOI: 10.21873/anticanres.16130
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