Hymecromone Research Articles

Abstract 3283: Role of HA, HAS3, UGT1A9 in metastatic renal cell carcinoma and sorafenib resistance

Abstract Introduction: Despite numerous available treatments, metastatic renal cell carcinoma (mRCC) remains highly lethal, with over 90% of patients succumbing within 5 years. Sorafenib (SF), a second-line treatment for mRCC, exhibits modest efficacy, and the underlying molecular basis for its failure is elusive. Hyaluronic acid (HA), implicated in promoting cell proliferation, invasion, and motility through CD44 and RHAMM receptors, represents a potential player in SF resistance. Hymecromone (HC), a non-toxic dietary supplement, is known to inhibit HA synthesis and downregulate CD44 and RHAMM mRNA levels. This study investigates the elusive mechanisms of SF resistance and evaluates the effectiveness of the HC+SF combination as a targeted therapy for advanced RCC, providing insights into a novel and potentially impactful treatment approach. Methods: HA levels were measured in normal and tumor kidney specimens using ELISA-like assay and IHC. Similarly, HAS3 and UGT1A9 (A9) levels and SF glucuronidation were measured by qPCR and a glucuronidation assay using tumor microsomes. Response to SF+HC combination was examined in A9 and HAS3 knockdown and overexpression cell lines (transfectants) and primary RCC spheroids by proliferation, apoptosis, motility, and invasion assays. Target analysis was performed by immunoblot. Treatment efficacy was evaluated in subcutaneous and orthotopic xenograft mouse models, with bioluminescence imaging. Results: SF failure in mRCC was linked due to its glucuronidation and inactivation by A9. Tumor microsomes from patients who developed mRCC glucuronidated SF 5-fold higher than from non-mRCC patients. A9 levels were elevated (8-10 folds) in mRCC patients’ tumor along with HA and HAS3 (5-10 folds) as compared to non-mRCC patients and normal kidney tissues. HC treatment downregulated A9 promoter activity and inhibited SF-glucuronidation in RCC cells. SF (5uM) and HC (20, 40ug/ml) combination inhibited RCC cell and primary tumor spheroid proliferation (>90%), motility/invasion (>80%), and induced apoptosis (5-fold) and downregulated HAS3 expression (>80%), and HA synthesis in RCC cells. While A9 and HAS3 transfectants both were resistant to SF+HC treatment, A9 and HAS3 - KO transfectants were sensitive to SF. Endothelial cells co-cultured with either A9 or HAS3 transfectants were resistant to SF+HC treatment. SF+HC upregulated apoptosis effectors, but downregulated CD44, RHAMM, and phospho-Met levels in EV but not in A9 or HAS3 transfectants. HC (100 or 200mg/kg) and SF (30mg/kg) oral combination abrogated Caki-1 tumor growth (>80%), without serum/tissue toxicity. HC+SF treatment decreased A9, HA and HAS3 levels, Ki67 index and microvessel density. Both A9 and HAS3 overexpressing tumors were resistant to treatment. Conclusion: This study demonstrates the mechanism of SF failure in mRCC and proposes a safe, oral adjunct combination for effective treatment. Citation Format: Anuj Kumar Sharma, Karina Aguilar, Chandramukhi S. Panda, Vinata B. Lokeshwar. Role of HA, HAS3, UGT1A9 in metastatic renal cell carcinoma and sorafenib resistance [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 3283.

Abstract 2065: Biomarker-targeted novel combination for renal cell carcinoma

Abstract INTRODUCTION: The survival of patients with metastatic renal cell carcinoma (mRCC) is < 10% at 5-years, despite treatment. Sorafenib (SF) is a second-line oral treatment which improves overall survival by 12-18%. However, mechanism for SF failure is not known. Hyaluronic acid (HA) promotes cell proliferation, invasion and motility through its receptors CD44 and RHAMM. Hymecromone (HC) is a non−toxic dietary supplement that inhibits HA synthesis and down regulates CD44 and RHAMM mRNA levels. In this study we examined the mechanism of SF resistance and the efficacy of HC+SF combination as a targeted therapy for advanced RCC. METHODS: HAS3 levels were measured in kidney specimens (normal=45; tumor= 86) by qPCR. HAS3 promoter was cloned and effect of SF and HC on HAS3 promoter activity was determined. Response of Vector-only (EV) or HAS3-overexpressing (HAS3) or HAS3-knockdown (HAS3-KO) transfectants of 786-O and Caki-1 cells, primary RCC spheroids and endothelial cells to SF+HC combination was examined in proliferation, apoptosis, motility and invasion assays. Immunoblotting was performed for target analysis. Treatment efficacy was evaluated in the Caki−1 subcutaneous (s.c.) and orthotopic xenograft models. RESULTS: HAS3 levels were 5-10-fold elevated in mRCC patients’ tumors as compared to non-mRCC patients and normal kidney tissues, respectively. HAS3 levels were an independent predictor of metastasis (P=0.043; sensitivity: 85.7%). Combination of SF (5 µM) with HC (20, 40 µg/mL) downregulated (>80%) HAS3 expression, HAS3 promoter activity and HA synthesis in RCC cells. HC+SF combination inhibited RCC cell and primary tumor spheroid proliferation > 90%, motility/invasion (>80%) and induced apoptosis (5-fold). While HAS3 transfectants were resistant to SF+HC treatment, HAS3-KO transfectants were sensitive to SF. HA addition also attenuated the effect of HC+SF observed in EV cells. Endothelial cells co-cultured with HAS3 transfectants were resistant to SF+HC treatment. SF+HC up-regulated apoptosis effectors, but downregulated CD44, RHAMM, and phospho-Met levels in EV but not in HAS3 transfectants. HC (100 or 200 mg/kg) and SF (30 mg/kg) oral combination abrogated Caki-1 tumor growth (>80%), without serum/tissue toxicity. HC+SF treatment decreased HAS3/HA levels, Ki67 index and microvessel density. HAS-overexpressing tumors were resistant to treatment. CONCLUSION: This is the first study to demonstrate that HAS3 overexpression in mRCC contributes to SF failure and that HAS3/HA downregulation is a mechanism to re-sensitize cells to SF. HC+SF is an orally bioavailable and non-toxic combination that may be an effective therapeutic strategy for mRCC. Citation Format: Jiaojiao Wang, Andre R. Jordan, Sarrah S. Lahorewala, Vinata B. Lokeshwar. Biomarker-targeted novel combination for renal cell carcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 2065.

Abstract 2080: Novel molecularly targeted combination for renal cell carcinoma

Abstract INTRODUCTION: Despite many available treatments, >90% of metastatic renal cell carcinoma (mRCC) patients die within 5 years. Due to its modest efficacy, Sorafenib (SF) is a second line treatment for mRCC. Molecular basis for SF failure is unknown. Hymecromone (HC) is a non−toxic orally bioavailable agent used in Europe for improving liver health. We examined the molecular basis of SF failure in clinical specimens and provided mechanistic evidence for why SF+HC could be a potent therapeutic combination against mRCC. METHODS: UGT1A9 (A9) levels and SF glucuronidation were measured in kidney specimens (45 normal; 86 tumor) by qPCR and a glucuronidation assay using tumor microsomes. Response to SF+HC combination in vector-only (EV), A9-overexpressing (A9) and A9-knockdown (A9-KO) 786-0 and Caki-1 transfectants, primary RCC spheroids, and endothelial cells was examined by proliferation, apoptosis, motility and invasion assays. Target analysis was performed by immunoblot. Treatment efficacy was evaluated in Caki−1 s.c. and orthotopic xenograft models, with bioluminescence imaging. RESULTS: A9 has been shown to glucuronidate and inactivate SF. A9 levels were 8-10-fold elevated in mRCC patients’ tumors compared to non-mRCC patients and normal kidney tissues. A9 levels were an independent predictor of metastasis (P=0.022; efficacy: 73%; sensitivity: 93.8%). Tumor microsomes from mRCC patients’ glucuronidated SF 5-fold higher than from non-mRCC patients. In RCC cells, HC treatment downregulated A9 levels, A9 promoter activity and inhibited SF-glucuronidation. SF (5 µM) and HC (20-40 µg/ml) combination inhibited RCC cell and primary tumor spheroid proliferation (>90%), motility/invasion (>80%), and induced apoptosis (5-fold). While A9 transfectants were resistant to SF+HC treatment, A9-KO cells were sensitive to SF alone. Endothelial cells co-cultured with A9 transfectants were resistant to SF+HC treatment. SF+HC upregulated apoptosis effectors, but downregulated CD44, RHAMM, and phospho-Met levels in EV but not A9 transfectants. HC (100 or 200 mg/kg) and SF (30 mg/kg) oral combination abrogated Caki−1 tumor growth (>80%) without serum/tissue toxicity. HC+SF treatment decreased A9 levels, Ki67 index and microvessel density. A9-overexpressing tumors were resistant to treatment. CONCLUSION: This is the first study to demonstrate that A9 overexpression in mRCC is likely a mechanism of SF failure and A9 downregulation can re-sensitize cells. HC+SF is an orally bioavailable and non-toxic combination that may be an effective therapeutic strategy for mRCC. Citation Format: Andre R. Jordan, Jiaojiao Wang, Luis E. Lopez, Daley S. Morera, Vinata B. Lokeshwar. Novel molecularly targeted combination for renal cell carcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 2080.

MP29-14 TARGETING HYALURONIC ACID FAMILY BIOMARKERS FOR TREATMENT OF RENAL CELL CARCINOMA

You have accessJournal of UrologyKidney Cancer: Basic Research II1 Apr 2014MP29-14 TARGETING HYALURONIC ACID FAMILY BIOMARKERS FOR TREATMENT OF RENAL CELL CARCINOMA Nicolas Ortiz, Michael Garcia-Roig, Travis Yates, Murugesan Manoharan, and Vinata Lokeshwar Nicolas OrtizNicolas Ortiz More articles by this author , Michael Garcia-RoigMichael Garcia-Roig More articles by this author , Travis YatesTravis Yates More articles by this author , Murugesan ManoharanMurugesan Manoharan More articles by this author , and Vinata LokeshwarVinata Lokeshwar More articles by this author View All Author Informationhttps://doi.org/10.1016/j.juro.2014.02.756AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookTwitterLinked InEmail INTRODUCTION AND OBJECTIVES Up to 25% of patients with renal cell carcinoma (RCC) have metastasis at initial diagnosis. Current targeted therapies (e.g., Sorafenib) are not curative and the 5-year survival of these patients is <10%. Molecular determinants of RCC growth and metastasis may be accurate prognostic markers for metastasis and could be targeted for therapy. We evaluated the expression of 3 genes (and their splice variants) as prognostic indicators. We also tested the therapeutic efficacy of the combination of hymecromone (HC), a non-toxic, orally bioavailable hyaluronic acid (HA) synthesis inhibitor and Sorafenib (SF). METHODS Tumor specimens were collected from RCC patients undergoing nephrectomy (n=81; T1, 40; T2+, 41; Fuhrman Grade 1, 5; Grade 2, 32; Grade 3, 25; Grade 4, 19). Nine patients developed metastasis within 15 months. Tissue-RNA was subjected to Q-PCR for HA family of markers (i.e. HAS3, CD44, RHAMM). mRNA levels were correlated with metastasis by logistic regression. Effect of HC, SF and HC+SF on cell proliferation, cell cycle and apoptosis was examined in RCC cells (Caki-1, 786-O, ACHN, 769-P) by cell counting, flow cytometry and Cell Death ELISA. Boyden chamber was used for motility assay. Effect on cell cycle, apoptosis, and HA-receptor levels was evaluated by immunoblotting and Q-PCR. RESULTS The mRNA levels of HAS3, CD44 and RHAMM were 3 to 7-fold elevated in RCC tissues from patients who developed metastasis when compared to those who did not develop metastasis during the follow up period. In univariate analysis age, grade, stage, CD44 and RHAMM levels significantly correlated with metastasis. In multivariate analysis, CD44 (< 0.0001), HAS3 (P=0.03) and RHAMM (P< 0.0001) significantly correlated with metastasis. Among the three markers, RHAMM had the highest accuracy (89.5%) in predicting metastasis. Combination of HC (15-30-μg/ml) and SF (1.6 to 3.2 μg/ml), at non-cytotoxic doses caused 60-90% inhibition in proliferation, motility and invasion after 48-72 h treatment. Only the combination induced apoptosis (3-4-fold). HC+SF combination down regulated CD44 and RHAMM mRNA levels by 2- and 68-fold, respectively and inhibited HA synthesis. In Caki-1xenograft, oral treatment with combination caused 100% inhibition of tumor growth. CONCLUSIONS HAS3, CD44 and RHAMM expression in RCC tissues is potentially an independent prognostic indicator for metastasis. HC+SF combination may be a potentially effective treatment for metastatic RCC. © 2014FiguresReferencesRelatedDetails Volume 191Issue 4SApril 2014Page: e310 Advertisement Copyright & Permissions© 2014MetricsAuthor Information Nicolas Ortiz More articles by this author Michael Garcia-Roig More articles by this author Travis Yates More articles by this author Murugesan Manoharan More articles by this author Vinata Lokeshwar More articles by this author Expand All Advertisement Advertisement PDF downloadLoading ...

168 DIETARY SUPPLEMENT HYMECROMONE BASED COMBINATION FOR RENAL CELL CARCINOMA

You have accessJournal of UrologyKidney Cancer: Basic Research (I)1 Apr 2013168 DIETARY SUPPLEMENT HYMECROMONE BASED COMBINATION FOR RENAL CELL CARCINOMA Nicolas Ortiz, Anaid Benitez, Travis Yates, and Vinata Lokeshwar Nicolas OrtizNicolas Ortiz Miami, FL More articles by this author , Anaid BenitezAnaid Benitez Miami, FL More articles by this author , Travis YatesTravis Yates Miami, FL More articles by this author , and Vinata LokeshwarVinata Lokeshwar Miami, FL More articles by this author View All Author Informationhttps://doi.org/10.1016/j.juro.2013.02.1548AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookTwitterLinked InEmail INTRODUCTION AND OBJECTIVES Approximately 17% of patients with renal cell carcinoma (RCC) present with metastasis upon diagnosis, rendering patients with a 5-year survival rate of 12%. Sorafenib (SF) was the first targeted therapy approved for RCC treatment, however it has modest efficacy. Hyaluronic acid (HA) is a non-sulfated glycosaminoglycan that promotes cell proliferation, invasion and motility through membrane receptors CD44 and RHAMM. High HA expression has been correlated with cancer progression and inflammatory diseases. Hymecromone (HC) is a non−toxic dietary supplement that inhibits HA synthesis and down regulates CD44 and RHAMM mRNA. In this study we examined whether HC, SF, or their combination inhibit RCC progression. METHODS Effect of HC, SF and HC+SF on cell proliferation, cell cycle and apoptosis was examined in RCC cells (Caki−1, 786−O) and endothelial cells (HMVEC−L, HUVEC) by cell-counting, flow cytometry and Cell Death ELISA. Boyden chamber was used for motility and invasion assays. Effect on cell cycle, apoptosis, and HA receptor levels were evaluated by immunoblotting and Q−PCR. Treatment efficacy was evaluated in the Caki−1 xenograft model. RESULTS While HC (0−30 ug/ml) and SF (0−3.2 ug/ml) individually were ineffective, their combination inhibited proliferation (>95%), motility (≥60%) and invasion (≥50%) in both RCC and endothelial cells. Combination of HC+ SF induced apoptosis (>8−fold) and caused cell cycle arrest. Consistent with these results, downregulation of Cyclins A, E1, D1, phospho−cdk2, Rb, STAT3, Mcl−1 and induced PARP cleavage, and Caspase 3 activation was also seen in combination treatment. HC+SF synergistically decreased the expression of HA receptors CD44 and RHAMM at the mRNA and protein levels. The oral administration of both HC (50−100mg/kg) and SF (30mg/kg) eradicated SF-resistant Caki−1 tumors whereas individual drugs were ineffective. No serum or tissue toxicity was observed in the treated groups. CONCLUSIONS Our findings show that a novel combination of SF with HC, a non−toxic oral supplement is potentially an effective treatment strategy for metastatic RCC. © 2013 by American Urological Association Education and Research, Inc.FiguresReferencesRelatedDetails Volume 189Issue 4SApril 2013Page: e69 Advertisement Copyright & Permissions© 2013 by American Urological Association Education and Research, Inc.MetricsAuthor Information Nicolas Ortiz Miami, FL More articles by this author Anaid Benitez Miami, FL More articles by this author Travis Yates Miami, FL More articles by this author Vinata Lokeshwar Miami, FL More articles by this author Expand All Advertisement Advertisement PDF DownloadLoading ...

99mTc-EHIDA hepatobiliary scintingraphy and treatment of obstructive jaundice in infants

Objective To discuss the diagnosis,differential diagnosis and treatment of obstructive jaundice in infants.Methods Infants with obstructive jaundice were examined by ECT to obtain dynamic images of the liver,biliary tract and intestine to find out the position and nature of the obstruction.Results Eight patients with delayed nuclide imagining were diagnosed as obstructive jaundice of cholestasis.Five of which with aggravated jaundice were cured by biliary irrigation and 3 by medication.For 16 patients with congenital biliary atresia,10 were exterior hepatic types and 6 interior hepatic types.They had the nuclide image of liver and biliary tract.Ten cases of the exterior hepatic types and four cases of the interior types received the Kasai operation.And bile was drained after the surgery.Seven patients survived and 4 of interior heaptic types died postoperatively.Due to bile fistula,3 patients died after the hepato-porto-jejunostomy for internal drainage.Two patients of interior hepatic types who had no operation died of the hepatic failure.Conclusion The nature and position of obstructive jaundice could be identified by nuclide imaging.Young patients with jaundice of cholestasis could be treated by antiinflamation,liver function protection,Hymecromone and Dexamethasone for two weeks.Once the serum bilirubin rises,operation would be indicated.In the case of congenital biliary atresia,operation should be done earlier on the basis of medication. Key words: Obstructive jaundice; Biliary atresia; 99mTc-EhIDA; Diagnosis