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1.
Dis Colon Rectum ; 2024 May 03.
Article in English | MEDLINE | ID: mdl-38701431

ABSTRACT

BACKGROUND: Creation of a tension free colorectal anastomosis after left colon resection or low anterior resection is a key requirement for technical success. The relative contribution of each of a series of known lengthening maneuvers remains incompletely characterized. OBJECTIVE: The aim of this study was to compare technical procedures for lengthening of the left colon prior to rectal anastomosis. DESIGN: A series of lengthening maneuvers was performed on 15 fresh cadavers. Average distance gained was measured for each successive maneuver, including (1) high inferior mesenteric artery ligation, (2) splenic flexure takedown, and (3) high inferior mesenteric vein ligation by the ligament of Treitz. SETTING: Cadaveric study. MAIN OUTCOME MEASURES: The pre-mobilization and post-mobilization position of the proximal colonic end was measured relative to the inferior edge of the sacral promontory. Measurements of the colonic length relative to the sacral promontory were taken following each mobilization maneuver. The inferior mesenteric artery, sigmoid colon and rectum specimen lengths were measured. The distance from the inferior border of the sacral promontory to the pelvic floor was measured along the sacral curvature. RESULTS: Average sigmoid colon resection length was 34.7 ± 11.1 cm. Prior to any lengthening, baseline reach was -1.3 ± 4.2 cm from the sacral promontory. Inferior mesenteric artery ligation yielded an additional 11.5 ± 4.7 cm. Subsequent splenic flexure takedown added an additional 12.8 ± 9.6 cm. Finally, inferior mesenteric vein ligation added an additional 11.33 ± 6.9 cm, bringing the total colonic length to 35.7 ± 14.7 cm. BMI and weight negatively correlated with length gained. LIMITATIONS: The study was limited by nature of being a cadaver study. CONCLUSIONS: Stepwise lengthening maneuvers allow significant additional reach to allow a tension free left colon to rectal anastomosis. See Video Abstract.

2.
Am Surg ; : 31348241256055, 2024 May 21.
Article in English | MEDLINE | ID: mdl-38770756

ABSTRACT

INTRODUCTION: Total neoadjuvant therapy (TNT) for patients with locally advanced rectal cancer (LARC) is now the standard of care. Randomized trials suggest the use of short-course radiotherapy (SCRT) and long-course radiotherapy (LCRT) are oncologically equivalent. OBJECTIVE: To describe pathologic outcomes after surgical resections of patients receiving SCRT versus LCRT as part of TNT for LARC. PARTICIPANTS: All patients with LARC treated at a single tertiary hospital who underwent proctectomy after completing TNT were included. Patients were excluded if adequate details of TNT were not available in the electronic medical record. RESULTS: A total of 53 patients with LARC were included. Thirty-nine patients (73.5%) received LCRT and 14 (26.4%) received SCRT. Forty-nine patients (92.5%) were clinical stage III (cN1-2) prior to treatment. The average lymph node yield after proctectomy was 20.9 for SCRT and 17.0 for LCRT (P = .075). Of the 49 patients with clinically positive nodes before treatment, 76.9% of those who received SCRT and 72.2% of those who received LCRT achieved pN0 disease after TNT. Additionally, there were no significant differences in rates of pathologic complete response between patients who received SCRT and LCRT, 7.1% and 12.8%, respectively (P = .565). CONCLUSION: Pathologic outcomes of patients with LARC treated with SCRT or LCRT, as part of TNT, may be similar. Further prospective trials are needed to assess long-term clinical outcomes and to determine best treatment protocols.

3.
J Surg Res ; 281: 37-44, 2023 01.
Article in English | MEDLINE | ID: mdl-36115147

ABSTRACT

INTRODUCTION: Preoperative endoscopic tattooing is an effective tool for intraoperative tumor localization in colon cancer. Endoscopic tattooing in rectal cancer may have unidentified benefits on lymph node yield, making it easier for pathologists to identify nodes during histopathologic assessment. There remains concern that tattoo ink may alter anatomical planes, increasing surgical difficulty. METHODS: Retrospective chart reviews from 2016 to 2021 of n = 170 patients presenting with rectal cancer were divided into two groups: with (n = 79) and without (n = 91) endoscopic tattoos. Demographics, operative details, tumor characteristics, prior chemoradiation, and pathologic details were collected. Primary outcome was total lymph node yield. Secondary outcomes were rates of adequate (> 12) nodes, margin status, and operative variables including operative time. RESULTS: No differences between pathologic stage, tumor height, high inferior mesenteric artery ligation, operative times, conversion rate, or surgical approach (open versus minimally invasive) were noted between groups. Receipt of neoadjuvant chemoradiation was less frequent in the endoscopic tattooing group (53.2% versus 76.9%, P ≤ 0.001). Total node number and rate of adequate lymph node yield were higher with endoscopic tattooing (20.5 ± 7.6 versus 16.8 ± 6.6 lymph nodes and 100.0% versus 83.5% adequate lymph node harvest, both P ≤ 0.001). Rates of positive circumferential and distal margins and complete total mesorectal excision were also similar. Regression analysis identified endoscopic tattooing (Incidence Risk Ratio 1.17, 95% confidence interval 1.04-1.31) and operative time more than 300 min (Incidence Risk Ratio 0.88, 95% confidence interval 0.77-0.99) had significant effects on lymph node harvest. Removal of patients with inadequate lymph node yield resulted in similar rates of total and positive lymph nodes. CONCLUSIONS: Endoscopic rectal tattooing is associated with increased lymph node yield (including after neoadjuvant chemoradiotherapy) without sacrificing oncologic or perioperative outcomes, although this effect is inconsistent when only considering patients with an adequate lymph node yield.


Subject(s)
Rectal Neoplasms , Tattooing , Humans , Tattooing/methods , Lymph Node Excision/methods , Retrospective Studies , Rectal Neoplasms/surgery , Rectal Neoplasms/pathology , Lymph Nodes/surgery , Lymph Nodes/pathology , Neoadjuvant Therapy , Neoplasm Staging
4.
Surg Endosc ; 36(3): 2121-2128, 2022 03.
Article in English | MEDLINE | ID: mdl-33890178

ABSTRACT

PURPOSE: "Endoscopically unresectable" benign polyps identified during screening colonoscopy are often referred for segmental colectomy. Application of advanced endoscopic techniques can increase endoscopic polyp resection, sparing patients the morbidity of colectomy. This retrospective case-control study aimed to evaluate the success of colon preserving resection of "endoscopically unresectable" benign polyps using advanced endoscopic techniques including endoscopic mucosal resection, endoscopic submucosal dissection, endoluminal surgical intervention, full-thickness laparo-endoscopic excision, and combined endo-laparoscopic resection. METHODS: A prospectively maintained institutional database identified 95 patients referred for "endoscopically unresectable" benign polyps from 2015 to 2018. Cases were compared to 190 propensity score matched controls from the same database undergoing elective laparoscopic colectomy for other reasons. Primary outcome was rate of complete endoscopic polyp removal. Secondary outcomes included length of stay, unplanned 30-day readmission and reoperation, 30-day mortality, and post-procedural complications. RESULTS: Advanced endoscopic techniques achieved complete polyp removal without colectomy in 66 patients (70%). Failure was most commonly associated with previously attempted endoscopic resection and occult malignancy. Compared with matched colectomy controls, endoscopic polyp resection resulted in significantly shorter hospital length of stay (1.13 ± 2.41 vs 3.89 ± 4.57 days; p < 0.001), lower unplanned 30-day readmission (1.1% vs 7.7%; p < 0.05), and fewer postoperative complications (4.2% vs 33.9%; p < 0.001). Unplanned 30-day reoperation (2.1% vs 4.4%; p = 0.34) and 30-day mortality (0% vs 0.6%; p = 0.75) trended lower. CONCLUSIONS: Endoscopic resection of complex polyps can be highly successful, and it is associated with favorable outcomes and decreased morbidity when compared with segmental colon resection. Attempting colon preservation using these techniques is warranted.


Subject(s)
Colonic Polyps , Case-Control Studies , Colectomy/methods , Colon/surgery , Colonic Polyps/pathology , Colonic Polyps/surgery , Colonoscopy/methods , Humans , Retrospective Studies
5.
Dis Colon Rectum ; 64(12): 1559-1563, 2021 12 01.
Article in English | MEDLINE | ID: mdl-34596631

ABSTRACT

BACKGROUND: Benign colon polyps are increasingly being detected because of improved colonoscopic screening and early detection of masses on the adenoma-to-carcinoma pathway. Full-thickness laparoendoscopic excision is a colon-preserving technique for endoscopically unresectable polyps consisting of endoscopically guided nonanatomic wedge colectomy. OBJECTIVE: This study aimed to evaluate the safety and success of full-thickness laparoendoscopic excision compared to segmental colectomy for complex polyps not amenable to endoscopic resection. DESIGN: This is a retrospective case-control study. SETTINGS: This study was conducted at a tertiary academic center. PATIENTS: A prospectively maintained institutional database identified 22 patients with benign complex polyps managed with full-thickness laparoendoscopic excision from 2015 to 2020. These patients were compared with 22 propensity score-matched controls from the same database that underwent laparoscopic segmental colectomy. MAIN OUTCOME MEASURES: Primary outcome was inpatient length of stay. Secondary outcomes included operative details and postoperative morbidities. RESULTS: Full-thickness laparoendoscopic excision was successful in all patients. Patients had a median age of 64 years (41-85), and 82% were men. Final pathology revealed complete excision of benign lesions in 20 of 22 patients and adenocarcinoma in 2 of 22. For the adenocarcinomas, 1 patient underwent subsequent elective colectomy without complications, and 1 patient declined surgery. Propensity score matching was successful for age, sex, BMI, ASA score, colon location, and prior abdominal surgery. Compared with controls, cases had significantly shorter operative time (89.5 minutes (46-290) vs 122 minutes (85-200), p = 0.009), length of stay (1 day (0-17) vs 3 days (1-8), p < 0.001), and reduced blood loss (5 mL (2-15) vs 25 mL (10-150), p < 0.001). Thirty-day morbidity (9.1% vs 27.3%, p = 0.240) was not significantly different. An unplanned 30-day reoperation was performed in 1 patient for suspected small-bowel obstruction. There was 1 mortality due to decompensated cirrhosis in the treatment group. LIMITATIONS: This study was limited by its single-institution retrospective design. CONCLUSIONS: Full-thickness laparoendoscopic excision is safe and successful compared with corresponding segmental colectomy for complex polyps. Favorable postoperative outcomes, including decreased operative time, length of stay, and blood loss, make it a useful approach for managing complex polyps throughout the colon.


Subject(s)
Colectomy/methods , Colonic Polyps/surgery , Colonoscopy/methods , Endoscopic Mucosal Resection/methods , Laparoscopy/methods , Adenocarcinoma/diagnosis , Adenocarcinoma/epidemiology , Adenocarcinoma/pathology , Adenocarcinoma/surgery , Adult , Aged , Aged, 80 and over , Blood Loss, Surgical/prevention & control , Case-Control Studies , Colonic Polyps/pathology , Combined Modality Therapy/methods , Early Diagnosis , Female , Humans , Intestinal Obstruction/epidemiology , Intestinal Obstruction/etiology , Intestinal Obstruction/surgery , Length of Stay/trends , Male , Mass Screening/standards , Middle Aged , Morbidity/trends , Operative Time , Postoperative Complications/epidemiology , Propensity Score , Reoperation/statistics & numerical data , Retrospective Studies , Safety
6.
J Surg Oncol ; 124(7): 1091-1097, 2021 Dec.
Article in English | MEDLINE | ID: mdl-34310720

ABSTRACT

BACKGROUND: Mesothelin is a cell surface glycoprotein overexpressed in 28%-58% of colorectal cancer (CRC). We hypothesized that CRC mesothelin expression contributes to peritoneal spread and that it is selectively overexpressed in those with peritoneal metastasis versus distant metastasis. METHODS: This case-controlled study involved mesothelin immunohistochemistry staining of tumor specimens from patients with metastatic CRC/appendiceal cancers between 2017 and 2019. Staining reactivity was graded from trace to 4+ (low ≤1+; high >1+). Staining patterns were characterized on global (focal/patchy/diffuse) and cellular (apical/cytoplasmic) levels. Immunostaining of normal mesothelial cells served as internal control. RESULTS: Thirty-one patients were identified: 11 peritoneal (study) and 20 distant metastasis (control). The control group did not include appendiceal cancers. The study group had greater proportion of high staining reactivity (55% vs. 5%; odds ratio [OR] = 20.4, 95% confidence interval [CI] 1.96-211.8). The study group had more diffuse (36% vs. 0%; OR = 22.2, 95% CI 1.1-465.3) and cytoplasmic staining patterns (73% vs. 28%; OR = 6.9, 95% CI 1.3-37.2). CONCLUSION: Mesothelin expression is higher in CRC/appendiceal cancers with peritoneal metastasis than those with distant metastasis. Immunohistochemistry staining patterns suggestive of propensity towards peritoneal metastasis include diffuse and cytoplasmic staining. Mesothelin may be a potential target for novel treatments of CRC/appendiceal carcinoma with peritoneal involvement.


Subject(s)
Appendiceal Neoplasms/metabolism , Colorectal Neoplasms/metabolism , GPI-Linked Proteins/metabolism , Peritoneal Neoplasms/metabolism , Peritoneal Neoplasms/secondary , Adenocarcinoma/metabolism , Adenocarcinoma/pathology , Adenocarcinoma/secondary , Aged , Appendiceal Neoplasms/pathology , Biomarkers, Tumor/metabolism , Case-Control Studies , Colorectal Neoplasms/pathology , Female , Humans , Immunohistochemistry , Male , Mesothelin , Middle Aged , Prevalence
7.
Clin Colon Rectal Surg ; 33(1): 16-21, 2020 Jan.
Article in English | MEDLINE | ID: mdl-31915421

ABSTRACT

Bleedings from small intestine account for 5% of all gastrointestinal bleeding. With advanced endoscopic tools, such as video capsule endoscopy and deep enteroscopy, accurate diagnosis and treatment is possible in majority of cases with low mortality and morbidity. Nonoperative management includes endoscopic hemostasis and angiographic embolization. Recurrence after initial treatment is relatively common. Surgery is reserved for the cases that are refractory to endoscopic or angiographic treatment, bleeding from tumor or mass lesions, or hemodynamic instability. At the time of surgical exploration, unless the lesion has been marked by endoscopic tattoo or clip, intraoperative enteroscopy is often necessary to localize the lesion.

9.
Proc Natl Acad Sci U S A ; 111(49): 17390-5, 2014 Dec 09.
Article in English | MEDLINE | ID: mdl-25422433

ABSTRACT

In this paper we discuss the transformation of a sheet of material into a wide range of desired shapes and patterns by introducing a set of simple cuts in a multilevel hierarchy with different motifs. Each choice of hierarchical cut motif and cut level allows the material to expand into a unique structure with a unique set of properties. We can reverse-engineer the desired expanded geometries to find the requisite cut pattern to produce it without changing the physical properties of the initial material. The concept was experimentally realized and applied to create an electrode that expands to >800% the original area with only very minor stretching of the underlying material. The generality of our approach greatly expands the design space for materials so that they can be tuned for diverse applications.

10.
J Biol Chem ; 289(36): 25306-16, 2014 Sep 05.
Article in English | MEDLINE | ID: mdl-25037223

ABSTRACT

The histone deacetylase inhibitor (HDACi) sodium butyrate promotes differentiation of colon cancer cells as evidenced by induced expression and enzyme activity of the differentiation marker intestinal alkaline phosphatase (ALPi). Screening of a panel of 33 colon cancer cell lines identified cell lines sensitive (42%) and resistant (58%) to butyrate induction of ALP activity. This differential sensitivity was similarly evident following treatment with the structurally distinct HDACi, MS-275. Resistant cell lines were significantly enriched for those harboring the CpG island methylator phenotype (p = 0.036, Chi square test), and resistant cell lines harbored methylation of the ALPi promoter, particularly of a CpG site within a critical KLF/Sp regulatory element required for butyrate induction of ALPi promoter activity. However, butyrate induction of an exogenous ALPi promoter-reporter paralleled up-regulation of endogenous ALPi expression across the cell lines, suggesting the presence or absence of a key transcriptional regulator is the major determinant of ALPi induction. Through microarray profiling of sensitive and resistant cell lines, we identified KLF5 to be both basally more highly expressed as well as preferentially induced by butyrate in sensitive cell lines. KLF5 overexpression induced ALPi promoter-reporter activity in resistant cell lines, KLF5 knockdown attenuated butyrate induction of ALPi expression in sensitive lines, and butyrate selectively enhanced KLF5 binding to the ALPi promoter in sensitive cells. These findings demonstrate that butyrate induction of the cell differentiation marker ALPi is mediated through KLF5 and identifies subsets of colon cancer cell lines responsive and refractory to this effect.


Subject(s)
Alkaline Phosphatase/metabolism , Cell Differentiation/drug effects , Epithelial Cells/metabolism , Histone Deacetylase Inhibitors/pharmacology , Kruppel-Like Transcription Factors/metabolism , Alkaline Phosphatase/genetics , Benzamides/pharmacology , Binding Sites/genetics , Blotting, Western , Butyric Acid/pharmacology , Cell Differentiation/genetics , Cell Line, Tumor , Colonic Neoplasms/genetics , Colonic Neoplasms/metabolism , Colonic Neoplasms/pathology , CpG Islands/genetics , DNA Methylation , Drug Resistance, Neoplasm/genetics , Gene Expression Profiling , HCT116 Cells , HT29 Cells , Humans , Kruppel-Like Transcription Factors/genetics , Oligonucleotide Array Sequence Analysis , Promoter Regions, Genetic/genetics , Protein Binding , Pyridines/pharmacology , Reverse Transcriptase Polymerase Chain Reaction
11.
Am J Physiol Gastrointest Liver Physiol ; 301(5): G856-64, 2011 Nov.
Article in English | MEDLINE | ID: mdl-21836055

ABSTRACT

Phosphatase and tensin homolog deleted on chromosome 10 (PTEN) is a negative regulator of phosphatidylinositol 3-kinase (PI3K) signaling that is frequently inactivated in colorectal cancer through mutation, loss of heterozygosity, or epigenetic mechanisms. The aim of this study was to determine the effect of intestinal-specific PTEN inactivation on intestinal epithelial homeostasis and tumorigenesis. PTEN was deleted specifically in the intestinal epithelium, by crossing PTEN(Lox/Lox) mice with villin(Cre) mice. PTEN was robustly expressed in the intestinal epithelium and maximally in the differentiated cell compartment. Targeted inactivation of PTEN in the intestinal epithelium of PTEN(Lox/Lox)/villin(Cre) mice was confirmed by genotyping, immunohistochemistry, and qPCR. While intestinal-specific PTEN deletion did not have a major effect on cell fate determination or proliferation in the small intestine, it did increase phosphorylated (p) protein kinase B (AKT) expression in the intestinal epithelium, and 19% of animals developed small intestinal adenomas and adenocarcinomas at 12 mo of age. These tumors demonstrated pAKT and nuclear ß-catenin staining, indicating simultaneous activation of the PI3K/AKT and Wnt signaling pathways. These findings demonstrate that, while PTEN inactivation alone has a minimal effect on intestinal homeostasis, it can facilitate tumor promotion upon deregulation of ß-catenin/TCF signaling, further establishing PTEN as a bona fide tumor suppressor gene in intestinal cancer.


Subject(s)
Adenocarcinoma/metabolism , Adenoma/metabolism , Epithelial Cells/metabolism , Intestinal Neoplasms/metabolism , Intestine, Small/metabolism , PTEN Phosphohydrolase/metabolism , Adenocarcinoma/genetics , Adenocarcinoma/pathology , Adenoma/genetics , Adenoma/pathology , Animals , Epithelial Cells/pathology , Intestinal Neoplasms/genetics , Intestinal Neoplasms/pathology , Intestine, Small/pathology , Mice , Mice, Knockout , PTEN Phosphohydrolase/genetics , Phosphorylation , Proto-Oncogene Proteins c-akt/genetics , Proto-Oncogene Proteins c-akt/metabolism , Signal Transduction/genetics , Wnt Proteins/genetics , Wnt Proteins/metabolism , beta Catenin/genetics , beta Catenin/metabolism
12.
J Clin Periodontol ; 37(3): 305-11, 2010 Mar.
Article in English | MEDLINE | ID: mdl-20041973

ABSTRACT

AIM: The aim of the present study was to investigate bone regeneration following ex vivo bone morphogenetic protein-2 (BMP-2) gene delivery using human gingival fibroblasts (HGFs) in rat calvarial defects. MATERIALS AND METHODS: An 8 mm craniotomy defect was created in Sprague-Dawley rats. The animals were divided into four groups: (1) non-grafted group, the defect was left empty; (2) collagen matrix group, the defect was filled with collagen matrix only; (3) HGF group, the defect was filled with non-transduced HGFs on collagen matrix; (4) BMP-2/HGF group, the defect was filled with BMP-2 gene-transduced HGFs on collagen matrix. Animals were sacrificed at 2 and 4 weeks after surgery, and micro-computed tomographic and histologic observations were performed. RESULTS: The BMP-2/HGF group showed promoted osseous healing of calvarial defects, as compared with the other groups. At both 2 and 4 weeks, regenerated bone area was significantly greater in the BMP-2/HGF group than the other three groups. Quite a few number of transplanted HGFs were observed within the regenerated bone tissues. CONCLUSIONS: The results of this study suggest that ex vivo BMP-2 gene delivery induces prominent bone regeneration in vivo and HGFs may be useful as target cells for ex vivo gene therapy.


Subject(s)
Bone Morphogenetic Protein 2/physiology , Bone Regeneration/physiology , Genetic Therapy/methods , Guided Tissue Regeneration/methods , Osseointegration/physiology , Adolescent , Animals , Bone Matrix/cytology , Bone Morphogenetic Protein 2/administration & dosage , Bone Morphogenetic Protein 2/genetics , Bone Regeneration/genetics , Bone Substitutes , Cells, Cultured , Craniotomy , Female , Fibroblasts/cytology , Fibroblasts/physiology , Fibroblasts/transplantation , Gene Transfer Techniques , Gingiva/cytology , Humans , Implants, Experimental , Male , Rats , Rats, Sprague-Dawley , Statistics, Nonparametric , Tissue Engineering/methods , Tissue Scaffolds , X-Ray Microtomography
13.
Cancer Res ; 69(19): 7811-8, 2009 Oct 01.
Article in English | MEDLINE | ID: mdl-19789347

ABSTRACT

Colorectal cancers (CRC) with microsatellite instability (MSI) have clinical, pathologic, genetic, and epigenetic features distinct from microsatellite-stable CRC. Examination of epidermal growth factor receptor (EGFR) mRNA and protein expression levels in a panel of colon cancer cell lines identified strong expression of EGFR in multiple cell lines with MSI. Although no relationship between EGFR overexpression and the length of a CA dinucleotide repeat in intron 1 was observed, a variant A13/A14 repeat sequence within the 3'-untranslated region (3'-UTR) of the EGFR gene was identified, which was mutated by either mononucleotide or dinucleotide adenosine deletions in 64% of MSI cell lines and 69% of MSI colon tumors. Using a Tet-Off system, we show that this mutation increases EGFR mRNA stability in colon cancer cells, providing a mechanistic basis for EGFR overexpression in MSI colon cancer cell lines. To determine whether this mutation is a driver or a bystander event in MSI colon cancer, we examined the effect of pharmacologic and molecular inhibition of EGFR in EGFR 3'-UTR mutant MSI cell lines. Cell lines with an EGFR 3'-UTR mutation and that were wild-type (WT) for downstream signaling mediators in the Ras/BRAF and PIK3CA/PTEN pathways were sensitive to EGFR inhibition, whereas those harboring mutations in these signaling mediators were not. Furthermore, in cell lines WT for downstream signaling mediators, those with EGFR 3'-UTR mutations were more sensitive to EGFR inhibition than EGFR 3'-UTR WT cells, suggesting that this mutation provides a growth advantage to this subset of MSI colon tumors.


Subject(s)
Colonic Neoplasms/genetics , ErbB Receptors/biosynthesis , Genes, erbB-1 , Mutation , Poly A/genetics , Repetitive Sequences, Nucleic Acid , 3' Untranslated Regions , Animals , Cell Line, Tumor , Colonic Neoplasms/enzymology , ErbB Receptors/genetics , Gene Amplification , Humans , Mice , Mice, SCID , Microsatellite Instability , RNA, Messenger/biosynthesis , RNA, Messenger/genetics , Ribonuclease III/antagonists & inhibitors , Ribonuclease III/genetics , Sequence Deletion
14.
Clin Cancer Res ; 15(8): 2797-807, 2009 Apr 15.
Article in English | MEDLINE | ID: mdl-19351753

ABSTRACT

PURPOSE: We examined the effects of G3139 on the interaction of heparin-binding proteins [e.g., fibroblast growth factor 2 (FGF2) and collagen I] with endothelial cells. G3139 is an 18-mer phosphorothioate oligonucleotide targeted to the initiation codon region of the Bcl-2 mRNA. A randomized, prospective global phase III trial in advanced melanoma (GM301) has evaluated G3139 in combination with dacarbazine. However, the mechanism of action of G3139 is incompletely understood because it is unlikely that Bcl-2 silencing is the sole mechanism for chemosensitization in melanoma cells. EXPERIMENTAL DESIGN: The ability of G3139 to interact with and protect heparin-binding proteins was quantitated. The effects of G3139 on the binding of FGF2 to high-affinity cell surface receptors and the induction of cellular mitogenesis and tubular morphogenesis in HMEC-1 and human umbilical vascular endothelial cells were determined. RESULTS: G3139 binds with picomolar affinity to collagen I. By replacing heparin, the drug can potentiate the binding of FGF2 to FGFR1 IIIc, and it protects FGF from oxidation and proteolysis. G3139 can increase endothelial cell mitogenesis and tubular morphogenesis of HMEC-1 cells in three-dimensional collagen gels, increases the mitogenesis of human umbilical vascular endothelial cells similarly, and induces vessel sprouts in the rat aortic ring model. CONCLUSIONS: G3139 dramatically affects the behavior of endothelial cells. There may be a correlation between this observation and the treatment interaction with lactate dehydrogenase observed clinically.


Subject(s)
Endothelial Cells/drug effects , Fibroblast Growth Factor 2/metabolism , Oligonucleotides, Antisense/pharmacology , Platelet-Derived Growth Factor/metabolism , Receptor, Fibroblast Growth Factor, Type 1/metabolism , Thionucleotides/pharmacology , Vascular Endothelial Growth Factor A/metabolism , Animals , Becaplermin , Cell Proliferation/drug effects , Collagen Type I/metabolism , Cytoskeleton/drug effects , Cytoskeleton/metabolism , Endothelial Cells/metabolism , Fibroblast Growth Factor 2/drug effects , Humans , Oligonucleotides, Antisense/genetics , Oligonucleotides, Antisense/metabolism , Proto-Oncogene Proteins c-bcl-2/antagonists & inhibitors , Proto-Oncogene Proteins c-bcl-2/genetics , Proto-Oncogene Proteins c-sis , Rats , Receptor, Fibroblast Growth Factor, Type 1/drug effects , Thionucleotides/genetics , Thionucleotides/metabolism
15.
Blood ; 112(10): 4343-52, 2008 Nov 15.
Article in English | MEDLINE | ID: mdl-18711003

ABSTRACT

Defibrotide (DF) is a mixture of porcine-derived single-stranded phosphodiester oligonucleotides (9-80-mer; average, 50-mer) that has been successfully used to treat severe hepatic veno-occlusive disease (sVOD) with multiorgan failure (MOF) in patients who have received cytotoxic chemotherapy in preparation for bone marrow transplantation. However, its mechanism of action is unknown. Herein, we show that DF and phosphodiester oligonucleotides can bind to heparin-binding proteins (eg, basic fibroblast growth factor [bFGF] but not vascular endothelial growth factor [VEGF] 165) with low nanomolar affinity. This binding occurred in a length- and concentration-dependent manner. DF can mobilize proangiogenic factors such as bFGF from their depot or storage sites on bovine corneal endothelial matrix. However, these molecules do not interfere with high-affinity binding of bFGF to FGFR1 IIIc but can replace heparin as a required cofactor for binding and hence cellular mitogenesis. DF also protects bFGF against digestion by trypsin and chymotrypsin and from air oxidation. In addition, DF binds to collagen I with low nanomolar affinity and can promote human microvascular endothelial cell-1 (HMEC-1) cell mitogenesis and tubular morphogenesis in three-dimensional collagen I gels. Thus, our data suggest that DF may provide a stimulus to the sinusoidal endothelium of a liver that has suffered a severe angiotoxic event, thus helping to ameliorate the clinical sVOD/MOF syndrome.


Subject(s)
Endothelial Cells/metabolism , Fibrinolytic Agents/pharmacology , Fibroblast Growth Factor 2/metabolism , Hepatic Veno-Occlusive Disease/metabolism , Neovascularization, Physiologic/drug effects , Polydeoxyribonucleotides/pharmacology , Animals , Bone Marrow Transplantation/adverse effects , Cattle , Cell Line, Transformed , Cornea/metabolism , Cornea/pathology , Endothelial Cells/pathology , Fibrinolytic Agents/therapeutic use , Hepatic Veno-Occlusive Disease/drug therapy , Hepatic Veno-Occlusive Disease/etiology , Hepatic Veno-Occlusive Disease/pathology , Humans , Multiple Organ Failure/drug therapy , Multiple Organ Failure/etiology , Multiple Organ Failure/metabolism , Multiple Organ Failure/pathology , Polydeoxyribonucleotides/therapeutic use , Protein Binding/drug effects , Receptor, Fibroblast Growth Factor, Type 3/metabolism , Vascular Endothelial Growth Factor A/metabolism
16.
Cancer Res ; 68(6): 1953-61, 2008 Mar 15.
Article in English | MEDLINE | ID: mdl-18339877

ABSTRACT

Cetuximab is a monoclonal antibody that targets the human epidermal growth factor receptor (EGFR). Although approved for use in EGFR-overexpressing advanced colorectal cancer, recent studies have shown a lack of association between EGFR overexpression and cetuximab response, requiring the identification of novel biomarkers predictive of response to this agent. To do so, 22 colon cancer cell lines were screened for cetuximab response in vitro and sensitive and resistant lines were identified. In sensitive cell lines, cetuximab induced a G(0)-G(1) arrest without inducing apoptosis. Notably, cetuximab-sensitive but not cetuximab-resistant cell lines were preferentially responsive to EGF-stimulated growth. Whereas neither EGFR protein/mRNA expression nor gene copy number correlated with cetuximab response, examination of the mutation status of signaling components downstream of EGFR showed that cell lines with activating PIK3CA mutations or loss of PTEN expression (PTEN null) were more resistant to cetuximab than PIK3CA wild type (WT)/PTEN-expressing cell lines (14 +/- 5.0% versus 38.5 +/- 6.4% growth inhibition, mean +/- SE; P = 0.008). Consistently, PIK3CA mutant isogenic HCT116 cells showed increased resistance to cetuximab compared with PIK3CA WT controls. Furthermore, cell lines that were PIK3CA mutant/PTEN null and Ras/BRAF mutant were highly resistant to cetuximab compared with those without dual mutations/PTEN loss (10.8 +/- 4.3% versus 38.8 +/- 5.9% growth inhibition, respectively; P = 0.002), indicating that constitutive and simultaneous activation of the Ras and PIK3CA pathways confers maximal resistance to this agent. A priori screening of colon tumors for PTEN expression status and PIK3CA and Ras/BRAF mutation status could help stratify patients likely to benefit from this therapy.


Subject(s)
Antibodies, Monoclonal/pharmacology , Antineoplastic Agents/pharmacology , Colonic Neoplasms/drug therapy , Mutation , PTEN Phosphohydrolase/biosynthesis , Phosphatidylinositol 3-Kinases/genetics , Antibodies, Monoclonal, Humanized , Cell Line, Tumor , Cetuximab , Class I Phosphatidylinositol 3-Kinases , Colonic Neoplasms/genetics , Colonic Neoplasms/metabolism , Colonic Neoplasms/pathology , Epidermal Growth Factor/pharmacology , ErbB Receptors/biosynthesis , ErbB Receptors/genetics , ErbB Receptors/metabolism , Erlotinib Hydrochloride , G1 Phase/drug effects , Gene Dosage , Genes, ras , HCT116 Cells , Humans , MAP Kinase Signaling System , Proto-Oncogene Proteins B-raf/genetics , Quinazolines/pharmacology , RNA, Messenger/biosynthesis , RNA, Messenger/genetics , Resting Phase, Cell Cycle/drug effects , ras Proteins/genetics
17.
Surgery ; 142(2): 222-7, 2007 Aug.
Article in English | MEDLINE | ID: mdl-17689689

ABSTRACT

BACKGROUND: We have previously published results indicating that decreased expression of CDK2-AP1 in MSI human colorectal cancer is associated with deletion mutations in the poly (T) 8 repeat sequence within the 3'-UTR of the CDK2-AP1 gene. In this study, we test the hypothesis that the del T mutation results in decreased CDK2-AP1 expression by causing reduced mRNA stability. METHODS: We introduced wild-type and mutant 3'-UTR sequences fused to a green fluorescent protein (GFP) gene separately into human CRC cell lines and quantified the expression of the GFP gene. Native CDK2-AP1 mRNA stability was measured in human CRC cell lines, using an actinomycin D assay and the mRNA structure folding software mfold 3.2. RESULTS: Mutant GFP-3'-UTR samples demonstrated significantly reduced GFP expression compared with wild-type GFP-3'-UTR as measured by both FACS and real-time PCR. Both the actinomycin D assay and mfold software demonstrated significantly reduced mRNA stability for the del T poly (T) 8 transcript compared with the wild type. CONCLUSIONS: In summary, these novel results support our hypotheses that the del T poly (T) 8 observed in the 3'-UTR of the CDK2-AP1 gene in human MSI CRC is functionally significant and results in decreased CDK2-AP1 expression. The results also indicate the mechanism of this decreased expression is caused at least in part by decreased mRNA stability.


Subject(s)
Colorectal Neoplasms/genetics , Gene Deletion , RNA Stability/genetics , Tumor Suppressor Proteins/genetics , 3' Untranslated Regions/genetics , Cell Line, Tumor , Colorectal Neoplasms/physiopathology , Gene Expression Regulation, Neoplastic , Green Fluorescent Proteins/genetics , Humans , Microsatellite Instability , Nucleic Acid Conformation , RNA, Messenger/chemistry , RNA, Messenger/genetics , RNA, Neoplasm/chemistry , RNA, Neoplasm/genetics
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