Andrew A. Shelton

Phenotypic characterization of human colorectal cancer stem cells

Recent observations indicate that, in several types of human cancer, only a phenotypic subset of cancer cells within each tumor is capable of initiating tumor growth. This functional subset of cancer cells is operationally defined as the “cancer stem cell” (CSC) subset. Here we developed a CSC model for the study of human colorectal cancer (CRC). Solid CRC tissues, either primary tissues collected from surgical specimens or xenografts established in nonobese diabetic/severe combined immunodeficient (NOD/SCID) mice, were disaggregated into single-cell suspensions and analyzed by flow cytometry. Surface markers that displayed intratumor heterogeneous expression among epithelial cancer cells were selected for cell sorting and tumorigenicity experiments. Individual phenotypic cancer cell subsets were purified, and their tumor-initiating properties were investigated by injection in NOD/SCID mice. Our observations indicate that, in six of six human CRC tested, the ability to engraft in vivo in immunodeficient mice was restricted to a minority subpopulation of epithelial cell adhesion molecule (EpCAM)high/CD44+ epithelial cells. Tumors originated from EpCAMhigh/CD44+ cells maintained a differentiated phenotype and reproduced the full morphologic and phenotypic heterogeneity of their parental lesions. Analysis of the surface molecule repertoire of EpCAMhigh/CD44+ cells led to the identification of CD166 as an additional differentially expressed marker, useful for CSC isolation in three of three CRC tested. These results validate the stem cell working model in human CRC and provide a highly robust surface marker profile for CRC stem cell isolation.

Single-cell dissection of transcriptional heterogeneity in human colon tumors.

Cancer is often viewed as a caricature of normal developmental processes, but the extent to which its cellular heterogeneity truly recapitulates multilineage differentiation processes of normal tissues remains unknown. Here we implement single-cell PCR gene-expression analysis to dissect the cellular composition of primary human normal colon and colon cancer epithelia. We show that human colon cancer tissues contain distinct cell populations whose transcriptional identities mirror those of the different cellular lineages of normal colon. By creating monoclonal tumor xenografts from injection of a single (n = 1) cell, we demonstrate that the transcriptional diversity of cancer tissues is largely explained by in vivo multilineage differentiation and not only by clonal genetic heterogeneity. Finally, we show that the different gene-expression programs linked to multilineage differentiation are strongly associated with patient survival. We develop two-gene classifier systems (KRT20 versus CA1, MS4A12, CD177, SLC26A3) that predict clinical outcomes with hazard ratios superior to those of pathological grade and comparable to those of microarray-derived multigene expression signatures.

Colon, rectum, and anus

The colon is one structural unit with two embryological origins. The cecum and right and midtransverse colons are of midgut origin and as such are supplied by the superior mesenteric artery (SMA). The distal transverse, splenic flexure, and descending and sigmoid colon are of hindgut origin and receive blood from the inferior mesenteric artery (IMA). The entire colon starts as a midline structure that rotates during development and attaches laterally to the right and left posterior peritoneum. The right and left colonic mesenteries are obliterated, fusing to the posterior peritoneum in these regions, leaving these portions of the colon covered by peritoneum on the lateral, anterior, and medial surfaces. The transverse and sigmoid colons, in contrast, are completely covered with peritoneum and are attached by long mesenteries, allowing for great variation in the location of these structures (Fig. 51.1).

Intensity-modulated radiation therapy versus conventional radiation therapy for squamous cell carcinoma of the anal canal.

The purpose of this study was to compare outcomes in patients with anal canal squamous cell carcinoma (SCCA) who were treated with definitive chemoradiotherapy by either intensity‐modulated radiation therapy (IMRT) or conventional radiotherapy (CRT).

Clinicopathologic and molecular features of sporadic early-onset colorectal adenocarcinoma: an adenocarcinoma with frequent signet ring cell differentiation, rectal and sigmoid involvement, and adverse morphologic features

Recent literature suggests an increasing incidence of colorectal carcinoma in young patients. We performed a histologic, molecular, and immunophenotypic analysis of patients with sporadic early-onset (≤40 years of age) colorectal carcinoma seen at our institution from the years 2000–2010 and compared these tumors to a cohort of consecutively resected colorectal carcinomas seen in patients >40 years of age. A total of 1160 primary colorectal adenocarcinomas were surgically resected for the years 2000 through 2010. Of these, 75 (6%) were diagnoses in patients ≤40 years of age of which 13 (17%) demonstrated abnormalities in DNA mismatch repair, 4 (5%) were in patients with known germline genetic disorders (two patients with familial adenomatous polyposis, one patient with juvenile polyposis, and one patient with Li-Fraumeni syndrome), and three patients (4%) had long-standing chronic inflammatory bowel disease. The sporadic early-onset colorectal carcinoma group comprised a total of 55 patients (55/1160, 5%) and were compared with a control group comprising 73 consecutively resected colorectal carcinomas with proficient DNA mismatch repair in patients >40 years of age. For the early-onset colorectal carcinoma group, most cases (33/55, 60%) were diagnosed between the age of 35 and 40 years of age. Compared with the control group, the early-onset colorectal carcinoma group was significantly different with respect to tumor location (P<0.007) with 80% (44/55 cases) identified in either the sigmoid colon (24/55, 44%) or rectum (20/55, 36%). Morphologically, early-onset colorectal carcinomas more frequently displayed adverse histologic features compared with the control colorectal carcinoma group such as signet ring cell differentiation (7/55, 13% vs 1/73, 1%, P=0.021), perineural invasion (16/55, 29% vs 8/73, 11%, P=0.009) and venous invasion (12/55, 22% vs 4/73, 6%, P=0.006). A precursor adenomatous lesion was less frequently identified in the early-onset colorectal carcinoma group compared with the control group (19/55, 35% vs 39/73, 53%, P=0.034). Of the early-onset colorectal carcinomas, only 2/45 cases (4%) demonstrated KRAS mutations compared with 11/73 (15%) of the control group colorectal adenocarcinomas harboring KRAS mutations, although this difference did not reach statistical significance (P=0.13). BRAF V600E mutations were not identified in the early-onset colorectal carcinoma group. No difference was identified between the two groups with regard to tumor stage, tumor size, number of lymph node metastases, lymphatic invasion, tumor budding, mucinous histology, or tumor-infiltrating lymphocytes. Both groups had similar recurrence-free (P=0.28) and overall survival (P=0.73). However, patients in the early-onset colorectal carcinoma group more frequently either presented with or developed metastatic disease during their disease course compared with the control colorectal carcinoma group (25/55, 45% vs 18/73, 25%, P=0.014). In addition, 8/55 patients (15%) in the early-onset colorectal carcinoma group developed local recurrence of their tumor while no patients in the control colorectal carcinoma group developed local recurrence (P<0.001), likely due to the increased incidence of rectal carcinoma in the patients with early-onset colorectal carcinoma. Our study demonstrates that colorectal carcinoma is not infrequently diagnosed in patients ≤40 years of age and is not frequently the result of underlying Lynch syndrome or associated with other cancer-predisposing genetic conditions or chronic inflammatory conditions. These tumors have a striking predilection for the distal colon, particularly the sigmoid colon and rectum and are much more likely to demonstrate adverse histologic factors, including signet ring cell differentiation, venous invasion, and perineural invasion.

Mechanical bowel preparation in intestinal surgery: a meta-analysis and review of the literature

IntroductionDespite several meta-analyses and randomized controlled trials showing no benefit to patients, mechanical bowel preparation (MBP) remains the standard of practice for patients undergoing elective colorectal surgery.MethodsWe performed a systematic review of the literature of trials that prospectively compared MBP with no MBP for patients undergoing elective colorectal resection. We searched MEDLINE, LILACS, and SCISEARCH, abstracts of pertinent scientific meetings and reference lists for each article found. Experts in the field were queried as to knowledge of additional reports. Outcomes abstracted were anastomotic leaks and wound infections. Meta-analysis was performed using Peto Odds ratio.ResultsOf 4,601 patients (13 trials), 2,304 received MBP (Group 1) and 2,297 did not (Group 2). Anastomotic leaks occurred in 97(4.2%) patients in Group 1 and in 81(3.5%) patients in Group 2 (Peto OR = 1.214, CI 95%:0.899–1.64, P = 0.206). Wound infections occurred in 227(9.9%) patients in Group 1 and in 201(8.8%) patients in Group 2 (Peto OR = 1.156, CI 95%:0.946–1.413, P = 0.155).DiscussionThis meta-analysis demonstrates that MBP provides no benefit to patients undergoing elective colorectal surgery, thus, supporting elimination of routine MBP in elective colorectal surgery.ConclusionIn conclusion, MBP is of no benefit to patients undergoing elective colorectal resection and need not be recommended to meet “standard of care.”

Outcomes of repeat colonoscopy in patients with polyps referred for surgery without biopsy-proven cancer

BACKGROUND Despite advances in endoscopic treatment, many colonic adenomas are still referred for surgical resection. There is a paucity of data on the suitability of these lesions for endoscopic treatment. OBJECTIVE To analyze the results of routine repeat colonoscopy in patients referred for surgical resection of colon polyps without biopsy-proven cancer. DESIGN Retrospective review. SETTING University hospital. PATIENTS Patients referred to a colorectal surgeon for surgical resection of a polyp without biopsy-proven cancer. INTERVENTIONS Repeat colonoscopy. MAIN OUTCOME MEASUREMENTS The rate of successful endoscopic treatment. RESULTS There were 38 lesions in 36 patients; 71% of the lesions were noncancerous and were successfully treated endoscopically. In 26% of the lesions, previous removal was attempted by the referring physician but was unsuccessful. The adenoma recurrence rate was 50%, but all recurrences were treated endoscopically and none were cancerous. Two patients were admitted for overnight observation. There were no major adverse events. LIMITATIONS Single center, retrospective. CONCLUSIONS In the absence of biopsy-proven invasive cancer, it is appropriate to reevaluate patients referred for surgical resection by repeat colonoscopy at an expert center.

Dartos Muscle Interposition Flap for the Treatment of Rectourethral Fistulas

PurposeRectourethral fistula is a rare complication of pelvic surgery, trauma, or inflammation. The many techniques for repairing these fistulas vary in their success rates. Our goal was to describe the use of a dartos muscle interposition flap for repair of these fistulas.MethodsWe performed a retrospective review of eight patients who underwent repair of a rectourethral fistula with a dartos muscle interposition flap. We describe the success rate and patient-related factors that may have affected success. The technique of dartos muscle interposition is described and compared with other previously described techniques.ResultsSix of eight patients had healing of their fistulas documented by follow-up cystogram.ConclusionsDartos muscle interposition is a straightforward technique that can result in successful fistula repair but should not be used in patients with risk factors for poor wound healing, such as an immunocompromised state or previous radiation therapy.

Predictors of postoperative urinary retention after colorectal surgery.

BACKGROUND: National quality initiatives have mandated the earlier removal of urinary catheters after surgery to decrease urinary tract infection rates. A potential unintended consequence is an increased postoperative urinary retention rate. OBJECTIVE: The aim of this study was to determine the incidence and risk factors for postoperative urinary retention after colorectal surgery. DESIGN: This was a prospective observational study. SETTINGS: A colorectal unit within a single institution was the setting for this study. PATIENTS: Adults undergoing elective colorectal operations were included. INTERVENTIONS: Urinary catheters were removed on postoperative day 1 for patients undergoing abdominal operations, and on day 3 for patients undergoing pelvic operations. Postvoid residual and retention volumes were measured. MAIN OUTCOME MEASURES: The primary outcomes measured were urinary retention and urinary tract infection. RESULTS: The overall urinary retention rate was 22.4% (22.8% in the abdominal group, 21.9% in the pelvic group) and was associated with longer operative time and increased perioperative fluid administration. Mean operative time for those with retention was 2.8 hours and, for those without retention, the mean operative time 2.2 hours (abdominal group 2 hours vs 1.4 hours, pelvic group 3.9 hours vs 3.1 hours, p ⩽ 0.02). Patients with retention received a mean of 2.7L during the operation, whereas patients without retention received 1.8L (abdominal group 1.9L vs 1.4L, pelvic group 3.6L vs 2.2L, p < 0.01). In the abdominal group, patients with and without retention also received different fluid volumes on postoperative days 1 (2.2L vs 1.7L, p = 0.004) and 2 (1.6L vs 1L, p = 0.05). Laparoscopic abdominal group had a 40% retention rate in comparison with 12% in the open abdominal group (p = 0.004). Age, sex, preoperative radiation therapy, preoperative prostatism, preoperative diagnosis, and level of anastomosis were not associated with retention. The urinary tract infection rate was 4.9%. LIMITATION: The lack of documentation of preoperative urinary function was a limitation of this study. CONCLUSIONS: The practice of earlier urinary catheter removal must be balanced with operative time and fluid volume to avoid high urinary retention rates. Also important is increased vigilance for the early detection of retention.

The Prognostic Significance of Pretreatment Hematologic Parameters in Patients Undergoing Resection for Colorectal Cancer.

Objectives: The prognostic value of several hematologic parameters, including platelet, lymphocyte, and neutrophil counts, has been studied in a variety of solid tumors. In this study, we examined the significance of inflammatory markers and their prognostic implications in patients with colorectal cancer (CRC). Materials and Methods: Patients with stage I-III CRC who underwent surgical resection at the Stanford Cancer Institute between 2005 and 2009 were included. Patients were excluded if they did not have preoperative complete blood counts performed within 1 month of surgical resection, underwent preoperative chemotherapy or radiation, had metastatic disease at diagnosis, or had another previous malignancy. We included 129 eligible patients with available preoperative complete blood counts in the final analysis. Results: A preoperative neutrophil-to-lymphocyte ratio of>3.3 was significantly associated with worse disease-free (DFS) and overall survival (OS) (P=0.009, 0.003), as was a preoperative lymphocyte-to-monocyte ratio of ⩽2.6 (P=0.01, 0.002). Preoperative lymphopenia (P=0.002) was associated with worse OS but not DFS (P=0.09). In addition, preoperative thrombocytosis was associated with worse DFS (P=0.006) and OS (P=0.010). Preoperative leukocytosis was associated with worse OS (P=0.048) but not DFS (P=0.49). Preoperative hemoglobin was neither associated with OS (P=0.24) or DFS (P=0.15). Conclusions: Pretreatment lymphopenia, thrombocytosis, a decreased lymphocyte-to-monocyte ratio, and an elevated neutrophil-to-lymphocyte ratio independently predict for worse OS in patients with CRC.