Proton Treatment Suppresses Exosome Production in Head and Neck Squamous Cell Carcinoma.

Proton therapy (PT) is emerging as an effective and less toxic alternative to conventional X-ray-based photon therapy (XRT) for patients with advanced head and neck squamous cell carcinomas (HNSCCs) owing to its clustered dose deposition dosimetric characteristics. For optimal efficacy, cancer therapies, including PT, must elicit a robust anti-tumor response by effector and cytotoxic immune cells in the tumor microenvironment (TME). While tumor-derived exosomes contribute to immune cell suppression in the TME, information on the effects of PT on exosomes and anti-tumor immune responses in HNSCC is not known. In this study, we generated primary HNSCC cells from tumors resected from HNSCC patients, irradiated them with 5 Gy PT or XRT, and isolated exosomes from cell culture supernatants. HNSCC cells exposed to PT produced 75% fewer exosomes than XRT- and non-irradiated HNSCC cells. This effect persisted in proton-irradiated cells for up to five days. Furthermore, we observed that exosomes from proton-irradiated cells were identical in morphology and immunosuppressive effects (suppression of IFN-γ release by peripheral blood mononuclear cells) to those of photon-irradiated cells. Our results suggest that PT limits the suppressive effect of exosomes on cancer immune surveillance by reducing the production of exosomes that can inhibit immune cell function.

Rapid purification and multiparametric characterization of circulating small extracellular vesicles utilizing a label-free lab-on-a-chip device.

Nano-scale extracellular vesicles are lipid-bilayer delimited particles that are naturally secreted by all cells and have emerged as valuable biomarkers for a wide range of diseases. Efficient isolation of small extracellular vesicles while maintaining yield and purity is crucial to harvest their potential in diagnostic, prognostic, and therapeutic applications. Most conventional methods of isolation suffer from significant shortcomings, including low purity or yield, long duration, need for large sample volumes, specialized equipment, trained personnel, and high costs. To address some of these challenges, our group has reported a novel insulator-based dielectrophoretic device for rapid isolation of small extracellular vesicles from biofluids and cell culture media based on their size and dielectric properties. In this study, we report a comprehensive characterization of small extracellular vesicles isolated from cancer-patients' biofluids at a twofold enrichment using the device. The three-fold characterization that was performed using conventional flow cytometry, advanced imaging flow cytometry, and microRNA sequencing indicated high yield and purity of the isolated small extracellular vesicles. The device thus offers an efficient platform for rapid isolation while maintaining biomolecular integrity.

Small extravesicular microRNA in head and neck squamous cell carcinoma and its potential as a liquid biopsy for early detection.

BACKGROUND: The objective was to assess secretion of small extracellular vesicular microRNA (exo-miRNA) in head and neck squamous cell carcinoma (HNSCC) according to human papillomavirus (HPV) status, and determine the translational potential as a liquid biopsy for early detection. METHODS: This study employed a combination of cell culture and case-control study design using archival pretreatment serum. Small extracellular vesicles (sEV) were isolated from conditioned culture media and human serum samples via differential ultracentrifugation. miRNA-sequencing was performed on each sEV isolate. RESULTS: There were clear exo-miRNA profiles that distinguished HNSCC cell lines from nonpathologic oral epithelial control cells. While there was some overlap among profiles across all samples, there were apparent differences in exo-miRNA profiles according to HPV-status. Importantly, differential exo-miRNA profiles were also apparent in serum from early-stage HNSCC cases relative to cancer-free controls. CONCLUSIONS: Our findings indicate that exo-miRNA are highly dysregulated in HNSCC and support the potential of exo-miRNA as biomarkers for HNSCC.

Gestational Perfluoroalkyl Substance Exposure and DNA Methylation at Birth and 12 Years of Age: A Longitudinal Epigenome-Wide Association Study.

BACKGROUND: DNA methylation alterations may underlie associations between gestational perfluoroalkyl substances (PFAS) exposure and later-life health outcomes. To the best of our knowledge, no longitudinal studies have examined the associations between gestational PFAS and DNA methylation. OBJECTIVES: We examined associations of gestational PFAS exposure with longitudinal DNA methylation measures at birth and in adolescence using the Health Outcomes and Measures of the Environment (HOME) Study (2003-2006; Cincinnati, Ohio). METHODS: We quantified serum concentrations of perfluorooctanoate (PFOA), perfluorooctane sulfonate (PFOS), perfluorononanoate (PFNA), and perfluorohexane sulfonate (PFHxS) in mothers during pregnancy. We measured DNA methylation in cord blood (n=266) and peripheral leukocytes at 12 years of age (n=160) using the Illumina HumanMethylation EPIC BeadChip. We analyzed associations between log2-transformed PFAS concentrations and repeated DNA methylation measures using linear regression with generalized estimating equations. We included interaction terms between children's age and gestational PFAS. We performed Gene Ontology enrichment analysis to identify molecular pathways. We used Project Viva (1999-2002; Boston, Massachusetts) to replicate significant associations. RESULTS: After adjusting for covariates, 435 cytosine-guanine dinucleotide (CpG) sites were associated with PFAS (false discovery rate, q<0.05). Specifically, we identified 2 CpGs for PFOS, 12 for PFOA, 8 for PFHxS, and 413 for PFNA; none overlapped. Among these, 2 CpGs for PFOA and 4 for PFNA were replicated in Project Viva. Some of the PFAS-associated CpG sites annotated to gene regions related to cancers, cognitive health, cardiovascular disease, and kidney function. We found little evidence that the associations between PFAS and DNA methylation differed by children's age. DISCUSSION: In these longitudinal data, PFAS biomarkers were associated with differences in several CpGs at birth and at 12 years of age in or near genes linked to some PFAS-associated health outcomes. Future studies should examine whether DNA methylation mediates associations between gestational PFAS exposure and health. https://doi.org/10.1289/EHP10118.

DNA methylation-derived systemic inflammation indices and their association with oropharyngeal cancer risk and survival.

BACKGROUND: Head and neck squamous cell carcinomas are associated with systemic inflammation (SI). We evaluated whether DNA methylation-derived SI (mdSI) indices are associated with oropharyngeal cancer risk and survival. METHODS: Ninety-four oropharyngeal squamous cell carcinoma (OPSCC) cases and 57 controls with DNA methylation data were included. Logistic regression analysis and survival analysis were performed to test the association of mdSI indices with OPSCC risk and survival. RESULTS: Higher methylation-derived neutrophil-to-lymphocyte ratio (mdNLR) was associated with increased risk of OPSCC (OR = 1.21, 95%CI: 1.11-1.40) while no association was found with methylation-derived lymphocyte-to-monocyte ratio (mdLMR). For 5-year overall survival, higher mdLMR was significantly associated with decreased risk of death (HR = 0.25, 95%CI: 0.10-0.64) while the converse was observed for mdNLR (HR = 2.48, 95%CI: 1.04-5.92). CONCLUSION: We observed an association between mdSI indices and OPSCC risk and 5-year overall survival. It is possible to use mdLMR as an independent prognostic factor for OPSCC.

Human Papillomavirus and Survival of Sinonasal Squamous Cell Carcinoma Patients: A Systematic Review and Meta-Analysis.

Human papillomavirus (HPV) is detectable in a subset of sinonasal squamous cell carcinoma (SNSCC), but the impact on patient outcomes is presently unclear due to a modest number of studies with limited statistical power. Therefore, we conducted a systematic review and meta-analysis to better clarify this relationship. A PubMed search was conducted to identify all studies reporting on overall (OS) or disease-free survival (DFS) for SNSCC by HPV status. Hazard ratios (HR) and corresponding 95% confidence intervals (CI) were extracted or, when not provided, indirectly estimated from each manuscript. Summary survival curves for 5-year OS and estimating survival probability by HPV status at pre-specified time intervals from study-specific Kaplan-Meier curves generated 2-year DFS. Log HRs and log CIs were combined across studies to generate summary estimates and a corresponding 95% CIs for OS and DFS. We identified ten unique studies reporting on OS and four for DFS. We observed a significant association between HPV and OS (summary HR = 0.51, 95% CI: 0.38-0.70) with relatively low heterogeneity between studies. These results indicate that HPV is a significant predictor of more favorable survival for SNSCC, and thus may be a useful biomarker for prognostication and, potentially, treatment modulation.

Firefighter occupation is associated with increased risk for laryngeal and hypopharyngeal squamous cell carcinoma among men from the Greater Boston area.

OBJECTIVE: Firefighters are exposed to a wide variety of carcinogens during the line of duty, including several associated with head and neck cancer. Existing studies assessing head and neck cancer risk with firefighting have predominately included occupational cohorts or registry data, which are limited by inability to adjust for smoking and alcohol consumption-major risk factors for head and neck cancer. Our objective was to assess the risk of head and neck cancer among men with an occupational history as a firefighter. METHODS: This work was conducted using male subjects from a large population-based case-control study of head and neck cancer from the greater Boston area using self-reported occupational history (718 cases and 905 controls). RESULTS: An occupational history as a firefighter was reported for 11 cases and 14 controls. Although no significant association was observed overall, we observed substantial increased risk for hypopharyngeal and laryngeal squamous cell carcinoma among professional municipal firefighters who had a light or no smoking history (OR=8.06, 95% CI 1.74 to 37.41), with significantly increasing risk per decade as a firefighter (OR=2.10, 95% CI 1.06 to 4.14). CONCLUSION: Professional municipal firefighters may be at increased risk for hypopharyngeal and laryngeal squamous cell carcinoma due to carcinogenic exposures encountered during the line of duty.

Chrysotile fibers in tissue adjacent to laryngeal squamous cell carcinoma in cases with a history of occupational asbestos exposure.

Asbestos describes a group of naturally occurring fibrous silicate mineral compounds that have been associated with a number of respiratory maladies, including mesothelioma and lung cancer. In addition, based primarily on epidemiologic studies, asbestos has been implicated as a risk factor for laryngeal and pharyngeal squamous cell carcinoma (SCC). The main objective of this work was to strengthen existing evidence via empirical demonstration of persistent asbestos fibers embedded in the tissue surrounding laryngeal and pharyngeal SCC, thus providing a more definitive biological link between exposure and disease. Six human papillomavirus (HPV)-negative laryngeal (n = 4) and pharyngeal (n = 2) SCC cases with a history working in an asbestos-exposed occupation were selected from a large population-based case-control study of head and neck cancer. A laryngeal SCC case with no history of occupational asbestos exposure was included as a control. Tissue cores were obtained from adjacent nonneoplastic tissue in tumor blocks from the initial primary tumor resection, and mineral fiber analysis was performed using a scanning electron microscope equipped with an energy dispersive X-ray analyzer (EDXA). Chrysotile asbestos fiber bundles were identified in 3/6 of evaluated cases with a history of occupational asbestos exposure. All three cases had tumors originating in the larynx. In addition, a wollastonite fiber of unclear significance was identified one of the HPV-negative pharyngeal SCC cases. No mineral fibers were identified in adjacent tissue of the case without occupational exposure. The presence of asbestos fibers in the epithelial tissue surrounding laryngeal SCC in cases with a history of occupational asbestos exposure adds a key line of physical evidence implicating asbestos as an etiologic factor.

Comprehensive mapping of the methylation landscape of 16 CpG-dense regions in oral and pharyngeal squamous cell carcinoma.

Aim: The goal of this study was to comprehensively interrogate and map DNA methylation across 16 CpG-dense regions previously associated with oral and pharyngeal squamous cell carcinoma (OPSCC). Materials & methods: Targeted multiplex bisulfite amplicon sequencing was performed on four OPSCC cell lines and primary non-neoplastic oral epithelial cells. Real-time quantitative polymerase chain reaction (RT-qPCR) was performed for a subset of associated genes. Results: There was clear differential methylation between one or more OPSCC cell lines and control cells for the majority of CpG-dense regions. Conclusion: Targeted multiplex bisulfite amplicon sequencing allowed us to efficiently map methylation across the entire region of interest with a high degree of sensitivity and helps shed light on novel differentially methylated regions that may have value as biomarkers of OPSCC.

Balancing yield, purity and practicality: a modified differential ultracentrifugation protocol for efficient isolation of small extracellular vesicles from human serum.

Ultracentrifugation remains the gold standard for isolation of small extracellular vesicles (sEV), particularly for cancer applications. The objective of this study was to determine if a widely used ultracentrifugation protocol for isolation of serum sEV could be modified to reduce the number of ultracentrifugation cycles and increase efficiency, while maintaining equal or better sample purity and yield. Serum was obtained from two healthy subjects. sEVs were isolated from 1 mL aliquots using three different ultracentrifugation protocols. Co-isolation of RNA carrier protein was assessed by performing Western blots for ApoA-I, ApoB, and Ago2. Small RNA-sequencing was performed on the sEV isolates, and differential detection of small ncRNA was compared across isolation protocols. Reduction from three- to two-ultracentrifuge cycles with no sucrose cushion resulted in a much higher sEV yield but also had the highest levels of lipoprotein and Ago2 contamination. However, the two-ultracentrifugation cycle protocol that incorporated a 30% sucrose cushion into the first cycle resulted in slightly higher sEV yields with lower levels of protein contamination compared to the lengthier three-ultracentrifugation cycle approach, therefore presenting a more efficient alternative approach for isolation of serum sEVs. It was also notable that there were some differences in sEV ncRNA cargo according to protocol, although it was less than expected given the differences in co-isolated RNA carrier proteins. Our results suggest that use of the modified serum sEV isolation protocol with two ultracentrifugation cycles and incorporating a 30% sucrose cushion offers a more efficient approach in terms of efficiency and purity.

Differential expression and prognostic value of long non-coding RNA in HPV-negative head and neck squamous cell carcinoma.

BACKGROUND: Long non-coding RNA (lncRNA) has emerged as a new avenue of interest due to its various biological functions in cancer. Abnormal expression of lncRNA has been reported in other malignancies but has been understudied in head and neck squamous cell carcinoma (HNSCC). METHODS: The lncRNA expression was interrogated via quantitative real-time polymerase chain reaction (qRT-PCR) array for 19 human papillomavirus (HPV)-negative HNSCC tumor-normal pairs. The Cancer Genome Atlas (TCGA) was used to validate these results. The association between differentially expressed lncRNA and survival outcomes was analyzed. RESULTS: Differential expression was validated for 5 lncRNA (SPRY4-IT1, HEIH, LUCAT1, LINC00152, and HAND2-AS1). There was also an inverse association between MEG3 expression (not significantly differentially expressed in TCGA tumors but highly variable expression) and 3-year recurrence-free survival (RFS). CONCLUSION: We identified and validated differential expression of 5 lncRNA in HPV-negative HNSCC. Low MEG3 expression was associated with favorable 3-year RFS, although the significance of this finding remains unclear.

Comprehensive microRNA-sequencing of exosomes derived from head and neck carcinoma cells in vitro reveals common secretion profiles and potential utility as salivary biomarkers.

Exosomes are nano-scale, membrane encapsulated vesicles that are released by cells into the extracellular space and function as intercellular signaling vectors through horizontal transfer of biologic molecules, including microRNA (miRNA). There is evidence that cancer-derived exosomes enable the tumor to manipulate its microenvironment, thus contributing to the capacity of the tumor for immune evasion, growth, invasion, and metastatic spread. The objective of this study was to characterize differential secretion of exosomal miRNA by head and neck squamous cell carcinoma (HNSCC) and identify a set of candidate biomarkers that could be detected in non-invasive saliva samples. We isolated exosomes from conditioned media from 4 HNSCC cell lines and oral epithelial control cells and applied miRNA-sequencing to comprehensively characterize their miRNA cargo and compare transcript levels of each HNSCC cell line to that of oral epithelial control cells. A candidate set of miRNA differentially secreted by all 4 HNSCC cell lines was further evaluated in saliva collected from HNSCC patients and healthy controls. We observed extensive differences in exosomal miRNA content between HNSCC cells when compared to normal oral epithelial control cells, with a high degree of overlap in exosomal miRNA profiles between the 4 distinct HNSCC cell lines. Importantly, several of the exosomal miRNA secreted solely by cancer cells in culture were detected at substantially elevated levels in saliva from HNSCC patients relative to saliva from healthy controls. These findings provide important insight into tumor biology and yields a promising set of candidate HNSCC biomarkers for use with non-invasive saliva samples.

Immune Response to HPV16 E6 and E7 Proteins and Patient Outcomes in Head and Neck Cancer.

IMPORTANCE: Pathology-based measures of human papillomavirus (HPV) status are routinely obtained in the care of head and neck cancer and are clearly associated with patient outcome for cancers of the oropharynx. However, it is unclear if HPV status is of high value for cancers of the larynx and oral cavity. In addition, it is possible to assess HPV infection using serology-based methods; however, the suitability of this pathology-independent measure for predicting patient outcome in head and neck cancer is unknown. OBJECTIVE: To investigate whether immunologic response to HPV16 is associated with patient survival across anatomic sites, independent of smoking and drinking history. DESIGN, SETTING, AND PARTICIPANTS: This was a population-based study of 1054 patients with head and neck cancer in the greater Boston, Massachusetts, area (1999-2003, 2006-2011). MAIN OUTCOMES AND MEASURES: All-cause survival in relation to HPV16 E6 and E7 seropositivity. RESULTS: The 1054 patients reflected the demographics of those treated in this timeframe (75% male; mean age, 59 years). Seropositivity was very strongly associated with improved survival overall (hazard ratio HR], 0.33; 95% CI, 0.24-0.45; P < .001), with no evidence that the magnitude of immune response, as assessed by titer levels, effected outcome. Seropositivity was associated with improved patient survival across all head and neck cancer sites: HR for oropharynx cancer, 0.26; 95% CI, 0.18-0.39; for oral cavity cancer, 0.45; 95% CI, 0.18-0.80; and for larynx cancer, 0.29; 95% CI, 0.10-0.85. In addition, the associations with seropositivity were similar across smoking and/or drinking exposure groups: HRfor low exposure, 0.52; 95% CI, 0.20-1.36; for moderate exposure, 0.42; 95% CI, 0.25-0.70; for heavy exposure, 0.51; 95% CI, 0.36-0.73. In a subset of 162 patients with both HPV serology and p16 immunohistochemical (IHC) measures available, both measures were similarly associated with survival in the oropharynx (HR for serology, 0.16; 95% CI, 0.03-0.47; for p16 measures, 0.16; 95% CI, 0.03-0.46), whereas only serology was associated with outcome when considering all head and neck cancer cases (HR for serology,0.49; 95% CI, 0.23-1.04; for p16, 0.65; 95% CI, 0.30-1.42). CONCLUSIONS AND RELEVANCE: Collectively, these data suggest that a positive serologic response to HPV16 oncoproteins may be the best approach to assess HPV-disease for clinical outcome because it is associated with survival for all types of disease and is a marker that is not dependent on pathology material.

CpG island methylation profile in non-invasive oral rinse samples is predictive of oral and pharyngeal carcinoma.

BACKGROUND: There are currently no screening tests in routine use for oral and pharyngeal cancer beyond visual inspection and palpation, which are provided on an opportunistic basis, indicating a need for development of novel methods for early detection, particularly in high-risk populations. We sought to address this need through comprehensive interrogation of CpG island methylation in oral rinse samples. METHODS: We used the Infinium HumanMethylation450 BeadArray to interrogate DNA methylation in oral rinse samples collected from 154 patients with incident oral or pharyngeal carcinoma prior to treatment and 72 cancer-free control subjects. Subjects were randomly allocated to either a training or a testing set. For each subject, average methylation was calculated for each CpG island represented on the array. We applied a semi-supervised recursively partitioned mixture model to the CpG island methylation data to identify a classifier for prediction of case status in the training set. We then applied the resultant classifier to the testing set for validation and to assess the predictive accuracy. RESULTS: We identified a methylation classifier comprised of 22 CpG islands, which predicted oral and pharyngeal carcinoma with a high degree of accuracy (AUC = 0.92, 95 % CI 0.86, 0.98). CONCLUSIONS: This novel methylation panel is a strong predictor of oral and pharyngeal carcinoma case status in oral rinse samples and may have utility in early detection and post-treatment follow-up.

Human papillomavirus serology and tobacco smoking in a community control group.

BACKGROUND: HPV infection is an established risk factor for oropharyngeal cancer, and it has been proposed that cigarette smoking may potentiate HPV infection in the oral epithelium. We sought to test the hypothesis that cigarette smoking increases HPV infection in an HPV16 serology study of cancer-free individuals. METHODS: Subjects were participants in a risk factor study for head and neck cancer, and were required to have no prior history of either HNSCC or any other cancer. Tobacco use and other risk factor data were gathered through interviewer-assisted questionnaires, while serology was conducted in a blinded fashion using a glutathione S-transferase capture enzyme-linked immunosorbent assay (ELISA) to detect antibodies against HPV16 L1, E1, E2, E4, E6 and E7 proteins. The differences in tobacco use by HPV serology were evaluated by ANOVA; and the reported odds ratios and 95% confidence intervals were determined by using unconditional logistic regression. RESULTS: We found no overall association of HPV16 serological markers with smoking. However, when the data were stratified by median age, smoking was positively associated with seropositivity for the HPV16 L1 capsid antigen in the younger controls while the older controls were less likely to be HPV16 L1 positive if they smoked (pinteraction < 0.002). There was no similar association of smoking and age with serological response to the early proteins (i.e E6, E7). CONCLUSIONS: Exposure to HPV16 capsid protein (L1) is increased among relatively younger adults who smoke and diminished among older smokers. However, this pattern is not accompanied by a differential susceptibility for active infection (as determined by the early gene proteins such as E6 and E7) among young and older smokers.

Obesity and head and neck cancer risk and survival by human papillomavirus serology.

PURPOSE: Previous studies examining the association of body mass index (BMI) with risk of and survival from head and neck squamous cell carcinoma (HNSCC) have been inconsistent, although an inverse association has been noted for obesity and risk of HNSCC in several studies. Previous studies have not examined whether these associations differ by human papillomavirus (HPV) status. METHODS: We utilized the resources of a population-based case-control study of HNSCC from the greater Boston area (959 cases and 1,208 controls were eligible for this analysis). Anthropometric history was collected through personal interviews, and HPV status was assessed using serology. We analyzed the association between BMI (assessed 5 years prior to disease incidence) and disease risk and survival using logistic regression and Cox proportional hazards regression, respectively. RESULTS: After adjusting for known risk factors, the association between obesity and overall risk of HNSCC was not significant (OR 0.79, 95 % CI 0.60-1.04). However, obesity (BMI ≥30 kg/m(2)) was inversely associated with HNSCC risk among HPV-seronegative cases (OR 0.48, 95 % CI 0.32-0.70), but not among HPV-seropositive cases (OR 0.91, 95 % CI 0.68-1.21). BMI was not associated with survival overall or by HPV status. However, being overweight (BMI 25-29.9 kg/m(2)) was associated with longer survival among HPV-seropositive smokers (HR 0.48, 95 % CI 0.31-0.74). CONCLUSIONS: Our findings are consistent with previous observations that obesity is inversely associated with the risk of HNSCC; however, this association appears to be confined to HPV-seronegative cases. Overall, obesity was not associated with HNSCC survival overall or by HPV status. IMPACT: Obesity is associated with risk of non-HPV HNSCC, but not HPV HNSCC.

Epigenetics of lung cancer.

Lung cancer is the leading cause of cancer-related mortality in the United States. Epigenetic alterations, including DNA methylation, histone modifications, and noncoding RNA expression, have been reported widely in the literature to play a major role in the genesis of lung cancer. The goal of this review is to summarize the common epigenetic changes associated with lung cancer to give some clarity to its etiology, and to provide an overview of the potential translational applications of these changes, including applications for early detection, diagnosis, prognostication, and therapeutics.

Novel DNA methylation targets in oral rinse samples predict survival of patients with oral squamous cell carcinoma.

OBJECTIVES: The objective of this study was to identify novel survival-associated biomarkers in oral rinse samples collected from patients with oral squamous cell carcinoma (OSCC). MATERIALS AND METHODS: We screened for putative survival-associated markers using publicly available methylation array data from 88 OSCC tumors. Cox models were then fit to methylation array data restricted to these putative loci in oral rinse samples of 82 OSCC patients from greater Boston. Pyrosequencing assays were designed for each locus that replicated in the oral rinse samples and applied to a validation set of oral rinse samples from another 61 OSCC patients. RESULTS: We identified 7 survival-associated methylation markers in oral rinse samples from OSCC patients, and have validated one, located in the body of GABBR1, by pyrosequencing. CONCLUSION: The 7 CpG loci identified through this study represent novel prognostic biomarkers for patients with OSCC that can be detected using a non-invasive oral rinse collection technique.

Let-7 microRNA-binding-site polymorphism in the 3'UTR of KRAS and colorectal cancer outcome: a systematic review and meta-analysis.

There is a small but growing body of literature regarding the predictive utility of a Let-7 microRNA-binding-site polymorphism in the 3'-untranslated region (UTR) of KRAS (KRAS-LCS6) for colorectal cancer outcome, although the results are conflicting. We performed a review and meta-analysis in an attempt to better clarify this relationship. A PubMed search was conducted to identify all studies reporting on KRAS let-7 microRNA-binding site polymorphism (LCS6; rs61764370) and colorectal cancer outcome. Hazard ratios (HR) and corresponding 95% confidence intervals (CI) were extracted or estimated from each manuscript. Log HRs and log CIs were combined across studies using the inverse-variance weight to calculate fixed- and random-effects summary estimates and corresponding 95% CIs for overall and progression-free survival. We did not observe any significant association between overall or progression-free survival, neither when considering all colorectal cancer patients nor for subgroup analyses (metastatic, anti-EGFR [epidermal growth factor receptor] treatment, or KRAS wild type). There was substantial heterogeneity across studies, overall and among subgroups analyzed. We have found no clear evidence to support an association between the KRAS-LCS6 genotype and overall or progression-free survival among colorectal cancer patients, even after conducting subgroup analyses by stage and anti-EGFR treatment status. This information helps to clarify the confusing body of literature regarding the clinical implications of the KRAS-LCS6 genetic variant on colorectal cancer outcomes, indicating that it should not be used at the present time to personalize therapeutic strategies (PROSPERO registration number: CRD42013005325).